Advisory Editorial Board

Bharat Patel

Griffith University, Australia

Chidchanok Lursinsap

Chulalongkorn University, Thailand

Chiu-Yue Lin

Feng Chia University, Taiwan

David J. Lurie

University of Aberdeen, UK

David Ruffolo

Mahidol University, Thailand

Hyoung Tae Choi

Kangwon National University, Korea

Pranom Chantaranothai

Khon Kaen University, Thailand

Somsak Pantuwatana

Burapha University, Thailand

Sukit Limpijumnong

Suranaree University of Technology, Thailand

Supot Hannongbua

Chulalongkorn University, Thailand

Suthat Yoksan

Srinakharinwirot University, Thailand

Vichai Reutrakul

Mahidol University, Thailand

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

Working Group Consultant Dean, Faculty of Science, Ubon Ratchathani University

Editor in Chief Chittakorn Polyon

Editors Nongkhran Sasom

Pornpan Pongpo

Saisamorn Lumlong

Sungwan Kanso

Udom Tipparach

Language Editors Bob Tremayne

Ian Thomas

Managing Staff Amornrat Wasuree

Apinya Pitaksa

Dutruthai Sahapong

Jiraporn Thongsud

Nanthana Pimpan

Sudarat Khamfai

Tutiyaporn Weerakul

Wanwisa Songserm

Art Work Kittipong Phitukwongyothin

Surasit Sutthikhampa

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

I

Editorial This is the 2nd volume (Number 1, January-June, 2011) of Science Journal Ubon Ratchathani University (SCJ) published by Faculty of Science, Ubon Ratchathani University. In this volume, there are 8 research articles and a review article with totally 71 pages. The most content of the volume consists of research articles and the review article, involving with researches in the fields of chemistry, materials science, physics, statistics, biology, and biotechnology. We would like to take this opportunity to express our deep appreciation to all authors and reviewers who for their significant contributions to make SCJ happen. We also hope that authors who submitted their manuscripts to SCJ will get a taste of achievement in publishing in an international journal. In the future, we aim to strive for the best quality in all published articles.

Editorial Team Science Journal Ubon Ratchathani University

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

II

Guide for Authors The Science Journal Ubon Ratchathani University (SCJ) is a peer reviewed journal publishing high quality articles. The SCJ accepts all articles dedicated to all aspects of sciences such as physics, chemistry, materials science, biology, biochemistry, biotechnology, microbiology, environmental science, and other basic sciences such as education science, mathematics, statistics and computer science, including information technology. The articles required are either research articles or review articles, is defined as follows: 1.

Review articles: an article which aims to present comprehensively already existing finding.

2.

Research articles: a regular article which aims to present new findings.

The SCJ considers only manuscripts that have not been published (or submitted simultaneously) at any language, elsewhere. The SCJ is issued both in electronic form (for free) and printed form as two volumes per year (free for the authors). The manuscript templates for both two article types should be prepared in english form, which can be downloaded from the SCJ homepage (http://scjubu.sci.ubu.ac.th). In the manuscript preparation, the article pages, including tables, figures and references, should be not over 12 pages. The sizes of figures in each article should be surely given high resolution enough to be clearly reviewed. The figures should be also used only in black-and-white. However, these figures could be prepared to be shown in colors in the online edition. Each article in the SCJ is reviewed by two or three reviewers. The review process is about 2-3 weeks. The revision article should be revised in 1-3 weeks, depending on each recommendation from a reviewer. The average reviewed, revised and managed time for each article published in the SCJ is about 2-3 months. In order to submit articles, authors should firstly register to obtain a username, and then login at the SCJ homepage (http://scjubu.sci.ubu.ac.th) to access the submission process. All information about the submitted articles would be informed by the SCJ editors ([email protected]) via e-mail of corresponding authors.

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

Contents Editorial

I

Guide for Authors

II

Auxins and Cytokinins Regulate Abscission and Physiological Changes of Flowers in Cut Dendrobium cv. Eiskul Inflorescences K. Rungruchkanont

1-11

Encapsulation of Acorus Calamus Linn. Extract by Polyurethane Microcapsules A. Paphonngam et al.

12-16

Forecasting Malaria Incidence Based on Monthly Case Reports and Climatic Factors in Ubon Ratchathani Province, Thailand, 2000 – 2009 W. Sriwattanapongse et al.

17-24

Gas Exchange Properties of Fresh Chillies and Their Applications in Mathematical Modelling of Packaging Systems W. Utto et al.

25-35

Synthesis and Characterization of Carbon Nanotubes Using a Natural Precursor: Turpentine Oil C.R. Bhattacharjee et al.

36-42

Neutron Interaction Cross-Sections for 27Al in the Energy Range from 0.2 to 22 MeV Using Optical Model Program A.K.M.R. Rahman et al.

43-52

Screening for Bioactive Compound Group in Parmotrema tinctorum’s Extract and Bioactivity Test of Dichloromethane Extract as Anti-tuberculosis Against Mycobacterium tuberculosis H37RV and Toxicity Against Artemia salina. I.K. Kusumaningrum et al.

53-59

Doxorubicin and Thousand Efforts to Get the Better Drugs N. Pongprom et al.

60-65

Natural Rubber bound 4-Aminodiphenylamine Antioxidant for Rubber Formulation Improvement P. Klinpituksa et al.

66-71

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

SCIENCE JOURNAL Ubonratchathani University

Sci. J. UBU, Vol. 2, No. 1 (January – June, 2011) 1-11

http://scjubu.sci.ubu.ac.th

Research Article

Auxins and Cytokinins Regulate Abscission and Physiological Changes of Flowers in Cut Dendrobium cv. Eiskul Inflorescences K. Rungruchkanont1,2* 1

Department of Horticulture, Faculty of Agriculture, Ubon Ratchathani University, Ubon Ratchathani 34190, Thailand. 2 Postharvest Technology Innovation Centre, Ubon Ratchathani University, Ubon Ratchathani 34190, Thailand. Received 13/12/10; Accepted 01/02/11

Abstract The effects of auxins and cytokinins on quality and physiological changes in cut Dendrobium cv. Eiskul inflorescences were studied at Ubon Ratchathani University, Thailand. The auxins tested were indoleacetic acid (IAA), indolebutylic acid (IBA), and naphthaleneacetic acid (NAA). The cytokinins tested were 6-benzylaminopurine (BAP; also called benzyladenine) and 6-furfurylaminopurine (kinetin). The hormones were supplemented in a vase solution, at 5, 25 and 50 mg L-1. All auxins tested extended the vase life of Dendrobium inflorescences by delaying senescence and inhibiting abscission of open flowers. The high concentrations of auxins were effective and increased ovary size. NAA, applied at 50 mg L-1 completely prevented abscission. This treatment could induce greening of the normally whitish pedicel, indicating that it reverses a number of developmental processes. Cytokinins had no effect on vase life and ovary size but BAP at 50 mg L-1 inhibited abscission of open flowers. The application of hormone treatments, IBA or BAP at 50 mg L-1 or a combination of 50 mg L-1 IBA and 50 mg L-1 BAP, prior to ethylene fumigation, prevented open flower abscission and decreased ACS activity in flowers. These results suggest that adding auxin hormones in the vase solution has the potential to improve cut flower quality and could be used as a commercial vase solution. Keywords: Abscission, Auxins, Cytokinins, Dendrobium, Vase solution.

placed in water, have a much shorter life than inflorescences that remain uncut. The older Dendrobium inflorescences, when cut and open flowers on the lower part of the cut inflorescences start to senesce earlier and show earlier abscission than the younger *Corresponding author. flowers higher up the inflorescence stalk. E-mail address: [email protected] 1. Introduction

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All rights reserved.

2

Auxins and Cytokinins Regulate Abscission and Physiological Changes of Flowers

The floral buds at the top of the inflorescence stalk exhibit early yellowing and also abscise. Inclusion of sugar and a suitable antibacterial compound in the vase water were found to delay early flower senescence, floral bud yellowing and floral bud abscission [1]. A good treatment consisted of 2% sucrose plus 200 mg L-1 8-hydroxyquinoline sulfate. In order to increase the efficiency of this vase solution, we further tested whether the inclusion of hormones in the vase water would prevent flower abscission and increase the vase life of inflorescences. Auxins are generally effective in inhibiting abscission, and it appears that a major endogenous hormone regulates abscission [2-5]. It has been concluded that the main effect of auxins is to decrease the sensitivity of the cells in the abscission zone to ethylene [4,6,7] Cytokinins are also known to decrease sensitivity of tissue to ethylene [8]. Cytokinins are endogenous regulators of sensecence [9]. For example, transgenic plants expressing the KNOTTED 1 (KN1) gene under the control of a senescence-specific promoter, showed delayed leaf senescence. This was accompanied by an increase of the cytokinin content in the leaves [10]. Similar experiments with the ipt gene in Petunia showed a delay of petal senescence. Isopentenyl transferase catalyzes a limiting step in cytokinin biosynthesis [8]. Much less has been known about the possible role of cytokinins on abscission. A combined treatment of gibberellins and a cytokinin has been reported to delay abscission of flowers [11] and petals [12], but the mechanism of cytokinin remains unclear. Thus far, the only report showing that cytokinins can delay flower abscission is the study by Sankhla et al. [13], who found that inclusion of thidiazuron, a cytokinin that is effective at very low concentrations, in the vase solution delayed flower abscission in cut Phlox paniculata inflorescences.

the quality and physiological changes of Dendrobium cv. Eiskul inflorescences. We also tested the effect of hormones under ethylene conditions. 2. Materials and Methods Plant Material. Cut inflorescences of Dendrobium cv. Eiskul were obtained from a commercial grower in Sakon Nakhon, Thailand. Inflorescences were transported 300 km at 25 °C to the laboratory at Ubon Ratchathani University, where they arrived within 5 h of harvest. Export grade inflorescences, with 6-7 open flowers and 4-5 flower buds, were selected for freshness and uniformity. Peduncles of individual inflorescences were recut in air, leaving 15 cm from the lowest open flower. Inflorescences were individually placed in 22 ml glass tubes containing 15 ml vase solution. The inflorescences were placed in a controlled room at 25 ±2°C and 75-80% relative humidity. Overhead daylight fluorescent lamps (TL-D, 36W/54, Philips) provided a photon flux density of about 10 μmoles m-2 s-1 at the level of the inflorescences, for 12 consecutive h per day.

Auxin and Cytokinin Treatments. The vase solutions all contained 2% sucrose plus 200 mg L-1 8-hydroxyquinoline sulfate (8-HQS). The auxins tested were indoleacetic acid (IAA; Fluka), indolebutyric acid (IBA; Fluka), and naphthaleneacetic acid (NAA; Merk). These chemicals were first dissolved in 0.1 N NaOH. The cytokinins tested were 6-benzylaminopurine (BAP; Fluka) and 6furfurylaminopurine (kinetin; Fluka) and were also first dissolved in 0.1 N NaOH. Final concentrations of auxins and cytokinins were 5, 25 or 50 mg L-1. Each treatment solution was adjusted to pH 4.0 in order to avoid differences in pH caused by NaOH. Control vase solutions were distilled water and 2% sucrose plus 200 mg L-1 8-hydroxyquinoline sulfate (8-HQS). Data were recorded daily for 45 d on petal senescence, bud yellowing, floral bud opening, floral bud In this study, we reported on the effects abscission, open flower abscission and vase of three auxins and two cytokinins on

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All rights reserved.

K. Rungruchkanont, Sci. J. UBU, Vol. 2, No. 1 (January – June, 2011) 1-11

life. Vase life of inflorescence was determined when senescence of open flower and flower bud occurred to 50%. Ovary diameter was measured at the upper end of pedicels by vernier caliper on day 40. Application of Hormones and Ethylene Treatments. IBA, BAP at 50 mg L-1 and a combination of IBA at 50 mg L-1 and BAP at 50 mg L-1 were added in vase solutions and used as hormone treatments. Inflorescences of Den. cv. Eiskul were put in three hormone treatments and two controls (same as above) for 24 h and then were treated with 0.25 μL L-1 ethylene for 24 h at 25 ºC. Open flower abscission was recorded daily for 15 d. ACC Synthase Activity Assay. Open flowers were collected from inflorescences that had been treated with IBA, BAP and a combination of IBA and BAP, two controls and then all were treated with ethylene. Treated tissues were collected at day 0, 1, 3, 5 and 7. For extraction and analysis of ACC synthase (ACS) activity, tissue (3g) was ground in 100 mM EPPS buffer (pH 8.5) containing 4 mM dithiothreol (DTT) and 0.5 mM pyridoxal phosphate (1:2 w/v), using mortar and pestle. The homogenate was centrifuged at 12,000 g for 20 min. The supernatant was placed in a dialysis bag and then soaked in a dialysis buffer containing 2 mM EPPS buffer, 0.1 mM DTT and 0.2 μM pyridoxal phosphate (1:10, v/v). The extract was dialyzed for 24 h. All steps were carried out at 4 oC. ACS activity was assayed according to method of Hoffman and Yang [14] by incubation of 0.4 ml extraction in 6ml tube with 50 μl of 0.5 mM Sadenosylmethionine and 90 μl of distilled water at 30 °C for 3 h. The ACC produced was extracted and analyzed according to Lizada and Yang [15] and modified by Hoffman and Yang [14]. Weighed flower tissues were ground in 9% trichloroacetic acid (TCA) (1:4, w/v) using a mortar and pestle. After holding for 12 h at 4 °C, the extract was centrifuged at 12000g for 20 min to remove insoluble cellular debris. The supernatant was used for ACC analysis. ACC content in floral tissue was measured

3

according to the method described by Lizada and Yang [15]. 100 μl of 10mM HgCl2 and 600 μl distilled water were added to 200 μl of the extract in a 6-ml tube sealed with a serum cap and held at 0 °C. Approximately 100 μl of a cold mixture of 5.25% NaOCl and saturated NaOH (2:1 v/v) were injected into the vial through the serum cap. The mixture was vortexed and incubated at 0 °C. The ethylene liberated was measured after 3 min incubation and the conversion factor determined by comparison with the production of ethylene from an internal ACC standard. The analysis was repeated three times for each treatment. Statistical Analysis. The experiments of auxins and cytokinins treatment were conducted at 6 replicate inflorescences. The experiment of hormones and ethylene treatment was conducted at 10 replicate inflorescences. The results were compared by analysis of variance, calculating Duncan's Multiple Range Test (DMRT). All data on abscission, the means between treatments were compared after calculating least significant difference (LSD). Experiments were repeated at least once at a later date, with similar results. The data presented refer to only one of these repeat experiments. 3. Results and Discussion Effects of Auxins and Cytokinins on Floral Senescence and Abscission. Inflorescences of Dendobium cv. Eiskul had a long vase life. The vase life in water control was 34.8 d, in HQS control it was 35.0 d whilst vase life in most auxin treatments, 45-54 d, was significantly higher than controls (Table 1). However, no difference in vase life was found among cytokinin treatments (Table 2). The average time to visible petal senescence of the flowers that were already open at the beginning of the experiments was 21.8 ± 2.6 d in the HQS control. Petal senescence in these flowers was delayed by 50 mg L-1 IAA (28.2 ± 4.8 d), 50 mg L-1 IBA (35.3 ± 5.05 d), 50 mg L-1 NAA (36.3 ± 7.9 d) and 50 mg L-1 BAP (32.7 ± 2.9 d) (data not shown).

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All rights reserved.

4

Auxins and Cytokinins Regulate Abscission and Physiological Changes of Flowers

Table 1. Effects of auxins and concentrations on physiological changes of Dendrobium. cv. Eiskul. Treatments

Vase Life (days)

Water 8-HQS 5 mg L-1 IAA 25 mg L-1 IAA 50 mg L-1 IAA 5 mg L-1 NAA 25 mg L-1 NAA 50 mg L-1 NAA 5 mg L-1 IBA 25 mg L-1 IBA 50 mg L-1 IBA F-test

34.8 d 35.0 d 37.8 d 48.2 ab 49.4 ab 45.0 bc 45.5 bc 48.2 ab 40.5 cd 45.0 bc 54.4 a *

Ovary Diameter (mm.) 3.56 c 3.58 de 3.58 de 3.85 bc 4.01 ab 3.72 cde 4.13 a 4.03 ab 3.77 cd 3.77 cd 3.97 ab *

% Bud Opening

% Bud Abscission

93.3 87.5 90.3 87.8 86.7 88.9 87.5 87.5 77.8 74.4 76.7 n.s.

37.5 24.2 25.0 14.4 15.6 19.5 23.6 12.5 18.1 18.1 23.3 n.s.

Means within a column not sharing the same letters were significantly different at P = 0.05 by DMRT; n.s. = not significant, * = significant at P = 0.05 and n= 6

Table 2. Effects of cytokinins and concentrations on physiological changes of Dendrobium. cv. Eiskul. Treatments

Vase life (days)

Water 8-HQS 5 mg L-1 BAP 25 mg L-1 BAP 50 mg L-1 BAP 5 mg L-1 Kinetin 25 mg L-1 Kinetin 50 mg L-1 Kinetin F-test

33.7 b 40.0 ab 40.3 ab 42.0 a 41.8 ab 40.5 ab 41.3 ab 39.0 ab *

Ovary diameter (mm.) 3.65 ab 3.60 ab 3.75 a 3.77 a 3.72 a 3.70 ab 3.66 ab 3.63 ab *

% Bud opening

% Bud abscission

75.0 83.3 100 95.8 94.4 75.6 86.1 89.2 n.s.

38.9 45.0 13.9 15.3 29.2 26.7 25.0 17.8 n.s.

Means within a column not sharing the same letters were significantly different at P = 0.05 by DMRT; n.s. = not significant, * = significant at P = 0.05 and n= 6

Abscission of open flowers was first found on day 20, and increased until day 45 (Figures 1 and 2). All auxins tested inhibited abscission of the open flowers. In each auxin tested, a higher concentration tended to have a larger inhibiting effect. Compared with the two other auxins tested, inhibition was

highest in the NAA treatments. At 50 mg L-1 NAA, abscission was completely absent until the end of the experiment (Figure 1). Kinetin had no effect on the abscission of open flowers (Figure 2). BAP, at 50 mg L-1 only tended to inhibited abscission but not significantly (Figure 2).

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All rights reserved.

K. Rungruchkanont, Sci. J. UBU, Vol. 2, No. 1 (January – June, 2011) 1-11

5

LSD0.05 = 15.34

100

Abscission (%)

80 60 40 20 0 -20 -5

0

5

10

15

20

25

30

35

40

45

Time (d)

Figure 1. Abscission of open flowers in Dendrobium cv. Eiskul inflorescences treated with auxin hormone. Distilled water control ( ■ ), 8-HQS control ( □ ), 5 mg L-1 IAA ( ▲ ), 25 mg L-1 IAA ( △ ), 50 mg L-1 IAA ( ▲ ), 5 mg L-1 NAA ( ◆ ), 25 mg L-1 NAA ( ◇ ), 50 mg L-1 NAA ( ◆ ), 5 mg L-1 IBA ( ● ), 25 mg L-1 IBA ( ○ ), 50 mg L-1 IBA ( ● ). Y axis indicates abscised flowers as a percentage of all open flowers. Means are the average of 6 inflorescences; the bar indicates LSD.

100

LSD0.05 = 23.68

Abscission (%)

80 60 40 20 0 -20 -5

0

5

10

15

20

25

30

35

40

45

Time (d)

Figure 2. Abscission of open flowers in Dendrobium cv. Eiskul inflorescences treated with cytokinin hormones. Distilled water control ( ■ ), 8-HQS control ( □ ), 5 mg L-1 BAP ( ▲ ), 25 mg L-1 BAP ( △ ), 50 mg L-1 BAP ( ▲ ), 5 mg L-1 Kinetin ( ◆ ), 25 mg L-1 Kinetin ( ◇ ), 50 mg L-1 Kinetin ( ◆ ). Y axis indicates abscised flowers as a percentage of all open flowers. Means are the average of 6 inflorescences; the bar indicates LSD.

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All rights reserved.

6

Auxins and Cytokinins Regulate Abscission and Physiological Changes of Flowers

Figure 3. The color of the ovary and pedicel in Dendrobium cv. Eiskul open flowers attached to cut inflorescences, after holding the inflorescence stems in vase solution containing various auxins at 50 mg L-1 for 40 days. The pedicels have been broken off at the abscission zone. Flower buds showed early yellowing. The average time of bud yellowing was not depending on treatment and was about 4.7 d. There was no difference in bud abscission in all treatments (Tables 1 and 2). The treatments also did not affect bud opening, with 74-100% bud opening at the end of experiment (Tables 1 and 2). The average time to visible petal senescence of opening buds in the water controls was 35.5 ± 1.9 d, in the HQS controls it was 40.7 ± 5.5 d while in the treatments with BAP it was 44.2 ± 4.2 d. All auxins at 50 mg L-1 further delayed petal senescence of the opened buds to about 48.4±3.8 d (data not shown). Effects of Auxins and Cytokinins on Ovary Growth and the Color of the Ovary and Pedicel. High concentrations of auxins treatment effected ovary diameter. On day 40 of vase life, flowers on inflorescences supplemented with IAA and NAA at 25 and 50 mg L-1 and IBA at 50 mg L-1 had larger ovary diameter than the controls (Table 1). The 50 mg L-1 NAA treatment also resulted in a marked greening of the ovary and pedicel (Figure 3). Much less greening of the ovary

was found in the 50 mg L-1 IBA treatment (Figure 3). Ovary size and pedicel color were not affected by cytokinins (Table 2). Effects of Hormones and Ethylene Treatments. Applications of IBA, BAP and IBA+BAP prior to ethylene treatment decreased flower abscission. Flowers treated with IBA and IBA+BAP had only 11 and 15 % abscission, respectively, which were significantly lower than the controls (Figure 4). Activities of ACS are shown in Figure 5. The ACS activity was low in flowers before hormones and ethylene were applied (before). The ACS activity increased rapidly in all treatments after ethylene was applied (day 0). However, flower treated with IBA and IBA+BAP had lower ACS activity than the other treatments. From day 1 to day 7, activity of ACS in flowers treated with all hormones slowly decreased whilst activity of ACS in control flowers increased. The activity of ACS in flowers of all treatments were related to flower abscission (Figures 4 and 5).

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All rights reserved.

K. Rungruchkanont, Sci. J. UBU, Vol. 2, No. 1 (January – June, 2011) 1-11

100

7

LSD0.05=20.8

Abscission (%)

80 60 40 20 0 -20 -1

0

1

3

5

7

9

11

13

15

Time (d) Figure 4.

Abscission of open flowers in Dendrobium cv. Eiskul inflorescences that were

treated with hormones prior to ethylene fumigation. Distilled water control ( ■ ), 8-HQS

control (◆ ), 50 mg L-1 IBA ( x ), 50 mg L-1 BAP ( ● ), 50 mg L-1 IBA + 50 mg L-1 BAP

Activities of ACS (nmol ACC/h/mg protein) .

( ▲ ). Y axis indicates abscised flowers as a percentage of all open flowers. Means are the average of 10 inflorescences; the bar indicates LSD.

12 10 8 6 4 2 0 -1

Before

day 0

day 1

day 3

day 5

day 7

Time Figure 5. ACC synthase activity in open flowers of Dendrobium cv. Eiskul that were treated with hormones prior to ethylene fumigation. Distilled water control ( ■ ), 8-HQS control

( ◆ ), 50 mg L-1 IBA ( x ), 50 mg L-1 BAP ( ● ), 50 mg L-1 IBA + 50 mg L-1 BAP ( ▲ ). Means are the average of 3 replications.

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All rights reserved.

8

Auxins and Cytokinins Regulate Abscission and Physiological Changes of Flowers

Three kinds of auxin, IAA, NAA and IBA, in vase solutions, could increase the vase life of Dendrobium cv. Eiskul (Table 1). The hormones could delay petal senescence and could decrease abscission of open flowers. The effects of auxin hormones were clearly detected at high concentration levels. NAA at 50 mg L-1 was able to completely prevent the abscission of open flowers (Figure 1) and it also had the effect on increasing ovary growth and promoting greening of pedicels (Table 1 and Figure 3). Synthetic auxins such as NAA and IBA are usually used in commercial agricultural practices, because they are more stable than IAA. IAA, the natural auxin, becomes degraded rapidly by light. In present tests, IAA was indeed less effective than IBA or NAA, but its effects were still considerable.

pollination induces developmental changes in the flower, such as corolla wilting, pigmentation changes, ovary and ovule development [4,21-23].

Auxins are known as hormones that can delay leaf yellowing in some species, for example in tulips [16]. This suggests that auxins can prevent net chlorophyll degradation. In present experiments, NAA at 50 mg L-1 was most effective treatment against flower drop, resulting in remarkable greening of both ovary and pedicel (Figure 3). Auxins therefore, not only are apparently able to prevent net chlorophyll degradation but they also seem to be able to promote net chlorophyll synthesis. Ovaries and pedicels are green when the floral buds are still young. They lose the green color during development into large buds and open flowers. This process was apparently reversed by the NAA treatment. Application of auxins such as NAA to the stigma of several orchid species induces ovary growth [17-19]. Auxin is the initiator of ovary growth and it acts through ethylene [20]. However, in this experiment we applied auxins into the vase solutions but we found the same result as application of auxins to the stigma.

Cytokinins were either ineffective (kinetin) or had a small effect compared to NAA (BAP). Thidiazuron, a cytokinin, however, has been found to reduce flower abscission in Phlox paniculata inflorescences [13]. We desired to corroborate this result, using a much less expensive cytokinin. Both in the work of Sankhla et al. [13] and in the present experiments the cytokinins tested did not completely prevent flower abscission.

Interestingly, the corollas of flowers that were applied with auxins in the vase solutions did not wilt as they happen in the pollination process (Figure 3). Normally,

Our results showed that the application of auxins in vase solutions retarded flower abscission of Dendrobium. Other reports have also shown that exogenously applied auxins can prevent or delay flower abscission. For example, a pulse treatment of 2,4dichlorophenoxyacetic acid (2,4-D) delayed floret abscission in Red Cestrum [24], the application of IAA to pedicels of Dendrobium “Miss Teen” after flower removal inhibited abscission of pedicels after ethylene treatment [7].

In order to confirm the effect of auxins and cytokinins on ethylene sensitivity of Dendrobium cv. Eiskul inflorescence, 50 mg L-1 IBA, 50 mg L-1 BAP and 50 mg L-1 IBA+ 50 mg L-1 BAP were applied to inflorescences prior to ethylene treatment. Inflorescences treated with three hormone treatments prior to ethylene treatment significantly decreased flower abscission compared to the controls. The 11% and 15% of flower abscission were found on IBA treatment and IBA + BAP treatment, respectively. They were lower than 31% of flower abscission found on BAP treatment (Figure 4). This result indicated that IBA had higher efficiency to control abscission than BAP and there was no interaction effect between IBA and BAP.

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All rights reserved.

K. Rungruchkanont, Sci. J. UBU, Vol. 2, No. 1 (January – June, 2011) 1-11

Ethylene is known as a gaseous plant hormone involved in senescence and abscission of plant organs [25,26]. In this experiment, flowers in control treatment that were treated with water and with 8-HQS prior to ethylene treatment had high flower abscission (68% and 44%, respectively) during 15 days (Figure 4). Flowers in control treatment without ethylene treatment had the same rate of abscission on day 35 (Figure 1). These results confirmed the role of ethylene on flower abscission. Nevertheless, the application of auxin and cytokinin under ethylene condition reduced the effect of ethylene. Auxins were reported to prevent flower abscission by decreasing the sensitivity of the cells in the abscission zone to ethylene [6,7]. Cytokinins are also known to decreased sensitivity of tissue to ethylene [8]. The application of exogenous ethylene can induce endogenous ethylene in plant tissue by autocatalytic process and senescence symptoms are developed. In ethylene biosynthesis, ACC synthase (ACS) is a key enzyme that catalyzes S-adenosylmethionine (SAM) to 1-aminocyclopropane-1-carboxylic acid (ACC). So the low ACS activity in plant tissues that were treated with ethylene implies less sensitivity of plant tissue to ethylene. In our results, the data of ACS activity in flowers showed that these hormones decreased sensitivity of cell to ethylene (Figure 5). ACS activity in flowers without ethylene application (before) was very low. The application of ethylene increased ACS activity in all treatments (day 0). However, IBA treated and IBA+BAP treated flowers showed lower ACS activity than flowers in the other treatments. Interestingly, BAP treated flowers at day 0 had high level of ACS activity as controls, but after 7 day the ACS activity in BAP treated flowers decreased to the same level as IBA treated and IBA+BAP treated flowers and were lower than the controls. The result of ACS

9

activity in hormone treated flowers was similar to a report using aminooxyacetic acid (AOA), an inhibitor of ethylene synthesis, to lower ACS activity in flowers that had been pollinated [18]. The data on auxins in the vase solutions are reminiscent of those in which auxin was applied in other ways. For an example, auxin applied to the pedicel of Dendrobium “Miss Teen”, after flower removal, inhibited early pedicel abscission [7]. In present experiment auxins were added in the vase solution, which is a simple and commercially viable way of application. The results indicate that auxins can be included in commercial practices to improve cut flower quality by including chemicals in the vase solution. 5. Conclusions A practically feasible treatment has been found to considerably extend vase life of Dendrobium inflorescences. All high concentrations of auxins tested delayed senescence and inhibited abscission of open flowers and also increased ovary size of flower. NAA, applied at 50 mg L-1 even completely prevented abscission. This treatment resulted in greening of the normally whitish pedicel. Cytokinins had no effect on vase life and ovary size but BAP at 50 mg L-1 could inhibit abscission. The applications of hormone treatments prior to ethylene fumigation prevented open flower abscission and decreased ACS activity. Acknowledgements This research was financially supported by Postharvest Technology Innovation Center and Siam Taiyoo Farm Co., Ltd. Thanks are due to Prof. Dr. Wouter van Doorn for his suggestions and Prof. Dr. Michael Hare for proof-reading the English.

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10

Auxins and Cytokinins Regulate Abscission and Physiological Changes of Flowers

References [1] Ketsa, S. (1986). Effect of sucrose, silver nitrate and 8-hydroxyquinoline sulfate on postharvest behavior of Dendrobium Pompadour flowers. In Proceeding of the Sixth Asean Orchid Congress Seminar, Bangkok, Thailand, pp. 124-9. [2] Jacobs, W.P. (1962). Longevity of plant organ: internal factors controlling abscission. Annu. Rev. Plant Physiol., 13, 403-36. [3] Carns, R.H. (1966). Abscission and its control. Anuu. Rev. Plant Physiol., 17, 295-314. [4] van Doorn, W.G., & Stead, A.D. (1997). Abscission of flowers and floral parts. J. Exp. Bot., 48, 821-37. [5] Al-Khalifah, N.S., & Alderson, P.G. (1999). The effect of auxins and ethylene on leaf abscission of Ficus benjamina, In A.K. Kenellis, C. Chang, H. Klee, A.B. Bleecker, J.C. Pech & D. Grierson (Eds.). Biology and Biotechnology of the Plant Hormone Ethylene II. (pp. 255-60). Dordrecht: Kluwer Academic Publishers. [6] Meir, S., Hunter, D.A., Chen, J., Halaly, V., & Reid, M.S. (2006). Molecular changes occurring during acquisition of abscission competence following auxin depletion in Mirabilis jalapa. Plant Physiol.,141, 160416. [7] Rungruchkanont, K., Ketsa, S., Chatchawankanphanich, O., & van Doorn, W.G. (2007). Endogenous auxin regulates the sensitivity of Dendrobium (cv. Miss Teen) flower pedicel abscission to ethylene. Funct. Plant Biol., 34, 885-94. [8] Chang, H., Jones, M.L., Banowetz, G.M., & Clark, D.G. (2003). Overproduction of cytokinins in petunia flowers transformed with PSAG12-IPT delays corolla senescence and decreases sensitivity to ethylene. Plant Physiol., 132, 2174-83. [9] Noh, Y., Quirino, B.F., & Amasino, R.M. (2004). Senescence and genetic engineering. In L.D. Nooden (Ed.). Plant Cell Death Processes (pp. 91-105). USA: Academic Press. [10] Ori, N., Juarez, M.T., Jackson, D., Yamaguchi, J., Banowetz, G.M., & Hake, S. (1999). Leaves senescence is delayed in tobacco plants expressing the maize homeobox gene knotted 1 under the control of a senescence-activated promoter. Plant Cell, 11, 1073-80. [11] Wien, H.C., & Zhang, Y. (1991). Prevention of flower abscission in bell pepper. J. Amer.

Soc. Hort. Sci., 116, 516-9. [12] Kim, H.J., & Miller, W.B. (2009). GA4+7 and BA enhances postproduction quality in pot tulip. Postharvest Bio.Techno., 51, 272-7. [13] Sankhla, N., Mackay, W.A., & Davis, T.D. (2003). Reduction of flower abscission and leaf senescence in cut phlox inflorescences by thidiazuron. Acta Hort., 628, 837-41. [14] Hoffman, N.E., & Yang, S.F. (1982). Enhancement of wound-induced ethylene synthesis by ethylene in preclimacteric cantaloupe. Plant Physiol., 69, 317-22. [15] Lizada, M.C.C., & Yang, S.F. (1979). A simple and sensitive assay for 1-aminocyclopropane-1-carboxylic acid. Anal. Biochem., 100, 140-5. [16] Kawa-Miszczak, L., Wegrzynowicz-Lesiak, E., & Saniewski, M. (2005). Retardation of tulip shoot senescence by auxin. Acta Hort., 669, 183-90. [17] Arditti, J., Jeffrey, D.C., & Flick, B.H. (1971). Post-pollination phenomena in orchid flowers. III. Effect and interactions of auxin, kinetin or gibberellin. New Phytologist, 70, 1125-41. [18] Ketsa, S., & Rugkong, A. (2000). The role of ethylene in enhancing the initial ovary growth of Dendrobium ‘Pompadour’ following pollination. J. Hortic. Sci. Biotechnol., 75, 451-4. [19] Zhang, X.S., & O’Neill, S.D. (1993). Ovary and gametophyte development are coordinately regulated following pollination by auxin and ethylene. The Plant Cell, 5, 40318. [20] Ketsa, S., Wisutiamonkul, A., & van Doorn, W.G. (2006). Auxin is required for pollination-induced ovary growth in Dendrobium orchids. Funct. Plant Biol., 33, 887-92. [21] Stead, A.D. (1992). Pollination-induced flower senescence: a review. Plant growth regul., 11, 13-20. [22] O’Neill, S.D. (1997). Pollination regulation of flower development. Annu. Rev. Plant Physiol. Plant Mol. Biol., 48, 547-74. [23] Halevy, A.H. (1995). The role of sensitivity to ethylene in pollination-induced corolla senscence syndrome. Acta Hort., 405, 210-5. [24] Abebie, B., Philosoph-Hadas, S., Lers, A., Goren, R., Huberman, M., Riov, J., & Meir, S. (2006). The differential effectiveness of two synthetic auxins in delaying floret abscission in red cestrum cut flowers depends on their transport and metabolism. In Proceedings 33rd PGRSA Annual Meeting,

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K. Rungruchkanont, Sci. J. UBU, Vol. 2, No. 1 (January – June, 2011) 1-11

Quebec, Canada, pp. 75-9. [25] Abeles, F.B., Morgan, W.P., & Salveit, M.E. (1992). Ethylene in Plant Biology. San Diego: Academic Press, Inc.

11

[26] van Doorn, W.G. (2002). Effect of ethylene on flower abscission: a survey. Ann. Bot., 89, 689-93.

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SCIENCE JOURNAL Ubonratchathani University

Sci. J. UBU, Vol. 2, No. 1 (January – June, 2011) 12-16

http://scjubu.sci.ubu.ac.th

Research Article

Encapsulation of Acorus Calamus Linn. Extract by Polyurethane Microcapsules A. Paphonngam, A. Kongdee*, K. Buttaraj Chemistry Program, Faculty of Science, Maejo University, Chiang Mai 50290, Thailand.

Received 10/03/10; Accepted 01/02/11

Abstract Acorus calamus Linn. or “Wann-Nam” in local name is a medicinal plant. Since extract from its leaves and rhizomes is used for medicinal purpose and exhibits anti-microorganism property. It will be used to finish cotton fabic as follows. Polyurethane (PU) microcapsules containing the extract was prepared from interfacial polymerization of diphenyl methylene diisocyanate (MDI) in organic phase and propylene glycol in aqueous phase, with the addition of the extract in water. Fourier Transform Infrared Spectroscopy (FTIR) and UV-Vis were used to characterize microcapsules and the extract. Scanning Electron Microscopy (SEM) was used to examine morphology of microcapsules. Antibacterial function against Stapphylococcus aureus and Salmonella thyphimurium of the extract and polyurethane microcapsules containing the extract will be tested. The encapsulated will be finished onto cotton fabrics to study anti-bacterial property before and after washing. Keywords: Polyurethane, Microcapsules, Encapsulation, A. calamus.

taken into consideration. Moreover, it is important to release the contents to the In recent years, microcapsules have been exterior of the microcapsule [1-4]. intensively used in various kinds of application fields such as recording material, A. calamus is a medicinal plant, its rhizome supplementary foods and industrial material. was used extensively as medicine composiIn many cases, two factors are required for tion. The main constituents of A. calamus are characteristics. The contents must be held in monoterpenes, sesquiterpenes, phenylpromicrocapsules stably until it is required. panoids, flavonoids and quinine. Crude Therefore, the interaction between the extract of A.calamus has been reported as microcapsule wall and the contents and the bactericide function agains Stapphylococcus dynamic strength of microcapsules must be aureus and Salmonella thyphimurium [5-11]. In this investigation, extract of A.calamus is used as the content of microcapsules due to *Corresponding author. its antibacterial function. Polyurethane E-mail address: arunee.k@ mju.ac.th 1. Introduction

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A. Paphonngaml et al., Sci. J. UBU, Vol. 2, No. 1 (January-June, 2011) 12-16

13

microcapsules were prepared to encapsulate Scanning Electron Microscopy (SEM) was the extract, they will be further used as used to observe surface of PU and encapsulated PU microcapsules using Scanning finishing agent for cotton fabric. electron microscope (JEOL 5410 LV). The samples were coated with gold by sputtering. 2. Materials and Methods The observation was performed with suitable Preparation of Extract. A.calamus was cut acceleration voltage and magnification. into small pieces and ground using miller. 100 ml of 80% ethanol was added into 10 g of A.calamus powder. Bioactive compounds containing in A.calamus was extracted by sonication at room temperature for 1 h. The mixture was filtered. Water in the filtrate was evaporated using rotary evaporator. The remained solution was then dried in an oven at 45 ºC for 24 h. Preparation of Microcapsules. Polyurethane microcapsules were prepared from interfacial polymerization of propylene glycol and diphenyl methylene diisocyanate (MDI) in different phases as the following procedures: O1 : 2 g of propylene glycol and 4 g of polyethylene glycol were dissolved in 90 ml of toluene. O2 : 0.75 g of diphenyl methylene diisocyanate (MDI) and 0.05 g of dibutyl tin dilaureate (DBDL) were dissolved in 15 ml of the solution O1. W : 5 g of plant extract in 100 ml of the water. W was added into O1. Then the mixture was vigorously stirred by a mechanical stirrer (IKA yellow line OST 20) at 2000 rpm for 5 min at room temperature. O2 was subsequently added to the primary emulsion and stirred at 700 rpm for 10 min. Stirring speed was subsequently lower to 500 rpm. PU microcapsules were allowed to form at 60 ºC for 4 h. The microcapsules were filtered and washed with 50% ethanol and water, then dried at 50 ºC for 12 h. Characterization of Microcapsules. Fourier Transform Infrared Spectroscopy (FTIR) was used to proof chemical structure of PU, encapsulated PU and dried extract. The experiments were performed on Fourier transform spectrometer (Perkin Elmer FTIR spectrum RX). The spectra of samples in KBr pellet were collected from 400 to 4000 cm-1.

Release Rate of Microcapsules. 0.25 g of microcapsules containing A. calamus extract was put into 250 ml distilled water at room temperature for 180 min. At 10 min interval, 25 ml of the mixture was collected. Supernatant obtaining from 3000 rpm centrifugation of the mixture for 5 min was used for spectrophotometric analysis at 240 nm with a UV-Vis spectrophotometer (Hitachi U-2900). 3. Results and Discussion Polyurethane microcapsules containing A. calamus extract were successfully prepared by interfacial polymerization at 60 ºC for 4 h. Slurry mixture were obtained during polymerization. White and yellow-brown powders of PU microcapsules and encapsulated PU microcapsules, respectively, were received after filtering and drying. Figure 1 illustrates FTIR spectra of A. calamus extract, polyurethane microcapsules and polyurethane microcapsules containing the extract. Spectrum of A.calamus extract showed a board absorption band at 3200–3800 cm-1 which corresponds to –OH of remained moisture in the extract. The bands at 2960 and 1680 cm-1 corresponds to =C-H and C=C stretching, respectively. While C-O-C stretching is observed at 1100 cm-1. The absorption bands of polyurethane microcapsules and polyurethane microcapsules containing the extract appear at 3450-3330 cm-1 which corresponds to N-H stretching. The bands at 3000-2900 cm-1 are =C-H stretching, the bands at 1710 and 1630 cm-1 are C=O stretching. N-H bending appears at 1600-1400 cm-1. C-O-C stretching is observed at 1100 cm-1 and =C-H bending at 850-600 cm-1. Indication of the extract encapsulated in PU microcapsules is the shifts of C=C band from 1630 to 1710 cm-1 and N-H band from 3360 to 3410 cm-1.

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14

Encapsulation of Acorus calamus Linn. Extract by Polyurethane Microcapsules

SEM images showed rather sphere microcapsules which are less than 10 µm in size, their surface is rough as seen in Figure 2. Release rate of A.calamus extract is illustrated in Figure 3. The rate is dramatically increased at the beginning due to increased absorbance of supernatant. Until 40 min, plateau of release graph is observed, however the graph is gradually declined after

immersing the capsules for 90 min. Release curve revealed swollen behavior of PU capsules in water. Upon swelling, pores of the capsules was opened, the extract was released until the maximum value was reached. Beyond 90 min, amount of the extract in the capsules was lower, thus, decrease in release amount was determined.

Figure 1. FT-IR spectra of (A) A.calamus extract, (B) polyurethane microcapsules, (C) polyurethane microcapsules containing the extract.

A

B

Figure 2. SEM image of polyurethane microcapsule containing the extract (A) x 2000 and (B) x 3500.

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A. Paphonngaml et al., Sci. J. UBU, Vol. 2, No. 1 (January-June, 2011) 12-16

15

Figure 3. Release rate of A.calamus extract from PU.

4. Conclusions

40 min immersed in water. Microcapsules containing A.calamus extract will be further used as finishing agent for cotton fabric. Washing resistance of the microcapsules and release of the extract from the capsules coated on cotton surface will be subsequently analyzed.

PU microcapsules containing A.calamus extract was successfully prepared by interfacial polymerization. Inclusion of the extract in polyurethane microcapsules was observed by the shifts of C=C band from 1630 to 1710 cm-1 and N-H band from 3360 t0 3410 cm-1. SEM images illustrated capsules surfaces and Acknowledgements size which is less than 10 µm. By using spectrophotometric determination at 240 nm, This research was financially supported by release amount of the extract was highest at the Thailand Research Fund (TRF-MAG).

References [1] Sawada, K., & Urakawa, H. (2005). Prepara tion of photosensitive color producing micro capsules: utilizing insitu polymerization method. Dyes and Pigments, 65, 45-9. [2] Kubota, Y., Suzuki, M., & Shinbo, K. (2001). Pressure and heat-sensitive recording material. Japan Patent. 174458. [3] Seki, S., & Hata, T. (2003). Heatsensitive recording material. United State Patent. 2003036478. [4] Halbrook, WB., & Wehr, MA., Thermosensitive recording material with preprinted indicia and thermal paper. Europe Patent.

1321307. [5] Valsaraj, R., Pushpangadan, P., Smitt, U.W., Adsersen, A., & Nyman, U. (1997). Antimicrobial screening of selected medicinal plants from India. Journal of Ethnopharmacology, 58, 75-83. [6] Dubey, R., Shami, T.C., & Bhasker Rao, K.U. (2009). Microencapsulation technology and applications. Defence Science Journal, 59, 82-95. [7] Saihia, D., Vroman, I., Giraud, S., & Bourbigotba, S. (2005).capsulation of ammonium phosphate with a polyurethane shell

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16

Encapsulation of Acorus calamus Linn. Extract by Polyurethane Microcapsules

part I: Microencoacervation technique. Reactive & Functional Polymers, 64, 127– 38. [8] Hirech, K., Payan, S., Carnelle, G., Brujes, L., & Legrand, J. (2003). Microencapsulation of an insecticide by interfacial polymerization. Powder Technology, 130, 324 –30. [9] Rodrigues, S.N., Fernandes, I., Martins, I.M., Mata, V.G., Barreiro, F., & Rodrigues, A.E. (2008). Microencapsulation of limon-

ene for textile application. Industrial & Engineering Chemistry Research, 47, 4142-7. [10] Park, S.J., Shin, Y.S., & Lee, J.R. (2001). Preparation and characterization of microcapsules of microcapsules containing lemon oil. Journal of Colloid and Interface Science, 241, 502-8. [11] Wiley, J., & Sons. (2005). Encyclopedia of Polymer Science and Technology: Microencapsulation, 12, 1-29.

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SCIENCE JOURNAL Ubon Ratchathani University

Sci. J. UBU, Vol. 2, No. 1 (January-June, 2011) 17-24

http://scjubu.sci.ubu.ac.th

Research Article

Forecasting Malaria Incidence Based on Monthly Case Reports and Climatic Factors in Ubon Ratchathani Province, Thailand, 2000 – 2009 W. Sriwattanapongse *, S. Me-ead , S. Khanabsakdi Department of Statistics, Faculty of Science, Chiang Mai University, Mueang, Chiang Mai, 50200 Thailand. Received 01/05/11; Accepted 30/06/11

Abstract Base on Malaria count data report from 2000-2009 in Ubon Ratchathani province of north-eastern Thailand, malaria incidence rates are computed by rain, mean temperature, minimum temperature, maximum temperature, humidity and month. Linear regression model, Poisson and Negative binomial GLM containing additive effects associated with the season of the year, climatic factors and the malaria incidence rates in the previous months provides a good fit to the data, and can be used to provide useful short-term forecasts. Although the season, rain, mean temperature, minimum temperature, maximum temperature, humidity effects are all highly statistically significant, by far the best predictor of the number of new cases occurring in any month is the disease incidence rate in the preceding month. Having a model that provides such forecasts of disease outbreak. Keywords: Ubon Ratchathani, Linear regression model, Poisson GLM, Negative binomial GLM.

1. Introduction Malaria is a biological phenomenon where all the three elements of the infection system, namely man, mosquito and parasite are influenced by various environmental variables. Approximately 40% of population of the world, mostly those living in the world’s poorest countries, is at risk of malaria. Every year, more than 500 million people become severely ill with malaria [1]. Malaria in Thailand is endemic in forest regions and most prevalent along the national *Corresponding author. E-mail address: [email protected]

borders, particularly on the border with Myanmar to the east. Although malaria cases and deaths have fallen substantially since 1999, the disease remains a considerable public health problem. The objective of this study is to identify the patterns of hospitaldiagnosed malaria incidences by using rain, mean temperature, minimum temperature, maximum temperature, humidity and month in Ubon Ratchathani province of northeastern Thailand. Ubon Ratchathani occupies an area of 16,112.650 square kilometers. It is away from Bangkok about 630 kilometer and it has population number of 1,803,754, and border Salavan and Champasak of Laos, to

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Forecasting Malaria Incidence Based on Monthly Case Reports and Climatic Factors

18

the south Preah Vihear of Cambodia. This province consists of the 25 districts : Mueang Ubon Ratchathani, Si Mueang Mai, Khong Chiam, Khueang Nai, Khemarat, Det Udom, Na Chaluai, Nam Yuen, Buntharik, Trakan Phuet Phon, Kut Khaopun, Muang Sam Sip, Warin Chamrap, Phibun Mangsahan, Tan Sum, Pho Sai, Samrong, Don Mot Daeng, Sirindhorn, Thung Si Udom, Na Yia, Na Tan, Lao Suea Kok, Sawang Wirawong and Nam Khun. Based on individual hospital case records routinely reported in the province from 20002009,incidence are computed by rain, mean temperature, minimum temperature, maximum temperature, humidity and month, and models for incidence rates are used to assess these climatic factors. Monthly disease counts in individual cells defined by rain, mean temperature, minimum temperature, maximum temperature, humidity and month are mostly small, so Poisson and negative binomial generalized linear models (GLM) are more statistically appropriate, and can be used to identify with unusually high disease occurrences [2].

maximum temperature group ( ≥ 3 1 .0 − < 3 3 .0 , ≥ 3 3 .0 − < 3 6 .0 a n d ≥ 3 6 .0 )

,

humidity group ( ≥ 60.0− < 65.0,≥ 65.0− < 70.0 , ≥ 70.0 − 75.0,≥ 75.0− < 80.0 and ≥ 80.0 )

and month, with rain group, mean temperature group, minimum temperature group, maximum temperature group, humidity group and calendar month as categorical determinants. Such incidence rates generally have positively skewed distributions so it is conventional to transform them by taking logarithms. And since monthly disease counts based on small regions are often zero, it is necessary to make some adjustment to avoid taking logarithms of 0: the method we use is to define outcome as n ijkt ⎞ ⎛ (1) y = ln 1 + K ijkt

⎜ ⎝

P

⎟ ⎠

where nijkt is the number of disease cases in the cell, P is the population at risk, and K is a specified constant. To allow for serial correlations in successive months, lagged incidence rates are included as additional determinants. Such an observation-driven model with m lags could take the form [3] yijkt = μ + αi + β j1 + β j 2 + β j 3 + ϕk +ηs + ∑l =1 γ l yijk ,t −l + ε ijkt m

2. Methods Data used in the current study are secondary data. First, it was taken from Vector Borne Infectious Diseases, 7th Disease Prevention Control, Ubon Ratchathani. Second, the climate data comprise of rain, temperature and humidity from the North-eastern meteorological center, Ubon Ratchathani. Incidence rates were computed as the number of cases per 10,000 residents in the district according to the 2000 Population and Housing Census of Thailand. The simplest model is based on linear regression with the outcome variable defined as the incidence rate in a cell indexed by rain group ( < 35.01,≥ 35.01 − 90.01 and > 90.01), mean temperature group ( ≥ 22.0 − < 25.0,≥ 25.0 − < 28.0 and ≥ 28.0 − < 32.0 ), minimum temperature group ( ≥ 11.20 − < 15.34,≥ 15.34 − < 19.48 and ≥ 19.48 ) ,

(2) where N ijkt is a random variable denoting the reported number of disease cases in rain group i, mean temperature group j1 , minimum temperature group j2, maximum temperature group j3 , humidity group k and month t for the region of interest and nijkt is the corresponding number observed, Yijkt is the outcome variable specified in Eq. (1) and yijkt the corresponding number observed, while ε ijkt comprises a set of independent normally distributed random variables with mean 0, and s = mod(t, 12). In this model we constrain the parameters so that α 1 =0, β 1 =0,

ϕk =0

and η1 = 0. While linear time trends

could be included in the model, they are less useful for short-term forecasting purposes in the presence of high

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W. Sriwattanapongse et al., Sci. J. UBU, Vol. 2, No. 1 (January-June, 2011) 17-24

19

serial correlations, and are not considered in distributions with means λ and 1 − π , t t the present study. Davis suggested respectively, with the following probability observation-driven models for time series function: counts N t based on the Poisson distribution

with mean λt where ln (λt ) is expressed as Prob[Nt = n] = an additive function of determinants and lagged observations on N t [4]. While these models are not appropriate for disease epidemics because they express the mean of the process at time t as an exponential function of lagged observations on the same process and are thus numerically unstable when substantial variations occur, they become stable when the lagged observation are replaced by logged incidence rates. Thus if pijkt is the population in rain group i and temperature group j ,humidity group k and month t and λijkt is the mean of N ijkt , a suitable generalized linear model based on the Poisson distribution could take the form: ln ( λijkt ) = ln ( pijkt ) + μ + α i + β j1 + β j 2 + β j 3 +

(3)

ϕ k + η s + ∑ l =1 γ l yijk ,t −l m

Poisson models for disease counts are often over-dispersed due to spatial or temporal clustering of cases [5], in which case the negative binomial distribution may be more appropriate. This distribution has an additional parameter γ and takes the form γ

Prob[Nt=n]

=

⎛ γ ⎞ ⎛ λt ⎞ Γ (n+γ ) ⎜ ⎟ ⎜ ⎟ Γ ( n + 1 )Γ ( γ ) ⎝ γ + λ t ⎠ ⎝ γ + λ t ⎠

n

(4)

λt is the conditional expected value of N t , but the 2 conditional variance is λt + λt γ [6]. The parameter γ is actually inversely related to

As for the Poisson model,

the over-dispersion, so that the Poisson model arises as the special case in the limit as γ → ∞ .Disease counts in months often have an excessive number of cells containing zero counts. For such situations, a zeroinflated Poisson model is given as a mixture of independent. Poisson and Bernoulli

π t + (1 −π t ) exp( − λt ) (1 −π t )

λt n n!

for n = 0

exp( − λt )

for n > 0

(5) where the logit of 1 − π t has the same functional form as the logarithm of

λt

given in

Eq. (4) , the conditional means of this distribution is λt (1 − π t ) and its conditional variance

is

λt (1 − π t )(1 + π t λt ) .

The

binomial model is also given by an equation similar to Equation (5), with the same mean, although in this case the conditional variance is λt (1 − π t )(1 + π t λt + λt ) . γ The need for a zero-inflated model instead of its corresponding simpler analogue may be assessed using a statistical test [7]. In conventional linear regression models where errors are assumed to be independent and normally distributed, the adequacy of the model can be assessed by plotting residuals, obtained from the observations simply by subtracting their conditional means, against corresponding normal scores. For count data where a GLM is fitted, (Pearson) residuals are defined by subtracting the conditional means from the counts and then dividing by the conditional standard deviations [8]. These residuals can be plotted against scores based on the appropriate asymptotic distribution. If the model is correct these residuals should follow a straight line with unit slope. For Poisson-distributed models, this asymptotic distribution is the normal distribution, there by normal scores can still be used. For negative binomial time series models, the asymptotic distribution of the standardized residuals can be derived using moment generating functions. The moment generating function for the negative binomial distribution given by Eq. (5) is [9] ⎛ λ λ ⎞ E [ exp ( θ N t ) ] = ⎜ 1 + t − t e θ ⎟ γ γ ⎝ ⎠

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−γ

(6)

20

Forecasting Malaria Incidence Based on Monthly Case Reports and Climatic Factors

so the moment generating function for the rates in June 2000 (1.32 per 10,000 cases/year) and the minimum rates in May standardized residuals is 2005 (0.03 per 10,000 cases/year). ⎡ ⎛ θ ( Nt − λt ) ⎞⎤ (7) = E ⎢exp ⎜ ⎟⎥ 2 Time series plots of average monthly malaria ⎢⎣ ⎝ λt + λt γ ⎠⎥⎦ −γ incident rates per 10,000 persons in Ubon ⎛ λt λt ⎞ 2 Ratchathani province are shown in Figure 1. ⎜1+ − exp θ λt + λt γ ⎟ exp ( −θ λt 1+ λt γ ) ⎝

γ

γ

(

)⎠

Using a Taylor expansion, the right-hand side of Equation reduces to corresponding zeroinflated negative ⎛ ⎞ θ θ2 ⎜1 − ⎟ − ⎜ γ 1 λ + 1 γ 2γ (1 + λt γ ) ⎟ t ⎝ ⎠

−γ

(

)

exp − θ γ + o(1 / λt )

where o(x ) is any function that tends to zero faster than x . Since the limit of (1 + x n )− n as n → ∞ as e , it follows that the limiting distribution of the Pearson residuals has moment generating function −x

Figure 1. Monthly distribution of the incidence of malaria in Ubon Ratchathani, Thailand, 2000–2009.

Regression Model. When the liner regression model given by Eq. (2) with m=3 lags and (1 − θ γ ) −γ exp(−θ γ ) K=10,000 is fitted, the lagged incidence rates account for the largest single This corresponds to a gamma distribution contribution to the r-squared statistic with shape parameter γ and scale parameter (80.26%) and rain, mean temperature, minimum temperature, maximum tempera1/ γ , shifted to the left by γ . ture, humidity are all highly statistically significant. Figure 2 shows plots of residuals 3. Results and Discussion versus predicted values (left panel) and normal scores (right panel). Distributions of Incidence Rates. During the study period from January 2000 to December 2009, number of 8,215 cases of malaria were diagnosed in Ubon Ratchathani province. The number of cases reported in a month for a particular rain group, mean temperature group, minimum temperature group, maximum temperature group, humidity group varied from 5 to maximum of 264 Figure 2. Plots of residuals with from linear (incidence rate 1.7735 per 10,000), in rain model for log-transformed incidence rates. group less than equal 22.0 to less than 25.0, minimum temperature group more than or Although the normal score plot indicates that equal 19.48, maximum temperature group the residuals are well approximated by a more than or equal 33.00 to less than 36.00, normal distribution, the zero counts are humidity group more than or equal 60.00 to conspicuous, constituting the downward less than 65.00 in August 2000. sloping line in the plot of residuals against predicted values. The largest residual 0.3786 Highest rates occurred in August 2000 corresponds to just 264 cases, reported in (1.77 per 10,000 cases/year) and the second

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

W. Sriwattanapongse et al., Sci. J. UBU, Vol. 2, No. 1 (January-June, 2011) 17-24

21

Table 1. Results for models fitted to Malaria incidence. Determinant

Linear model

Constant Rain Group:

Mean temperature Group:

Minimum temperature Group:

Poisson GLM

Neg. Binomial GLM Coeff St.Error

Coeff

St.Error

Coeff

St.Error

1

0.6851 0

0.0893 1

0.2379 0

0.1009 1

0.0949 0

0.2941 1

2

-0.0854

0.0533

-0.3712

0.0700

-0.3415

0.1789

3

-0.2565

0.0763

-1.2796

0.1014

-1.2042

0.2563

1

0

1

0

1

0

1

2

-0.1386

0.0430

-0.3221

0.0400

-0.2989

0.1381

3

-0.1566

0.0919

-0.3962

0.1231

-0.3674

0.3084

1

0

1

0

1

0

1

2

0.0069

0.0381

-0.0111

0.0483

0.0097

0.1287

3

-0.0012

0.0743

-0.1519

0.1108

-0.1861

0.2579

Maximum temperature Group: 1

0

1

0

1

0

1

2

-0.0910

0.0496

-0.4680

0.0697

-0.4392

0.1685

3

-0.1636

0.0604

-0.6459

0.0785

-0.6785

0.2038

1

0

1

0

1

0

1

2

-0.1223

0.0434

-0.3110

0.0449

-0.2871

0.1411

3

-0.0502

0.0754

-0.0147

0.0924

-0.0122

0.2506

4

0.0564

0.1079

0.6541

0.1498

0.6253

0.3646

5 January

-0.0172 0

0.1136 1

0.2819 0

0.1598 1

0.2541 0

0.3850 1

February

0.0691

0.0486

0.1999

0.0531

0.2149

0.1591

March

-0.0024

0.0594

-0.1177

0.0725

-0.1634

0.1988

April

-0.0126

0.0635

-0.2853

0.0780

-0.1762

0.2132

May

-0.0070

0.0881

-0.0576

0.1220

-0.0677

0.3021

June

0.0864

0.0863

0.3434

0.1176

0.3550

0.2942

July

0.1092

0.0872

0.3955

0.1166

0.4309

0.2966

Humidity Group:

Month:

AR Lag:

df :93

August

0.0246

0.0835

0.1456

0.1160

0.1330

0.2855

September

-0.0389

0.0833

-0.0004

0.1162

-0.1400

0.2853

October

-0.0776

0.0827

-0.2615

0.1187

-0.2564

0.2846

November

-0.0821

0.0728

-0.2968

0.1002

-0.2739

0.2472

December

-0.0201

0.0497

-0.0832

0.0583

-0.0467

0.1643

1

0.2494

0.0901

0.7073

0.0929

1.0136

0.2944

2

0.0553

0.0919

0.3572

0.0978

0.3667

0.3017

3

-0.1574

0.0814

-0.5392

0.0865

-0.5729

0.2679

R-square:0.8026

Deviance:847.98

Deviance:124.52

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

22

Forecasting Malaria Incidence Based on Monthly Case Reports and Climatic Factors

2000. Table 1 gives the estimate coefficients reported in October 2000, respectively, the and their standard errors both for this linear Poisson GLM predicts 143.8206 for August model and for the corresponding GLMs 2000 giving the Pearson residual based on the Poisson and negative distribu264 − 143.8206 tions. = 10.0212 143.8206

Comparing the linear model based on the logtransformed incidence rates with the GLMs based on the disease counts, the results are similar in some respects but quite different in others. Each model has a seasonal peak during the period from May to August.

In contrast, even though the negative binomial GLM predicts a higher predicted number of cases for this cell (51.1895), it gives a smaller Pearson residual. Using the estimated dispersion parameter γ =10.36, the corresponding Pearson residual is

After the mosquito larvae deposited during 142 − 51.1895 = 5.2073. the rainy season become active. These effects 51.1895 51.1895(1 + ) however have smaller standard errors in the 10.36 GLMs. The autocorrelation coefficients are also similar in each model, although only two But the second higher predicted number of lagged terms are needed in the Poisson cases for this cell (102.3529) is model. 264 −102.3529 102.3529(1 +

102.3529 ) 10.36

= 4.84407.

The high residual deviance for the Poisson model is substantially higher than the number of degree of freedom, whereas the negative The negative binomial model also shows binomial GLM gives a much lower residual departure from a appropriate fit in the deviance. residuals plot. Although the higher residuals do not suggest any unusual pattern they are of Figure 3 compares the plots of Pearson interest because they could herald the residuals against their asymptotic scores beginnings of more serious diseases based on the two GLMs. The over-dispersion outbreaks. in the Poisson model is seen in the steep rise in the curve away from the unit-slope line for Some care is needed with the choice of the the residuals greater than zero. constant K, for the value chosen in the analysis (K=10,000), the sum of predicted disease for negative binomial is 8296.634 compared with 8,215 observed malaria cases in the 120 cells (10 ± 12 months). From Table 1, the plus and minus sign of coefficient would have increase and decrease Figure 3. Plots of Pearson residuals after that effect of each factors to malaria incident fitting GLMs to Malaria rates. model in that area case. There is also a highest outlier among the residuals from the Poisson model, corresponding to 264 malaria cases reported in August 2000 (at the height of the 2000 epidemic).Based on 108 case reported in the same rain, mean temperature, minimum temperature, maximum temperature, humidity in September 2000 and 118 cases

4. Conclusions We have shown that malaria is a serious health problem in Ubon Ratchathani province. The linear and poisson model provides an appropriate fit to rain, mean temperature, minimum temperature, maximum temperature, humidity, and month. The

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

W. Sriwattanapongse et al., Sci. J. UBU, Vol. 2, No. 1 (January-June, 2011) 17-24

23

will certainly affect the abundance and distribution of infectious disease and will cause changes in the epidemiology of infectious diseases. The ability of mankind to react or adapt is dependent upon the magnitude and speed of the change. The outcome will also depend on our ability to recognize epidemics early, to contain them According to this study, the prevalence of effectively, to provide appropriate treatment, malaria was high in rain group of less than and to commit resources to prevention and 35.01, mean temperature group of 22.00- research [14]. 25.00, minimum temperature group of more than 19.48, maximum temperature Strategies for reducing malaria involve not group of 33.00-36.00, humidity group of only gaining a better understanding of the 60.00-65.00 in August 2000. We found that risk factors for the disease and how it is the prevalence of malaria had decreased in propagated, but also choosing where and the study period. Devi investigated Pearson’s when to allocate limited resources for treating correlation analysis that relationship between newly diagnosed cases and preventing further climate variables and malaria transmission infections. Having a model that provides [10]. In addition, Bi et al was to explore the accurate short-term forecasts of disease impact of climate variable on the outbreaks, even if the model is based purely transmission of malaria and indicate that on statistical data analysis, can provide a climatic variables should be considered as useful basis for such resource allocation. For possible predictors for regions with similar example, it might involve allocating geographic, climatic, and socio-economic mosquito nets to families in specific. conditions to those of Shuchen County [11]. Hoshen and Morse developed a mathe- Acknowledgements matical-biological model that may be suitable for the simulation of malaria forecasts based We would like express our gratitude to the on seasonal weather forecasts [12]. Gagnon 7th Disease Prevention Control, Ubon et al. found that there was a statistically Ratchathani for allowing us to use the disease significant relationship between El Niño and data from their office. malaria epidemics [13]. Global warming probability plot in this study was shown as an instrument for verifying the distributional assumption of a model and an effective way of capturing any unexpected increase in malaria counts. The Provincial Public Health office should severance increase in malaria count.

References [1] WHO (2007), Malaria Bulletin [2] Kaewsompak, S., Boonpradit, S., Choopradub, C. & Chaisuksan, Y. (2005). Mapping acute febrile illness incidence in Yala province, Songklanagarind J. Medicine, 23(6),455-62. [3] Sriwattanapongse W, Kuning M, Jansakul N. (2008). Malaria in North – Western Thailand,. Songklanakarin J Sci Technol; 30(2):207-14. [4] Davis, R., Dunsmuir, W., & Streett, S. (2003). Observation-driven models for Poisson counts, Biometrical 90(4), 777-90.

[5] Ruru, Y., & Barrios, E.B.(2003). Poisson regression models of malaria incidence in Jayapura: Indonesia, The Philippine Statistician, 52(1-4), 27-38. [6] Jansakul, N., & Hinde, J.P. (2004). Linear mean-variance negative binomial models for analysis of orange tissue-culture data, Songklanakarin J. Science and Technology, 26(5), 683-96. [7] Vuong, Q. H. (1989).Likelihood ratio tests for model selection and non-nested hypotheses. Econometrica 57, 307–33.

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24

Forecasting Malaria Incidence Based on Monthly Case Reports and Climatic Factors

[8] Kedem, B., & Fokianos, K. (2002). Regression Models for Time Series Analysis, UK: John Wiley & Son. [9] Wackerly, D.D., Mendenhall, W.,& Schaeffer, R.L. (1996). Mathematical statistics with applications, 5th edition, Duxbury Press. [10] Devi, N. P., & Jauhari, R .K. (2006). Climatic variables and malaria incidence in Dehradun,Uttaranchal, India, J.Vector Borne Disease, 43(1), 21–8. [11] Bi, P., Tong, S., Donald, K., Parton, K. &Ni, J. (2003). Climatic variables and transmission of malaria: a 12-year data analysis in

Shuchen County, China, Public Health Reports 1, (118), 65-72. [12] Hoshen, M.B., & Morse, A.P., (2004). A weather-driven model of malaria transmission, Malaria J., 3, 32-46. [13] Gagnon, A.S., Smoyer-Tomic, K.E.,& Bush, A.B. (2002). The El Niño Southern Oscillation and malaria epidemics in South America, International J. Biometeorology, 46(2), 81-9. [14] Khasnis, A.A., & Nettleman, D. M. (2005) Global Warming and Infectious Disease, Archives of Medical Research, 36, 689–96.

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SCIENCE JOURNAL Ubon Ratchathani University

Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 25-35

http://scjubu.sci.ubu.ac.th

Research Article

Gas Exchange Properties of Fresh Chillies and Their Applications in Mathematical Modeling of Packaging Systems W. Utto1*, T.R. Robertson2, D.J. Tanner3 1

2

Faculty of Agriculture, Ubon Ratchathani University, Thailand. School of Engineering and Advanced Technology, Massey University, Palmerston North, New Zealand. 3 ZESPRI International Ltd., New Zealand. Received 06/01/12; Accepted 15/06/12

Abstract Key postharvest physiological properties including respiration rates and skin permeances under a range of storage temperature (5 to 30ºC) were characterised and then incorporated together with the dimensionless Biot number (Bi) in a mathematical model for packaging design. Respiration rates dramatically increased as storage temperatures were increased whilst skin permeances, as previously unidentified evidence, were slightly increased. The major pathway for gas exchanges of chillies was dominated by movement through the stem area. Calculated Bi values were substantially less than 0.1 indicating uniform concentrations of CO2 and O2 in chilli cavity (air). Utilisation of the steady-state model to express gas exchange between fruit and immediate environment can reasonably be justified. Keywords: Chillies, Respiration rates, Skin permeances to oxygen and carbon dioxide, Biot number.

1. Introduction Fresh chillies are one of the most economically important vegetables worldwide [1], however chillies are highly perishable under ambient storage atmosphere (~21% O2 and ~0.03% CO2) and temperatures [2-4]. Postharvest physiological changes in fresh produce during storage, handling and entering marketing chains cannot be stopped, but they can be slowed within certain limits. There are several biological processes, particularly respiration that lead to reduction in quality attributes, i.e. appearance, texture or flavour. The rapid *Corresponding author. E-mail address: [email protected]

changes in these attributes can rapidly lower the marketability of fresh produce [5]. The O2 required for respiration and the CO2 produced within the fruit, move to and from the fruit by a process of diffusion. Diffusion is a spontaneous process leading to the net movement of a material from one region to an adjacent one, from high concentration to low concentration [6]. The rate of gas diffusion depends on the properties of the gas molecules, the magnitude of the gradient (between fruit and environment), the physical properties of the intervening barriers such as the skin of the fruit and the temperature of immediate environment. The presence of

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

26

Gas Exchange Properties of Fresh Chillies and Their Applications in Mathematical Modeling

barriers such as the fruit skin and the consumption of O2 through respiration, creates a decrease in the concentration of O2 within the plant organ, while the production of CO2 creates an increase in CO2 within the plant organ [7].

routes through the fruit surface. Skin ' permeances to O2 and CO2 ( PO' 2 and PCO , 2 respectively) are similar when gas diffusion ' is dominated by pores, whilst if PCO is much 2 higher than PO' 2 , the major gas diffusion route is through the skin cuticle [10].

Although there is a number of studies on postharvest management of fresh chillies, information on their physiological properties, particularly skin permeances to O2 and CO2 has not been reported. The objective of the current study was to measure the respiration rate and skin permeances of chillies at temperatures normally used for their storage and handling.

Solanaceous fruits such as capsicums (including chillies) tomatoes and eggplants lack stomata or lenticels and as a result have much lower permeances than fruit and vegetables with these pores [11]. In peppers gas exchange is thought to mainly occur through the stem end [10]. Permeances to O2 and CO2 of bell pepper have been reported to be relatively similar and range from 580 pmol⋅s-1⋅m-2⋅Pa-1 (cv. Tasty) [12] to 2. Theory 790 pmol⋅s-1⋅m-2⋅Pa-1 (cv. Samanta), [10]. The resistance that the skin presents to gas However the skin permeances to O2 and CO2 exchange can be described as the permeance of hot chilli peppers have not been reported. of a plant organ to a gas. This is the rate at which the gas can diffuse in or out of the 3. Materials and Methods organ per unit of surface area for a given gradient of partial pressure. Permeances to Fresh Green Chillies. For all experiments, gas vary depending on the type of freshly harvested, mature field-grown green commodity, cultivar, part of the plant, chillies (Capsicum annuum L. cv. ‘Cayenne’) severity of preparation, and stage of maturity. were obtained directly from local growers Permeance appears to be little affected by (Feilding, New Zealand). Chillies were temperature [8,9]. The magnitude of washed and allowed to dry at room permeance to O2 and CO2 combined with the temperature (≈20°C) for 1 hour, following respiration rates, determines the internal O2 Lee et al. [13]. They were then dried with a i ( p Oi 2 ) and CO2 ( p CO ) concentrations for paper kitchen towel to remove any residual 2 surface water before being separated, at any given postharvest condition. An increase random for further experiments. in storage temperature leads to a rapid rise in respiration rate, but skin permeance does not Measurements of Chilli Respiration Rate. respond as rapidly [12]. This leads to a Respiration rates were determined by i decline p Oi 2 and an increase in p CO . measurement of the change in partial pressure 2 Internal O2 of fruit with very low skin of CO2 (following Johnston et al. [14]). permeance can therefore decline rapidly at Individual chillies were sealed in 500ml high storage temperatures and can lead to opaque plastic jars and 1 ml headspace fermentation [10]. Fruit skin permeance to samples were removed through a septum at gases comprises the combined permeance of the top of the container immediately and one pores (stem scars, cut end of peduncles, hour after sealing. For CO2 analysis, gas surface punctures and other discontinuities of samples were injected into a gas analyser the outer integument) and cuticle fitted with a miniature infrared CO2 (Analytical Development (interspersed with stomata and/or lenticels) transducer Company, Hoddeson, UK) using O2-free N2 [7]. The total fruit skin permeance to CO2 -1 as the carrier gas at 30 ml min ; data collectand O depends on the dominant diffusion 2

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

27

W. Utto et al., Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 25-35

ion was via a Hewlett Packard integrator (model 3396A). This was calibrated with external CO2 standards (certified as βstandard by B.O.C. Gases New Zealand Ltd). Ambient pressure data were collected using a pressure transducer (Barigo Electronic Altimeter, Barigo Barometerfabrik GMBH, D-7730 Villingen Schwenningen, Germany). Fruit surface temperature was logged (Squirrel model 1206, Grant Instruments, Cambridge, U.K.) using thermistor probes (FF type, U bead, ±0.2ºC; Grant Instruments, Cambridge, U.K.) attached onto the skins of selected fruits. Respiration rate was then calculated according to Eq. (1). rCO2 =

Vnet ⋅ Δp CO2

(1)

R ⋅ T fr ⋅ M fr ⋅ t

where rCO2 is rate of CO2 generation (mol· -1

-1

kg ·s ), Vnet is net volume as difference between fruit and jar volumes (m3), Δp CO2 is

Auckland, New Zealand). Once the cannulae were attached, fruit were separated into 4 groups and then equilibrated for 24 hr overnight in 4 different temperatures. Afterward, steady-state internal CO2 ( pCO2 ) and O2 ( p O2 ) at each storage temperature were determined by sampling 100 μl from the fruit cavities through the cannulae using a gas-tight syringe (Hamilton Gastight®, Hamilton Co., US). Values for chilli skin permeance to CO2 and O2 were then calculated using Eqs. (2) and (3), respectively, following Chen et al. [12]. Pfr' ,CO2 =

(p

Pfr' ,O2 =

(

rCO2 cav CO2

env − p CO 2

rO2 pOenv 2

−

pOcav 2

)

⋅

M fr

)

⋅

M fr

A fr

A fr

(2)

(3)

where P fr' ,CO2 , Pfr' ,O2 is chilli skin difference of partial pressure of CO2 to CO2 and O2, respectively quantified over the analysis time (Pa), R is permeance -1 -2 -1 3 -1 -1 (mol·s ·m ·Pa ), rCO2 , rO2 is rate of CO2 gas constant (8.314 Pa·m ·mol ·K ), T fr is of O2 consumption, fruit temperature (K), M fr is total mass of generation and rate -1 cav env respectively (mol·s ·kg-1), pCO , pCO is 2 2 fruit (kg), and t is analysis time (s). To determine fruit volume, a system similar to CO2 partial pressure in fruit cavity and the displacement platform-scale apparatus immediate environment, respectively (Pa), described by Mohsenin [15] was used. p Ocav , p Oenv is O2 partial pressure in fruit 2 2 Respiration rates of fresh chillies were cavity and immediate environment, determined at 5, 10, 20 and 30°C (averaging respectively (Pa), and A fr is surface area of 30 replicates per storage temperature). fruit (m2). Measurements of Cilli Skin Permeances to CO2 and O2. Fresh green chillies were used to Determinations of A fr were conducted determine internal atmosphere and skin following Clayton, Amos, Banks, & Morton permeances to O2 and CO2 at 5, 10, 20 and [16]. The estimated surface area of an 30°C (averaging 20 replicates per storage average chilli is 0.0034 ± 0.0004 m2 temperature). Cannulae (14 gauge stainless (measured with 30 chilli fruits). It should be steel needles, cut down to 2-cm length) were noted that the area of stem end was not taken inserted through the fruit wall into the cavity into account as this was assumed to be small, of the fruit (note this is a similar method as compared to the whole fruit. Rates of CO2 that utilised by Chen et al. [12] to measure skin permeances to CO2 and O2 of bell pepper). The connection between cannula and skin was sealed gas-tight using epoxy adhesive (5 min cure; Areldite®, Ciba-Geigy,

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

28

Gas Exchange Properties of Fresh Chillies and Their Applications in Mathematical Modeling

generation were referred to as rCO2 measured 4. Results and Discussion in experiments of respiration rates. Rates of O2 consumption were estimated from Eq. (4) following Talasila & Cameron [17], where the respiration quotient ( RQ ) presents the rCO2 to rate of O2 consumption ( rO2 ) at a

Respiration Rate. Respiration rates ( rCO2 ). of green ‘Cayenne’ chillies increased exponentially with increasing temperature (Figure 1). Values for respiration rate increased about 20 fold from 0.07 μmol⋅kg-1⋅s-1 to 1.34 μmol⋅kg-1⋅s-1 as temperature increased from 5 to 30°C. The estimates for respiration rate at reference max temperature ( rCO ) and activation energy 2 , ref

given temperature. Under oxidative respiration conditions RQ can reasonably be assumed to be unity. This assumption was adopted in the present work, because transfer rate measurements were conducted under ( Ea r max ) are given in Table 1, following ambient atmospheric conditions. CO2

Hertog et al [19] and Chen et al. [12]. The rCO2 variation in respiration rate of fresh chillies rO2 = (4) RQ increased with increasing storage temperatures (Figure 1). The exponential All collected data were expressed according relationship between temperature and to the units proposed by Banks, Cleland, respiration rate for fresh green chillies Cameron, Beaudry, & Kader [18]. The data confirmed that rCO2 is governed by on respiration rates and skin permeances to temperature [20]. The results observed in the O2 and CO2 were analysed using temperature current experiment coincide very well with dependence according Arrhenius’s law from literature data. For example Lee et al. [13] Eq. (5). The reference temperature for reported that estimated maximum rate of CO 2 Arrhenius’s law was in all cases fixed at evolution ( rCO2 ) of green chillies (Capsicum 15°C (288.15 K), following Chen et al. [12]. The non-linear equations were applied annum L. cv ‘Nogkwang’) at 10ºC was ~0.19 -1 -1 directly, using SAS (version 8; Statistical μmol·kg ·s . Krajayklang et al. [21] reported Analysis System), without transformation to that rate of CO2 production ( rCO2 ) of green data or equations. chillies (Capsicum annum L. cv ‘Caysan

k = k ref ⋅ e

Ea ⎛ 1 1 − ⎜ − R ⎜⎝ T Tref

⎞ ⎟ ⎟ ⎠

SPS705’) at 22ºC was ~0.47 μmol·kg-1·s-1.

(5)

where Ea is Energy of activation (J⋅mol-1), k is interested rate (exact unit depends on the rate), k ref is the interested rate at arbitrarily chosen reference ( Tref ), Tref is reference temperature (288.15 K, (15°C)), and T is measured temperature (K). Figure 1. Temperature effect on respiration of green ‘Cayenne’ chillies for temperatures ranging from 5 to 30°C. Solid line was fitted using Eq. (5).

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

W. Utto et al., Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 25-35

29

There is no literature data on Ea r max of green several researchers [3, 13, 24] have reported CO2 that chilling injury can occur in chillies or ‘Cayenne’ chillies and/or other types of hot peppers at temperatures lower than 7°C, this peppers. However, as the Ea r max of bell was not seen in this study. Mencarelli et al. CO2 reported that chilling injury peppers (Capsicum annum L. cv ‘Tasty’) [12] [25] susceptibilities of small peppers ( Capsicum is less than that of hot chillies, the respiration rate of bell peppers is less sensitive to frutescens L.) appeared to depend on stage of temperature than that of hot chillies. maturity. Peppers harvested at yellow stages Furthermore, the respiration rate of bell were likely be more sensitive to chilling injury at 5ºC, compared to fruits harvested at peppers (~0.01 μmol⋅kg-1⋅s-1, at 15°C) [12] is green and red stages. Mercado, Quesada, significantly lower than that of hot ‘Cayenne’ Valpuesta, Reid, & Cantwell [26] reported -1 -1 chillies (~0.32 μmol⋅kg ⋅s ). Considering that mature green bell peppers (Capsicum both Ea r max and respiration rate, the frutescens L. cv. ‘Galaxy’) kept at 5ºC under CO2 biological properties of hot peppers might be ambient atmosphere for 20 days showed expected to be more sensitive to temperature severe chill-induced decay (refer to calyx and surface decay), compared to full red peppers difference. which were still marketable. Mohammed et . [27] reported that there were no apparent al Chen et al. [12] reported that the energies of activation for bell pepper respiration (39869 differences in chilling injury susceptibility and 30947 J·mol-1, for rate of CO2 generation between two local hot pepper cultivars (i.e. and O2 consumption, respectively) were very not specified) grown in Trinidad. Incidences close to the energies of activation for LDPE of chilling injury of both cultivars were permeabilities to the respiratory gases (31751 observed when fruits were kept at 2-4ºC. and 30450 J·mol-1, for permeabilities to CO2 These were reported through visual and O2, respectively). In such a case, the gas appearances (such as calyx discolouration conditions inside the package were almost and surface browning) and biochemical insensitive to temperatures between 0 and property changes (i.e. electrolyte leakages 30°C. This could be beneficial for and bioelectrical resistances). Based on the maintaining the appropriate atmospheric gas literature evidence, it may be assumed that composition of modified atmosphere the chilli cultivars used for this study and packaging of chillies, as in developing their stages in maturity may have enabled countries such as Thailand and Vietnam, them to withstand chilling injury decay at 5ºC much of the time chillies and other storage temperature. horticultural products are subjected to varying conditions and are often stored and transported under high temperatures [22]. Keeping fresh green chillies at 5°C minimised the respiration rate (Figure 1), thus low temperature storage can be used for prolonging the shelf life of green ‘Cayenne’ chillies. However, some studies reported that the pepper (Capsicum annum L. cv. ‘Galben Superior’) is relatively susceptible to chilling injury at temperatures below 7°C thereby increasing the respiration rate and limiting the shelf life [23].

Skin Permeances to CO2 and O2. The skin permeance to CO2 and O2 ( Pfr' ,CO2 , Pfr' ,O2 , respectively) of chillies increased exponentially with increasing temperature (Figure 2). The estimates for activation energy and skin permeance to both CO2 and O2 at the reference temperature ( P fr' ,CO2 , ref and

Pfr' ,O2 , ref , respectively) are given in Table 2, following Chen et al. [12].

In the present work, there was no rise in respiration rate observed at 5°C. While

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

30

Gas Exchange Properties of Fresh Chillies and Their Applications in Mathematical Modeling

Table 1. Parameter estimates and standard errors (SE) resulting from non-linear regression analysis of the data on the effects of temperature on respiration. a max rCO 2 , ref

b

(SE)

R2 c

nd

69813 (3.11)

95.90

111

CO2

0.32 (4.06) a

Ear max a (SE)

max The standard errors (SE) are expressed as a percentage, relative to the estimated values. b CO , ref 2

r

production rate (μmol–kg-1–s-1) at reference temperature max maximum CO2 production rate ( CO , ref ); 2 observations.

r

c

R2

Tref

(= 15°C);

Ear max

is the maximum CO2

is the energy of activation (J–mol-1) of the

CO2

is the percentage variance accounted for by regression; d

In Figure 2, values for and increased only 2-fold as temperature increased from 5 to 30ο C, as compared to the 20-fold increase in respiration rate (Figure 1). This can be explained by the fact that the energy of activation for the CO2 and O2 skin permeances were comparably lower than the respiration activation energies of the two gases (Figure 1 and Table 1). The largest variations in and were found in fruits stored at 10 οC.

n is the number of

and immediate environment taking place in air through gaps underneath the stem plate. Similar findings on such main gas exchange pathways of peppers were reported by other researchers [10,28]. Recently Taksinamanee et al. [4] studied stomata of hot chilli pepper (Capsicum annum L. cv ‘Super hot’) using Scanning Electron Microscopy and reported that no stomata on the chilli surface was found. However the stomata were abundant at the pendicle areas which were postulated by Taksinamanee et al. [4] as an important water loss pathway from chillies. Based on evidence in the literature, it therefore could be suggested that the major pathway for gas exchange of green ‘Cayenne’ chillies occurs through the stem end, and as a consequence their skin permeances to CO2 and O2 are considered reasonably temperature independent.

Average energies of activation of common Figure 2. Temperature affects on skin films such as LDPE [9,12,29], polypropylene permeance to A) O2 ( Pfr' ,O2 , pmol⋅s-1⋅m-2⋅Pa-1); (PP) polyvinylchloride (PVC) and cellulose acetate [9] are 31100-39720, 46900, 56300 and B) CO2 ( P fr' ,CO2 , pmol⋅s-1⋅m-2⋅Pa-1), of and 21000 J⋅mol-1, respectively. These are green ‘Cayenne’ chillies held at temperatures higher than the activation energy of chilli between 5 and 30°C. Solid lines were fitted skin permeances (to both CO2 and O2) which are in a range of 18000-20000 J⋅mol-1 (Table using Eq. (5). 2). Hence plastic films are more sensitive to Chen et al. [12] found that skin permeances any temperature changes than gas of peppers was relatively temperature permeances of chilli skin. These incidences independent and postulated such findings as suggest that careful designs of MA plastic results of gaseous diffusion between fruits film packaging systems for horticultural

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31

W. Utto et al., Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 25-35

products are required when temperature fluctuations become significant. In such cases, active packaging technology would be incorporated into the systems for maintaining optimal MA conditions. For example example active CO2 scavenger systems could be utilised so as to reduce CO2 pressures when temperatures rise. Applications of active packaging systems for horticultural products can be found in Utto et al. [30] and Scully and Horsham [31].

value, with temperature increasing from 5 to 30°C. Whilst, the internal partial pressure of i ) increased approximately 9 times CO2 ( p CO 2 (Figure 3). As storage temperature is i increases increased, p Oi 2 declines and p CO 2 in chillies. This could be explained by the different degrees in which rate of respiration and skin permeances responds to the changes of temperatures. Because the energy of activation of respiration is larger than that of skin permeance in response to an increasing in temperature, respiration rate will increase faster than skin permeance. As a results, the level of internal O2 partial pressure progressively decreased, whilst the internal CO2 partial pressure continuously accumulated with increasing temperature.

In addition to skin permeances to O2 and CO2, the research findings suggested that changes occurred to the O2 and CO2 partial pressures inside the chilli cavity as a consequence of changes in environmental temperature (Figure 3). The internal partial pressure of O2 ( p Oi 2 ) declined to half its

Table 2. Parameter estimates and standard errors (SE) resulting from non-linear regression analysis of the data on the effects of temperature on chilli skin permeance to O2 and CO2. Gas (i)

P 'fr ,i ,ref

O2 CO2

a

(SE)b

185 (0.96) 197 (0.80)

Ea i (SE)c

R2

19948 (3.07) 18798 (2.74)

93.70 94.89

'

d

n

e

80 80

Pfr' ,i ,ref

is the skin permeance to gas i, (O2 and CO2), (pmol⋅s-1⋅m-2⋅Pa-1) at reference temperature Tref (15°C); b The standard ' errors (SE) are expressed as a percentage, relative to the estimated values; c Ea i is the energy of activation (J⋅mol-1) of skin

a

permeance to gas i;

d

R2

is the percentage variance accounted for by regression; e

n

is the number of observations.

In Figure 3, the total internal partial pressure i for O2 and CO2 ( p Oi 2 plus p CO ) was 2 approximately 21.57 ± 0.30 kPa. When the sum of internal partial pressures of O2 and CO2 was at or close to 21, almost the same values for the gases in air, it could be concluded that the route of gas exchange was through air [10,29]. Figure 3. Variation in A) internal partial pressure of O2 ( p Oi 2 ) and B) internal partial i pressure of CO2 ( p CO ) of green ‘Cayenne’ 2

chillies held at 5 and 30°C.

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32

Gas Exchange Properties of Fresh Chillies and Their Applications in Mathematical Modeling

Applications to Mathematical Modelling for Eqs. (7) and (8). The assumption that the Packaging Systems: A Case of Calculating concentration is uniform within a phase is Biot Number (Bi) for Justifying Uniformity of reasonably accurate when Bi < 0.1 [36]. Gas Concentration in Package Headspace. Mathematical models for predicting O2 and CO2 in MA package atmospheres have been developed principally based on mass balances of gaseous transport through packaging film and gas exchange between package atmosphere and product (hence respiration rate) [32-34]. A steady-state mass transfer model is one common approach used to describe the gas exchange because of its mathematical simplicity [32,35]. The generic equation for this approach is shown in Eq. (6) in which modelling CO2 exchange is an example.

Bi fr ,CO2 = Bi fr ,O2 =

k 'fr ,CO2 L fr

(7)

D cav fr ,CO2 k 'fr ,O2 L fr

(8)

D cav fr ,O2

where Bi fr ,CO2 Bi fr ,O2 is Biot numbers of CO2 and O2, respectively (dimensionless), k 'fr ,O2 is skin permeance to O2 (m·s-1) (note unit changed for consistency with the overall unit), L fr is characteristic dimension of fruit cav (m), and D cav fr ,CO2 , D fr ,O2 is diffusivity of

(

env N 'fr ,CO2 = rCO2 M fr − k 'fr ,CO2 A fr C cav fr , CO2 − C fr , CO2

) (6)

where

N 'fr ,CO 2

CO2 and O2 in fruit cavity (air) (m2·s-1). Value of L fr was determined by dividing fruit volume ( V fr ) by their respective surface

is rate of CO2 exchange area ( A fr ). In the present work, L fr is

m, giving an estimated (mol·s-1), k 'fr ,CO 2 is skin permeance to CO2 approximately 0.0025 -6 3 V fr as 8.75 × 10 m and A fr as reported (m·s-1) (note unit changed for consistency earlier. The calculation of Bi numbers for cav env with the overall unit), and C fr ,CO , C fr ,CO CO2 and O2 are summarised in Table 3. 2 2 is CO2 concentration in the fruit cavity and immediate environment, respectively All Bi values were substantially less than -3 (mol·m ). The key assumption of the steady- 0.1. This implies that the external resistance state model of gas exchange is the fruit (contributed by the skin permeances) is much internal atmosphere composition beneath the larger than the diffusive resistance of CO2 skin is considered uniform. Justifications for and O2 in the chilli cavity (air). A uniform this assumption therefore may be required. concentration of these gases in the cavity can To do so, Merts [32] utilised the thus be reasonably presumed and the dimensionless Biot number ( Bi ) to justify utilisation of the steady-state model to the uniformity of internal fruit concentration. express gas exchange between fruit and Bi is the ratio of external to internal immediate environment can reasonably be resistances to mass transfer across the fruit practical. skin contributed by the surface (e.g. skin permeance) and internal fruit properties (e.g. diffusivity), respectively. The Biot number of CO2 and O2 can be calculated according to

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33

W. Utto et al., Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 25-35

Table 3. Estimated Biot numbers for chilli gas exchanges of CO2 and O2.

k 'fr ,CO2

Temp. (ºC)

(m·s-1) 3.28 × 10-7 4.54 × 10-7 5.98 × 10-7 7.08 × 10-7

5 10 20 30 a

a

k 'fr ,O2

a

(m·s-1) 3.11 × 10-7 3.51 × 10-7 5.70 × 10-7 6.83 × 10-7

D cav fr ,CO2

b

(m2·s-1) 1.43 × 10-5 1.47 × 10-5 1.56 × 10-5 1.66 × 10-5

D cav fr ,O2

b

(m2·s-1) 1.80 × 10-5 1.85 × 10-5 1.97 × 10-5 2.09 × 10-5

Bi fr ,CO2

c

5.92 × 10-5 7.92 × 10-5 9.83 × 10-5 1.10 × 10-4

Bi fr ,O2

c

4.45 × 10-5 4.86 × 10-5 7.43 × 10-5 8.39 × 10-5

average chilli skin permeances to CO2 and O2 at individual temperatures; b Approximate mass diffusivities of CO2 and O2 in air

estimated using the correlation proposed by Fuller, Schettler, and Giddings [37]; c Characteristic dimension of fruit ( L fr ) is averagely 0.0025 m

5. Conclusions The present work identified respiration rates and skin permeances to O2 and CO2 of fresh chillies as well as extent to which storage temperatures have effects of on these two variables. Research findings substantiate hypothesis and theoretical knowledge on that the storage temperature is a key factor affecting gas exchanges properties of fresh chillies. The skin permeance information specifies key gas exchanging pathway which occurs through chilli stem ends. By

incorporating values of skin permeances into dimensionless Bi calculations could assist justifications for uniformities of gaseous concentrations in chilli cavity, and appropriateness of modelling steady-state gas exchange between the cavity and an immediate environment. Acknowledgements This project was financially supported by VSR Packaging Ltd. (Te Atatu, Auckland, New Zealand).

References [1] Bosland, P.W. and Votava, E.J., 1999 Peppers: vegetable and spice capsicums, (pp. 1-13) Peppers: vegetable and spice capsicums. [2] Lownds, N.K., Banaras, M., and Bosland, P.W. 1994. Postharvest water loss and storage quality of nine pepper (Capsicum) cultivars. HortScience, 29, 191-93. [3] Wall, M.M. and Berghage, R.D. 1996. Prolonging the shelf-life of fresh green chile peppers through modified atmosphere packaging and low temperature storage. Journal of Food Quality, 19, 467-77. [4] Taksinamanee, A., Srilaong, V., Uthairatanakij, A., and Kanlavanarat, S. 2006. Comparison of hydro-cooling and forced-air cooling on stomata closing at the pedicel of red hot chili cv. 'Superhot'. Acta Horticulturae, 712, 829-33. [5] Kays, S.J., 1991 Metabolic processes in harvested products, (pp. 75-142) Postharvest Physiology of Perishable Plant Products. Van Nostrand Reinhold: New York.

[6] Geankoplis, C.J., 1993 Principles of unsteadystate and convection mass transfer, (pp. 42687) Transport Processes and Unit Operations. Prentice Hall: New Jersey. [7] Kays, S.J., 1991 Movement of gases, solvents, and solutes, within harvested products and their exchange between the product and its external environment, (pp. 409-55) Postharvest Physiology of Perishable Plant Products. Van Nostrand Reinhold: New York. [8] Cameron, A.C., Boylanpett, W., and Lee, J. 1989. Design of modified atmosphere packaging systems - modeling oxygen concentrations within sealed packages of tomato fruits. Journal of Food Science, 54, 1413-16. [9] Cameron, A.C., Talasila, P.C., and Joles, D.W. 1995. Predicting film permeability needs for modified atmosphere packaging of lightly processed fruits and vegetables. HortScience, 30, 25-34.

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34

Gas Exchange Properties of Fresh Chillies and Their Applications in Mathematical Modeling

[10] Banks, N.H. and Nicholson, S.E. 2000. Internal atmosphere composition and skin permeance to gases of pepper fruit. Postharvest Biology and Technology, 18, 3341. [11] Blanke, M.M. and Holthe, P.A. 1997. Bioenergetics, maintenance and transpiration of pepper fruit. Journal of Plant Physiology, 150, 247-50. [12] Chen, X.Y., Hertog, M.L.A.T.M., and Banks, N.H. 2000. The effect of temperature on gas relations in MA packages for capsicums (Capsicum annuum L., cv. Tasty): an integrated approach. Postharvest Biology and Technology, 20, 71-80. [13] Lee, K.S., Woo, K.L., and Lee, D.S. 1994. Modified atmosphere packaging for green chili peppers. Packaging Technology and Science, 7, 51-58. [14] Johnston, J.W., Hewett, E.W., Hertog, M., and Harker, F.R. 2002. Temperature and ethylene affect induction of rapid softening in 'Granny Smith' and 'Pacific RoseTM' apple cultivars. Postharvest Biology and Technology, 25, 257-64. [15] Mohsenin, N.N., 1986 Physical characteristics, (pp. 79-127) Physical Properties of Plant and Animal Materials: Structure, Physical Characteristics, and Mechanical Properties. Gordon and Breach Science Publishers Inc.: New York. [16] Clayton, M., Amos, N.D., Banks, N.H., and Morton, R.H. 1995. Estimation of apple fruit surface area. New Zealand Journal of Crop and Horticultural Science, 23, 345-349. [17] Talasila, P.C. and Cameron, A.C. 1997. Freevolume changes in flexible, hermetic packages containing respiring produce. Journal of Food Science, 62, 659-64. [18] Banks, N.H., Cleland, D.J., Cameron, A.C., Beaudry, R.M., and Kader, A.A. 1995. Proposal for a rationalized system of units for postharvest research in gas-exchange. HortScience, 30, 1129-31. [19] Hertog, M.L.A.T., Peppelenbos, H.W., Evelo, R.G., and Tijskens, L.M.M. 1998. A dynamic and generic model of gas exchange of respiring produce: the effects of oxygen, carbon dioxide and temperature. Postharvest Biology and Technology, 14, 335-49. [20] Hardenburg, R.E., Watada, A.E., and Wang, C.Y., 1986 Summary of respiration and ethylene production rates, (pp. The Commercial Storage of Fruits, Vegetables, and Florist and Nursery Stocks U.S. Dept. of Agriculture, Agricultural Research Service: Washington, D.C.

[21] Krajayklang, M., Klieber, A., and Dry, P.R. 2000. Colour at harvest and post-harvest behaviour influence paprika and chilli spice quality. Postharvest Biology and Technology, 20, 269-78. [22] Nissen, R.J., George, A.P., Broadley, R.H., Hetherington, S., and Newman, S.M. 2006. Developing improved supply chains for temperate fruits in transitional Asian economies of Thailand and Vietnam. Acta Horticulturae, 335-42. [23] Burzo, I., Amariutei, A., and Craciun, C. 1994. Effect of low temperature on some physiological and ultrastructural changes of sweet pepper, eggplants and pod beans. Acta Horticulturae, 598-07. [24] Kader, A.A., Zagory, D., and Kerbel, E.L. 1989. Modified atmosphere packaging of fruits and vegetables. Critical Reviews in Food Science and Nutrition, 28, 1-30. [25] Mencarelli, F., Botondi, R., and Moraglia, D. 1989. Postharvest quality maintenance of new varieties of tomato, pepper and eggplant with small size fruits: preliminary results. Acta Horticulturae, 235-41. [26] Mercado, J.A., Quesada, M.A., Valpuesta, V., Reid, M., and Cantwell, M. 1995. Storage of bell peppers in controlled atmospheres at chilling and nonchilling temperatures. Acta Horticulturae, 134-42. [27] Mohammed, M., Wilson, L.A., and Gomes, P.I. 1992. Postharvest losses and quality changes in hot peppers (Capsicum Frutescens L.) in the roadside marketing system in Trinidad. Tropical Agriculture, 69, 333-40. [28] Bower, J., Patterson, B.D., and Jobling, J.J. 2000. Permeance to oxygen of detached Capsicum annuum fruit. Australian Journal of Experimental Agriculture, 40, 457-63. [29] Beaudry, R.M., Cameron, A.C., Shirazi, A., and Dostal-Lange, D.L. 1992. Modifiedatmosphere packaging of blueberry fruit: effect of temperature on package O2 and CO2. Journal of the American Society for Horticultural Science, 117, 436-41. [30] Utto, W., Mawson, A.J., Bronlund, J.E., and Wong, K.K.Y. 2005. Active packaging technologies for horticultural produce. Food New Zealand, 5, 21-32. [31] Scully, A.D. and Horsham, M.A., 2007 Active packaging for fruits and vegetables, in C.L. Wilson, Editor (pp. 57-71) Intelligent and Active Packaging for Fruits and Vegetables. CRC Press: Boca Raton. [32] Merts, I., Mathematical Modelling of Modified Atmosphere Packaging Systems for

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W. Utto et al., Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 25-35

Apples. 1996, Massey University: Palmerston North, New Zealand. [33] Yam, K.L. and Lee, D.S., 1995 Design of modified atmosphere packaging for fresh produce, in M.L. Rooney, Editor (pp. 55-73) Active Food Packaging. Blackie Academic & Professional: Glasgow. [34] Mannapperuma, J.D. and Singh, R.P., 1994 Modeling of gas exchange in polymeric packages of fresh fruits and vegetable, in R.P. Singh and F.A.R. Oliveira, Editors (pp. 43758) Minimal Processing of Foods and Process Optimization : An Interface. CRC Press: Boca Raton, FL. [35] Ben-Yehoshua, S. and Cameron, A.C., 1989 Exchange determination of water vapour, carbon dioxide, oxygen, ethylene and other gases of fruits and vegetables, in H.F.

35

Linskens and J.F. Jackson, Editors (pp. 17793) Modern Methods of Plant Analysis: Volume 9-Gases in Plant and Microbial Cells. Spinger-Verlag: Berlin. [36] Geankoplis, C.J., 1993 Principles of unsteady-state heat transfer, (pp. 330-80) Transport Processes and Unit Operations. Prentice Hall: New Jersey. [37] Johnson, A.T., 1999 Mass transfer, (pp. 494700) Biological Process Engineering : An Analogical Approach to Fluid Flow, Heat Transfer, and Mass Transfer Applied to Biological Systems. Wiley: New York.

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SCIENCE JOURNAL Ubon Ratchathani University

Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 36-42

http://scjubu.sci.ubu.ac.th

Research Article

Synthesis and Characterization of Carbon Nanotubes Using a Natural Precursor: Turpentine Oil C.R. Bhattacharjee1, A. Nath2*, D.D. Purkayastha1, B. Mukherjee3, Maheswar Sharon4, Madhuri Sharon5 1

Department of Chemistry, Assam University, Silchar 788011, Assam, India. 2 Department of Chemistry, S. S. College, Hailakandi, Assam, India. 3 Department of Chemistry, Pendharkar College, Dombivli, Maharashtra, India. 4 N.S.N. Research Centre for Nanotechnology and Bionanotechnology, Jambhul Phata, Ambernath (W) 421 505, Maharashtra, India. 5 MONAD Nanotech Pvt Ltd, A-702 Bhawani Towers, Powai, Mumbai 400 076, India. Received 29/04/11; Accepted 28/11/11

Abstract Pure Carbon nanotubes (CNTs) have been efficiently synthesized from Turpentine oil using Fe nanoparticles as a catalyst and also without the use of catalyst by simple Chemical Vapour Deposition (CVD) method at 900 °C. Turpentine oil (C10H16), a plant-based precursor was used as a source of carbon and argon as a carrier gas. The CNTs have been grown directly inside the quartz tube. The as grown carbon nanotubes were characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM), and X- Ray diffraction (XRD) studies. The XRD data indicated the presence of a high degree of crystallanity and the graphitic nature of the synthesized tubes. These tubes have been found to have outer diameters between 12 and 15 nm. Keywords: Carbon nano tubes, Chemical vapour deposition, Catalyst. *Corresponding author. E-mail address: abhijitnath1974@ rediffmail.com

1. Introduction During the past few years, carbon nanotubes have attracted much attention because of their unique physico-chemical and mechanical properties [1,2], as well as the potential applications such as in electron field emitters, biosensors, electrode material for solar cells, fuel cells and so on [3-7]. Carbon nanotubes

as light-weight and high performance materials are expected to bring major changes in the automobile sector by replacing steel or aluminum body and in the medicine area as a smart drug deliver [8]. From the application point of view it is highly desirable to synthesize wellordered arrays of nanotubes at low cost.

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C.R. Bhattacharje et al., Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 36-42

In general, carbon nanotubes are mainly synthesized by arc discharge, laser ablation, and chemical vapor deposition method [9– 11]. Compared with other methods, chemical vapor deposition (CVD) is a very effective technique because of low cost production, controllability, and the viability in highly dense synthesis of nanotubes at lower temperature on various substrates [12,13]. Recently, vertically aligned carbon nanotubes [14], multiwalled carbon nanotubes [15], and single-walled carbon nanotubes [16] were synthesized by this simple method using natural precursor as a carbon feedstock. The advantage of the natural precursors over conventional precursors (methane, acetylene, alcohol, benzene, etc.) lies in the reproducibility [14–16]. So far, there have been few reports about the synthesis of carbon nanotubes from natural precursors. In the present investigation, attempts were made to prepare the carbon nanotubes using turpentine oil as a carbon feedstock. 2. Materials and Methods The synthesis of CNTs was carried out using the CVD method. The process involved pyrolytic degradation of oil at suitable temperatures and atmosphere free from oxygen. Turpentine oil, derivable from the resin of pine trees, used as the carbon source has a purity of ~99.5% (Sigma Aldrich). Fe nanoparticles were used as catalyst to grow the CNTs. The Fe nano particles were synthesized from analytical-grade ferric nitrate, Fe(NO3)3·9H2O (Sigma Aldrich). A quartz boat loaded with oil was kept in a lower-temperature zone while the catalyst particles in another boat at a highertemperature zone of a horizontal quartz tube (1m long with an inner diameter of 25 mm) in a dual-zone furnace (300 mm long). The outer part of the quartz tube was attached with a water bubbler. The reaction parameters such as argon (Ar) gas and its flow rate, temperature, and duration of heating, were set per requirement. In a typical experiment, the horizontal quartz tube containing quartz boats was first flushed with Ar gas in order to eliminate air from the tube.

37

Then the gas was allowed to flow with a flow rate of 6 ml/min. The furnace was then switched on to attain the set temperature of 900 °C at a rate of 7 °C /min. When the desired temperature was reached, the furnace was left on for a set duration of 3 hours and then allowed to cool down to room temperature under the Ar gas flow. After cooling, the carbon materials were taken out and powdered, then weighed to calculate the yield.

Figure 1. A self explanatory schematic diagram of pyrolysing unit used for preparation of carbon materials from turpentine oil. Synthesis of Catalyst . In a typical method of synthesis, the solutions of ferric nitrate, Fe(NO3)3.9H2O and urea were prepared separately in deionized water keeping the molar ratio at 1:10. Then the two solutions were mixed in a beaker and stirred with a magnetic stirrer at room temperature until a homogeneous solution was obtained. The mixed solution was then transferred into a round-bottomed flask, and the flask was sealed and heated to a temperature of 400oC for 1.5 hours. Upon completion of the reaction, the product was washed with deionized water till it was neutral, to remove the possible absorbed ions and chemicals in order to inhibit agglomeration. The resulted precipitate, in the form of metal hydroxide, was dried and calcined in at 400oC for 1 hour to obtain Fe2O3 nanoparticles. The synthesized Fe2O3 nano particles were then reduced in a H2 gas environment at 7000C for 2 hours in a CVD apparatus. The schematic of the synthesis is represented as the following:

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38

Synthesis and Characterization Of Carbon Nanotubes Using a Natural Precursor: Turpentine Oil

O

Fe(NO3)3.9H2O

4000 C

+

Metal : Urea =1:10

H2N

Fe2O3 nanoparticles

NH2

Step 1. Synthesis of Fe2O3 nano particles Fe2O3 nanoparticles

7000 C H2 gas

Fe nanoparticles

Step 2. Reduction of Fe2O3 nano particles

distinction between the catalyzed and uncatalyzed carbon formation is in the uncatalysed reaction which exhibited the formation of CNT along with amorphous carbon, whereas the reactions using Fe as catalyst formed less amorphous carbon as almost pure nano tubes are formed (Figures 5(a) and 5(b)). SEM Studies. The SEM micrograph of the carbon material derived from turpentine oil using Fe nanoparticles as catalyst was shown in the Figure 2. The image shows the possible existence of carbon nanotubes.

Characterization of the CNTs. Powder X-ray diffraction (XRD) measurements were carried out on a Bruker AXS D8-Advance powder X-ray diffractometer using the Cu-Kα radiation (λ=1.5418Å) with a scan speed 2°/min. Transmission electron microscopy images were obtained on a JEOL, 9JSM100CX transmission electron microscope (TEM) with an accelerating voltage of 100kV. The sample powders were dispersed in ethanol, under sonication and TEM grids were prepared using a few of drops the dispersion followed by drying in air. Scanning electron microscopy (SEM) images Figure 2. SEM micrographs of carbon were obtained on a JEOL, JSM scanning obtained from catalyzed reaction. electron microscope (SEM). 3. Results and Discussion The yield of the carbon materials was found to be in the range of 30-33%. The densities of the carbon synthesized from turpentine oil with or without the catalyst were in the range of 0.21 to 0.26. The results show that the density of the synthesized materials is much lower than that of the graphitic carbon and amorphous carbon [17]. The morphology and internal structure of the synthesized carbon materials were investigated through the SEM and TEM studies. Turpentine oil on pyrolysis at 9000C in presence of Fe nanoparticles as catalyst produced tubes of 12-15 nm in diameters while the unanalyzed reaction yielded tubes of 20-30 nm in diameter. The important

TEM Studies. The TEM micrographs of the CNTs synthesized by catalytic CVD of turpentine oil using of (Æ”using” or “by use of”) Fe nanoparticles as nucleating agent are presented in Figures 3(a) and 3(b). The diameters of the tubes were recorded in the range of 12-15 nm. The TEM images of the carbon obtained from unanalyzed reactions also indicate the presence of branched CNTs (Fig.4a and 4b) having diameters in the range of 20-30 nm. The TEM micrographs given in Figures 5(a) and 5(b) indicate the presence of of Fe nano particles. The particles are less dispersed and show agglomeration with dimension was in the range 30-35 nm.

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C.R. Bhattacharje et al., Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 36-42

(a)

39

(b)

Figure 3. TEM micrographs of carbon from catalysed reaction of turpentine oil (a) scale 500 nm (b) scale 50 nm.

(a)

(b)

Figure 4. TEM micrographs of carbon from uncatalysed reaction (a) scale 100 nm (b) scale 71.43 nm.

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40

Synthesis and Characterization Of Carbon Nanotubes Using a Natural Precursor: Turpentine Oil

(b)

(a)

20

40

60

80

40

(103)

(101)

(100) 100

2 th eta (d eg re es)

(102)

Intensity(a.u)

(110)

(004)

(101)

(100)

Intensity(a.u)

(002)

(002)

Figure 5. TEM images of Fe particles (a) scale 200 nm (b) scale 100 nm.

60

80

100

2 th e ta (d e g r e e s)

(a)

(b)

Figure 6. XRD pattern of carbon (a) in presence of Fe nano particle as catalyst (b) without catalyst.

XRD Studies. The X-Ray diffraction (XRD) pattern of the material obtained from catalysed and uncatalysed reactions are presented in Figures 6(a) and 6(b), respectively. The XRD patterns of the samples reveal the characteristics of the graphitized carbon [17-19]. The (002) graphitic lines of the samples were observed at 2θ = 25.8° corresponding to an inter-planar spacing of about 0.343 nm which is usually attributed to CNTs [19]. The patterns also

indicate high degree of crystalinity in case of the catalyzed reaction, which suggests low content of amorphous carbon and impurities by using catalysts of Fe nanoparticles. 4. Conclusions Carbon nanotubes were obtained by pyrolysis of turpentine oil, an unconventional natural precursor, as a carbon source. The diameters of the CNTs from catalysed reactions was

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C.R. Bhattacharje et al., Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 36-42

about 12-15 nm while those from the uncatalysed reactions were about 20-30nm. This research is still ongoing and the preliminary results show that it is promising. The ecologically advantageous and regenerative material may be an effective precursor for the growth of CNTs. This method can be scaled up for mass production.

41

Acknowledgements The financial support provided by the UGC (NERO) is much appreciated. The technical supports given by IITG and SAIF, NEHU are also acknowledged. We are also thankful to the authorities of SICES Ambernath.

References [1] Dresselhaus, M. S. (1992). Down the straight and narrow. Nature, 358, 195 [2] Saito, R., Fujita, M. Dresselhaus, G., & Dresselhaus, M.S. (1992). Electronic structure of chiral graphene tubules. Appl. Phys. Lett. 60, 2204-6. [3] Kumar, M., Kakamu, K., Okazaki, T. & Ando, Y. (2004). Field emission from camphor-pyrolyzed carbon nanotubes. Chem. Phys. Lett., 385, 161-5. [4] Sawatsuk, T., Chindaduany, A., Saekung, C., Prafontep, S. & Tumcharern, G. (2009). Dye - sensitized solar cells based on TiO2 - MWCNTs composite electrodes: Performance improvement and their mechanisms. Diam. Relat. Mater., 18, 524-7. [5] Maiyalagan, T., Viswanathan, B., & Varadaraju, U. V. (2005). Nitrogen containing carbon nanotubes as support for Pt – Alternate anodes for fuel cell applications. Electrochem. Commun., 7, 905-12. [6] Wang, S. G., Qing Zhang, R., Yoon, S. F., Ahm, J., Yang, G. J., Tian, J. Z., Li, J. Q., & Zhou, Q. (2003). Multi- Walled carbon nanotubes for the immobilization of enzyme in glucose biosensors. Electrochem. Commun., 5, 800-3. [7] Tan, T. T., Sim, H.S., Lau, S.P., Yang, H.Y., Tanemura, M., Tanaka, J., et al. (2006). X- Ray generation using carbonnanofiber-based flexible field emitters. Appl. Phys. Lett., 88, 103105-3. [8] Karthikeyan, S. & Mahalingam, P., (2010). Studies of Yield and Nature of Multi - Walled Carbon Nanotubes Synthesized by Spray Pyrolysis of Pine Oil at Different Temperature. Int. J. Nanotech. Appl.4, (3), 189-7 [9] Journet, C., Maser, W. K., Bernier, P. Loiseau, A., de la Chapelle, M.

L., Lefrant, S., Deniard, P., Lee, R., & Fisher, J. E. (1997). Large - scale production of single - walled carbon nanotubes by the electric-arc technique. Nature, 388, 756-8 [10] Guo, T., Nikoleav, P., Thess, A., Colbert, D.T., & Smalley, R. E. (1995). Catalytic growth of single - walled nanotubes by laser vaporization. Chem. Phys. Lett., 243, 49-4 [11] Cassell, A. M., Franklin, N. R., Tombler, T.W., Chan, E. M., Han, J. & Dai, H., (1999). Directed Growth of Free- StandingSingle - Walled Carbon Nanotubes. J. Am. Chem. Soc. 121, 7975 -6. [12] Pan, Z. W., Xie, S. S., Chang, B. H., Wang, C. Y., Lu, L., Wu, W., Zhou, W. Y., Li, W. Z., & Qian, L. X., (1998).Very long carbon nanotubes. Nature, 394, 631-2. [13] Fan, S. S., Chapline, M. G., Franklin, N. R., Tombler, T. W. Cassell, A. M. Regular & Dai, H., (1999). Self-oriented arrays of carbon nanotubes and their field emission properties. Science, 283, 512- 4. [14] Afre, R.A. Soga, T. Jimbo, T. Kumar, M. Ando, Y. & Sharon, M., (2005). Growth of vertically aligned carbon nanotubes on silicon and quartz substrate by spray pyrolysis of a natural precursor: Turpentine oil. Chem. Phys. Lett., 414, 6-0. [15] Tfre, R. A. Soga, T. Jimbo, T. Kumar, M. Ando, Y. Sharon, M. Somani, P. R. & Umeno, M., (2006). Carbon Nanotubes by spray pyrolysis of turpentine oil at different temperatures and their studies. Microporous Mesoporous Mater., 96, 184-0.

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42 [16]

Synthesis and Characterization Of Carbon Nanotubes Using a Natural Precursor: Turpentine Oil

Ghosh, P. Soga, T. Afre, R. A. & Jmbo, T., (2008). Simplified synthesis of single-walled carbon nanotubes from a botanical hydrocarbon: Turpentine oil. J. Alloys Compd., 462, 289-3. [17] Zhang, D. Shi, L. Fang, J. & Dai, K., (2005). Preparation and modification of carbon nanotubes Materials Letters, 59, 4044-7. [18] Kong, Q. & Zhang, J., (2007). Synthesis

of straight and helical carbon nanotubes from catalytic pyrolysis of polyethylene Polymer Degradation and Stability, 92(11), 2005- 0. [19] Afolabi, S. Abdulkareem, A. S. & Iyuke, S. E., (2007). Synthesis of carbon nanotubes and nanoballs by swirled floating catalyst chemical vapour deposition method. J. Exp. Nanosci., 2(4), 269-7.

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SCIENCE JOURNAL Ubon Ratchathani University

Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 43-52

http://scjubu.sci.ubu.ac.th

Research Article

Neutron Interaction Cross-Sections for 27Al in the Energy Range from 0.2 to 22 MeV Using Optical Model Program A.K.M.R. Rahman* Department of Physics, Chittagong University, Chittagong 4331, Bangladesh.

Received 06/09/11; Accepted 28/11/11

Abstract Neutron total and differential elastic scattering cross-sections for 27Al nucleus were calculated from different global spherical optical potential (SOP) sets for different neutron energies ranging from 0.2 MeV to 22 MeV using the well-known computer program SCAT-2 along with an IBM-PC-AT. In addition, the angular distributions of elastically scattered neutrons at different neutron energies were calculated. The results were compared with those of the experimental data obtained from the EXFOR data file of NEA data bank. The best-fit potential parameters were thereby selected. It was observed that the best fit to the experimental values of the total cross-sections are obtained by SOP parameters of Fu. Ketrick for 27Al. Furthermore, the variations of the total cross-sections as a function of real potential depth V0 and real radius parameter r0 were calculated to observe the sensitivity of these parameters towards the cross-sections. Keywords: Optical model potential, SOP parameter, Interaction cross section, Angular distribution.

1. Introduction The interactions of nucleons with the nuclei can be interpreted by different theoretical models. But no individual model or formula can explain the nuclear interaction crosssections for the entire energy range. However, the average total neutron crosssections can be interpreted by using the optical model, first proposed by Bethe [1] *Corresponding author. E-mail address: [email protected]

and then modified by many investigators [2-5]. They have shown that the total and elastic scattering cross-sections can be well fitted by the optical model potential with suitably adjusted parameters. Optical potentials are widely used in the DistortedWave-Born-Approximation(DWBA) analysis and in supplying transmission coefficient for Hauser-Feshbach statistical theory. The most important task is to determine the optical model parameters [6]. A lot of empirical information exists on its parameterizations [7]. Some investigations have been devoted

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44

Neutron Interaction Cross-Sections for 27Al in the Energy Range from 0.2 to 22 MeV

to its energy dependence [8-11]. A very well established procedure is derivation of the optical model from a realistic nucleonnucleon interaction using Brueckner theory [1,5,12-14]. There are two approaches to optical model, the microscopic approach and the phenomenological approach. Among these two approaches, the optical model has long been known to provide an excellent phenomenological description of nucleonnucleus elastic scattering [2,15-21] for medium and heavy mass nuclei.

Here, f (r,Rx,ax) = [1+exp((r-Rx)/ax)]-1 is the Woods-Saxon form factor. 'x' indicates R(for real) and I' (for imaginary). g(r,RI,aI) = exp[(r-RI)/(aI)][1+exp((r-RI)/aI]-2 is the derivative Woods-Saxon form factor with RI = rI A1/3. f(r,RI',aI') = exp[-{(r- RI)/aI}2] is the Gaussian form factor with RI = rI A1/3.

h(r,Rso,aso) = 1/r(d/dr)f(r,Rso,aso)= (1/raso) exp[(r-Rso)/aso][1+exp((r-Rso)/aso)]-2 is the The present work contains a study of Thomas form factor with Rso = rso A1/3 neutrons interactions with 27Al in the energy range 0.2 MeV to 22 MeV. Hence phenomenological approach is deployed. The 3. Materials and Methods neutron total cross-section σ t , shape elastic In our present calculations, eight sets of scattering cross-section σsc, and compound parameters have been used. These are obtained from the literature. A discussion on nucleus cross-section σc for 27Al were each of them is being made here. These calculated for different SOP sets. Then the global spherical optical parameterizations are total cross-sections σ t were compared with presented in Table 1. But some of them are the experimental data obtained from the used with small change of several sets of in SCAT-2 computer references [22, 23]. The angular distributions parameterizations 27 program for 27 Al. The numerical values of the of Al for different energies were calculated parameters of the optical potential can be and are compared with the experimental data obtained approximately by elementary supplied by IAEA. Variations of σ t and σ c arguments. Since the potential is the as a function of real potential depth V0 , real extension to the positive energies of the radius rR were also calculated to observe the potential of the simple shell model and its parameter sensitivity. depth can be calculated from the Fermi gas model, using the relation between the depth and the radius. The depth of the imaginary 2. Theory part of the potential can be estimated from O. Bersillon [2] used the general form of the semi-classical arguments and knowledge of the nuclear matter distribution gives r = optical potential as: 1.2fm and a = 0.65fm. Finally, the spin-orbit U(r) = V (r) - V f(r,R ,a )-i[-4W g(r,R ,a )+ splitting Vso ≈ 8MeV .

Gs G Wvf(r,RI',aI')]+CsoVsoh(r,Rso,aso)( l ⋅ S ) c

r

R

R

I

I

(1)

where the terms represent the coulomb potential, the real volume potential, imaginary surface potential, imaginary volume potential and real spin orbit potential, respectively. This form of the optical potential is used in SCAT-2 [FORTRAN] computer program in our research works.

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45

A.K.M.R. Rahman., Sci. J. UBU, Vol. 2, No. 1 (January–June, 2011) 43-52

Table 1. Global spherical optical parameterizations. Parameter set

Potential

r1

Set 2,

VR = 47.01- 0.2657E - 0.0018E2

Wilmore,

WD = 9.52 - 0.53E

Hodgson, 1964

Vso = 7.0

× 10 × 10 1.322-7.6 × 10

E ≤ 15MeV ; A>40 Set 4,

η = 13.0 - 0.25E - 12 η

VR = 56.3 - 0.32E - 24

Becchetti, WD

Greenlees, 1969

Wv = 0.22E - 1.56

E40 Set 7,

Vg = 71.0

1.182+1.93

Madland,

WD = 7 + 0.4E

1.21

1978,

Vso = 7

Actinide

β = 0.85

E15

1.295

0.59

E ≤ 15

1.295

0.59

E >15

1.295

0.59

1.01

0.75

Wv = - 4.3 + 0.38E Vso = 6.2 Set 10, Fu.

VR = 49.747- 0.429E - 0.0003E2

1.287

0.56

Ketrick

VR = 11.8 - 0.21E

1.345

0.47

Vso = 6.2

1.12

0.47

Set 12,

VR = 45.45 - 0.22E

1.265

0.65

Bersillon et al.

WD = 2.28 + 0.47E

E < 10

1.235

0.5

209

Bi

WD = 6.9 - 0.45E

E > 10

1.235

0.5

-5

E > 10

Fe

10 eV < E
40-fold specificity when patients with Doxorubicin (75 mg/m2 by an evaluated against an antigen negative cell line intravenous bolus every 3 weeks) showed and non-binding control antibody-drug that CAELYX® was significant less conjugations. Both drug-linkers gave myelosuppressive, only 3 (6%) patients had conjugates that showed >70-fold more potent grad 3 and 4 neutropenia, versus 33 (77%) on Doxorubicin. Therefore, than Doxorubicin when evaluated on the patients CAELYX® has equivalent activity to basis of drug concentration. Doxorubicin in soft-tissue sarcoma with an improved toxicity profile [16]. However, the pharmacokinetics of pegylated liposomal 3

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64

Doxorubicin and Thousand Efforts to Get the Better Drugs

Doxorubicin differs from conventional Doxorubicin as pegylation protects liposomes from uptake by phagocytic cells. Pegylated liposomal Doxorubicin has a considerably slower clearance from the circulation (0.04 l/h/m2) and a prolonged β half-life (55 h) compared to conventional Doxorubicin. This results lead to the development of nonpegylated liposomal drug such as MyocetTM (Cephalon) which is Doxorubicin encapsulated in a non-pegylated liposomal membrane of phosphatidylcholine and cholesterol. It is approved in combination with cyclophos-phamide as a first-line treatment of metastatic breast cancer, in Europe and Canada. The pharmacokinetics of non-pegylated liposomal Doxorubicin (NPLD) differs from both conventional Doxorubicin and pegylated liposomal Doxorubicin [17]. Plasma levels of total Doxorubicin are substantially higher with NPLD than its conventional Doxorubicin, while the peak plasma levels of free Doxorubicin are lower with NPLD. The clearance (5.1 ± 4.8 l/h) is much slower than conventional Doxorubicin (46.7 ± 9.6 l/h), but not as slow as pegylated liposomal Doxorubicin [18]. Recently, Doxorubicin derivative that could be given locally and concentrated to the draining lymphatic basin of the breast was developed. The conjugation of Doxorubicin and a lymphatic carrier such as hyaluronan (HA) have been investigated [19]. The results showed that HA–DOX (13) by subcutaneous injection weekly would have great benefit over standard or metronomic dosing regimens both in terms of patient compliance 6. Conclusion Doxorubicin was discovered in the mid of the 1950s. It is among the most effective drugs since then. However, the problems of cardiotoxicity and drug resistance lead to the investigation for more potent and less toxic derivatives of Doxorubicin. Many synthetic derivatives and prodrugs have been

and tolerance, but also with regard to potential decreased toxicity and improved efficacy. Further translational efforts will focus on optimizing dose frequency and completing preclinical proofs of this concept. OH NH2

O

OH

O

OH

OH

O

O O

O N H

HN O O HO

NH HO O HO O OH

OCH3

N

OH O NHAc

n

13: HA-ADH-DOX

Engineered drug delivery systems for cancer treatment aim to increase the efficacy of chemotherapeutic agents while minimizing the interactions with healthy sites in the body by modifying their bio-distribution and controlling the rate at which the agent is released. The production of nanoparticles constituting of the grafted biodegradable and biocompatible copolymer P(MePEGCA-coHDCA) and incorporating Doxorubicin was investigated. The results show that the use of confined impinging jets mixer (CIJM) can help in properly controlling the operating parameters and the operating conditions, in turn improving the final nanoparticle properties, proving the great potentials of this promising technique that allows for continuous production and is easily scalable for release. Further investigation of these systems is currently under study with particular attention to the design of more effective drug release tests and to the use of nanocapsules as an alternative to nanoparticles [20]. synthesized. But, until now Doxorubicin is still the standard drug for cancer chemotherapy. On the other hand, many groups have focused on the development of drug delivery system in order to send Doxorubicin to a specific tumor site and reduce the effects on normal cells. Furthermore, the biomolecular technology can be a useful tool to make drugs more specific and less toxic to the normal cells.

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N. Pongprom, Sci. J. UBU, Vol. 2, No. 1 (January –June, 2011) 60-65

65

References [1] Di Marco, A., Gaetani, M., Orezzi, P., et al. (1964). “Daunomycin,” a new antibiotic of the rhodomycin group. Nature, 201, 706-707. [2] Dubost, M., Ganter, P., Maral, R., et al. (1964). Rubidomycin: A new antibiotic with cytostatic properties. Cancer Chemother. Rep., 41, 35-36. [3] Arcamone, F., Cassinelli, G., Fantini, G., et al. (1969). Adriamycin, 14-hydroxyldaunomycin, a new antitumor antibiotic from S. peucetius var. caesius. Biotechnol. Bioeng. 11, 1101-10. [4] Weiss, R. B. (1992). The anthracyclines: Will we ever find a better Doxorubicin? Seminars in Oncology, 19(6), 670-86. [5] Jain, K. K., Casper, E. S., Geller, N. L., et al. (1985). A prospective randomized comparison of epiruicin and doxorubicin in patient with advance breast cancer. J. Clin. Oncol., 3, 818-26. [6] Maligress, P. E., Nicolaou, K. C. & Wrasidlo, W. (1993). A new designed tumor selective daunomycin derivative. Bioorg. Med. Chem. Lett., 3, 1051-4. [7] Izawa, T. & Kato K. (1995). Design and synthesis of an antitumor prodrug released by the reaction with sulfhydryl compounds. Bioorg. Med. Chem. Lett., 5, 593-6. [8] Olsufyeva, E. N., Tevyashova, A. N., Trestchalin, I. D., Preobrazhenskaya, M. N., Platt, D. & Klyosovb, A. (2003). Synthesis and antitumor activity of new D-galactosecontaining derivatives of doxorubicin, Carbohydrate Research, 338, 1359-67. [9] Bhupender, S., St. Jean, C. N., Mandal, D., Kumar, A. & Parang, K. (2011). Fatty acyl amide derivatives of doxorubicin: Synthesis and in vitro anticancer activities, Eur. J. Med. Chem., 46, 2037-42. [10] Niculescu-Davaz, I. & Springer, C. J. (1997). Antibody-directed enzyme prodrug therapy (ADEPT): a review. Adv. Drug Del. Rev., 26, 151-72. [11] Florent, J-C., Dong, X., Gandel, G. et al. (1998). Prodrugs of anthracyclines for use in antibody-directed enzyme prodrug therapy. J. Med. Chem., 41, 3572-81. [12] Svensson, H. P., Vrudhula, V. M., Emswiler, J. E. et al. (1995). In Vitro and In Vivo Activities of a Doxorubicin Prodrug in Combination with Monoclonal Antibody βLactamase Conjugates. Cancer Res. 55, 2357-65.

[13] Helen, J., Rossa, H. J., Hartb, L. L., et al. (2006). A randomized, multicenter study to determine the safety and efficacy of the immunoconjugate SGN-15 plus docetaxel for the treatment of non-small cell lung carcinoma, Lung Cancer, 54, 69-77. [14] Jeffrey, S. C., Nguyen, M. T., Andreyka J. B., Meyer, D. L., Doronina, S.O. & Senter, P. D. (2006). Dipeptide-based highly potent doxorubicin antibody conjugates, Bioorg., Med. Chem. Lett., 16, 358-62. [15] Swenson, C. E., Perkins, W. R., Roberts, P. & Janoff, A. S. (2001). Liposome technology and the development of Myocet(TM) (liposomal doxorubicin citrate). The Breast, 10(2), 1-7. [16] Judson, I., Radford, J. A., Harris, M. et al. (2001) Randomised phase II trial of ® pegylated liposomal doxorubicin (DOXIL / ® CAELYX ) versus doxorubicin in the treatment of advanced or metastatic soft tissue sarcoma: a study by the EORTC Soft Tissue and Bone Sarcoma Group. Eur. J. Cancer, 37, 870-7. [17] Batist, G., Ramakrishnan, G., Rao, C. S. et al. (2001). Reduced cardiotoxicity and preserved antitumor efficacy of liposome-encapsulated doxorubicin and cyclophosphamide compared with conventional doxorubicin and cyclophosphamide in a randomized, multicenter trial of metastatic breast cancer. J. Clin. Oncol., 19(5), 1444-54. [18] Leonard, R. C. F., Williams, S., Tulpule, A., Levine, A. M. & Oliveros, S. (2009) Improving the therapeutic index of anthracycline chemotherapy: Focus on liposomal doxorubicin (MyocetTM), The Breast, 18, 218-24. [19] Cai, Sh., Thati, Sh., Bagby, T. R., et al. (2010) Localized doxorubicin chemotherapy with a biopolymeric nanocarrier improves survival and reduces toxicity in xenografts of human breast cancer, J. Controlled Release, 146, 212-8. [20] Lincea, F., Bolognesi, S., Stellab, B., Marchisioa, D. L. & Dosio, F. (2011). Preparation of polymer nanoparticles loaded with doxorubicin for controlled drug delivery. Chem. Eng. Res. Design, 89, 2410-9.

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SCIENCE JOURNAL Ubon Ratchathani University

Sci. J. UBU, Vol. 2, No. 1 (January-June, 2011) 66-71

http://scjubu.sci.ubu.ac.th

Research Article

Natural Rubber bound 4-Aminodiphenylamine Antioxidant for Rubber Formulation Improvement P. Klinpituksa1,3*, A. Hamad2,3,C. Nakason2,3 , A. Kaesaman2,3 1

Department of Science, 2 Department of Rubber Technology and Polymer Science, 3 Center of Excellence in Natural Rubber Technology, Faculty of Science and Technology, Prince of Songkla University, Pattani 94000, Thailand. Received 02/02/10; Accepted 01/02/11

Abstract This research work was aimed to prepare the natural rubber bound 4-aminodiphenylamine (NRbound-4-ADPA) for replacing typical antioxidant used in the conventional rubber formulation. Epoxidized natural rubber with 30 %mol epoxide (ENR-30) was first prepared by performic epoxidation using high ammonia concentrated latex (60% DRC), Teric N-10 stabilizer, HCOOH/H2O2 at 50 oC for 6 hr and 15 min. Reaction of ENR-30 with 4-aminodiphenylamine (4-ADPA) was then carried out in toluene solution at 80 oC for 3, 6, 12 and 12 hrs. The content of 4-ADPA bound to natural rubber molecules was determined by using standard curve of ENR-30/4-ADPA physical mixing method. It was found that the level of bound 4-ADPA was 1.0, 1.48, 2.61 and 3.95 phr, respectively. The molecular characteristics of ENR and NR-bound-4-ADPA were also investigated by means of IR spectroscopy. The absorption bands at 870 cm-1 and 1596 cm-1 corresponding to epoxide group and secondary amine in NR-bound-4-ADPA were observed, respectively. The antioxidant capability of NR-bound-4-ADPA and conventional 4-ADPA antioxidant in natural rubber vulcanizates was then studied. Improvement of ageing resistance of NR-bound-4-ADPA was observed. That is the NR-bound-4-ADPA showed superior oxidation resistance than that of the conventional antioxidant. Keywords: Natural rubber, Natural rubber bound antioxidant, Ageing resistance.

conventional antioxidant. For instance, macromolecular antioxidant by reaction of There have been many attempts to improve 4-anilinoaniline with epoxidized polyisopreageing properties of rubbers by chemical ne [1], epoxidized natural rubber latex with modification rubbers/polymers with a aromatic amines [2], natural rubber-bound p-phenylenediamine antioxidant [3], chlorinated paraffin wax-bound p-phenylene*Corresponding author. diamine antioxidant [4], and oligomer-bound E-mail address: [email protected] antioxidants [5]. 1. Introduction

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P. Klinpituksa et al., Sci. J. UBU, Vol. 2, No. 1 (January-June, 2011) 66-71

The main objective of this work was to prepare natural rubber bound to 4aminodiphenylamine (4-ADPA) via epoxidized natural rubber. The natural rubber bound 4-aminodiphenylamine (NR-bound-4ADPA) was then formulated with NR in order to investigate the ageing properties compared with the application of typical antioxidant in the same recipe. 2. Theory Oxidative degradation of organic materials is a natural and continuous processes. It occurs in varying degrees at various rates depending upon the environmental stresses placed upon material. Natural and synthetic polymers are particulary susceptible to degradative changes resulting from interaction with molecular oxygen. These changes can manifest themselves as drastics swings in tensile strength, elongation, hardness, tack and discoloration. Oxidation can be slowed by chain-breaking antioxidants, which prevent initial formation of free radicals. Antioxidations work by deactivating the sites by decomposing the hydroperoxide or by terminating of the free radical reaction. In rubber processing, it generally needs to add an antioxidant to protect oxidation from oxygen in the atmosphere. Disadvantages of using a typical antioxidant include toxicity, leaching and volatility. The efficiency of polymer bound antioxidant is expected to be higher than that of organic low molecular antioxidants because of their lower volatility and of their better compatibility with polymer. Polymer bound aromatic antioxidants containing amino group are of additional interest because of less toxicity problems [1]. 3. Materials and Methods Materials. Epoxidized natural rubber (ENR) was first prepared using high ammonia concentrated latex (60% DRC) manufactured by Yala latex Industry Co., Ltd, Yala Thailand. Hydrogen peroxide and formic acid were supplied by Ried-de Haen

67

Germany. Teric N-10 was purchased from Huntsman, Austria. 4-aminodiphenylamine (4-ADPA) was supplied by Fluka chemical Switzerland Ltd. Compounding ingredients, such as zinc oxide, stearic acid were supplied by Imperial Chemical Co., Ltd., Thailand. N-tert-Butyl-2-benzothiazole sulfonamide (TBBS) was commercial grade supplied by Flexsys America L.P., U.S.A. Toluene and acetone were used to purified the natural rubber were manufactured by Lab- Scan Ltd, Ireland. Methods. Preparation of Epoxidized Natural Rubber. Epoxidized natural rubber with 30%mol (ENR-30) was first prepared by epoxidation reaction of high ammonium concentrated natural latex (60% DRC) with HCOOH/H2O2, Teric N-10 stabilizer at 50 oC for 6 hr and 15 min as previously described elsewhere [6]. FTIR spectroscopy and 1HNMR spectroscopy were then used to characterize the ENR-30 [7]. Preparation of NR-Bound-4-ADPA. A solution of 5 g ENR-30 in 100 ml of toluene in a 250 mL round bottom flask equipped with mechanical stirrer was immersed in water bath at 80 ◦C. A solution of 5 g phenol and 5 g 4-ADPA in 50 mL of toluene were then added. The reaction mixture was kept at 80 o C for 3, 6, 12 and 24 hr. The NR-bound-4ADPA was obtained by precipitation in acetone, and dried in hot oven at 50oC. The product was then purified by dissolve in toluene, reprecipitated in acetone and dried. Determination of 4-ADPA Content. NRbound-4-ADPA obtained at various reaction times of experiments was characterized by means of FT-IR spectroscopy (Omnic ESP Magna-IR 560 spectrophotometer, Nicolet). Samples were dissolved in CHCl3, casted on KBr cell and evaporated CHCl3 out with aired dryer. The samples were tested after completion of the blank spectrum scanning. The FT-IR equipment was operated with a resolution of 4 cm-1 and scanning range from 4000 to 400 cm-1. The absorbance ratios at 1596 cm-1 and 1375 cm-1 corresponding to -

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68

Natural Rubber Bound 4-Aminodiphenylamine Antioxidant for Rubber Formulation

NH group of 4-ADPA and -CH3 group of natural rubber (cis-1,4-polyisoprene unit) was determined. The amounts of 4-ADPA bound to natural rubber molecule were determined from the standard curve which was constructed by physical mixing method of ENR-30/4-ADPA [8].

Table 1. Formulation of NR-bound-4-ADPA vulcanizates. Ingredients NR-bound-4-ADPA ZnO Stearic acid TBBS Sulfur

Quantities (phr) 100 6.00 0.50 0.70 3.5

Table 2. Formulations of NR and ENR-30 vulcanizates with the conventional antioxidant (4-ADPA). Ingredients NR (ENR-30) ZnO Stearic acid TBBS 4-ADPA Sulfur

Quantities (phr) 100 6.00 0.50 0.70 1.00 3.50

Transmittance (Arbitary scale)

ADPA obtained at various reaction times are given in Figure 1. The spectra of NR-bound4-ADPA samples clearly show new absorption bands at 1596 cm-1 corresponding to the -NH bending vibration of 4-ADPA and the intensities of this peak increased with increasing 4-ADPA content in NR-bound-4ADPA. In our work, we are interested in -1 -1 Ageing Resistance NR-Bound-4-ADPA. The absorbance ratio of 1596 cm and 1376 cm antioxidant capability of NR-bound-4-ADPA (-CH3 deformation of natural rubber constituent) in order to monitor the progress compared with NR and ENR-30 using a of reaction at various reaction times. typical conventional 4-ADPA antioxidant in rubber vulcanizates was formulated as shown in Tables 1 and 2. 3 hr

6 hr

12 hr

24 hr

1596 1376

3900

3400

2900

Characterization of NR-bound-4ADPA. FTIR spectra of NR-bound-4-

1400

900

400

Figure 1. FTIR spectra of NR-bound-4ADPA at various reaction times. A possible reaction for preparation of NRbound-4-ADPA from natural rubber is illustrated in Figure 2. n Natural Rubber (NR) HCOOH/H2O2 50oC, 6.15 hr O

m

Epoxidised Natural Rubber (ENR) N

NH2

H

4-Aminodiphenylamine (4-ADPA) Phenol 80oC, 24 hr HN

O

n-m

4. Result and Discussion

1900

Wavenumber(cm-1)

n-m

Samples were moulded in an electrically heated hydraulic pressed at 150 oC. Dumbbell shape pieces were punched out of these compression mould along the mill grain direction. The tensile properties of vulcanizates were tested using a Tensile Testing Machine [9].

2400

m-p

OH

NH

p

Natural Rubber bound 4-Aminodiphenylamine (NR-bound 4-ADPA)

Figure 2. Possible reaction of preparation of NR-bound-4-ADPA from NR and 4-ADPA.

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69

P. Klinpituksa et al., Sci. J. UBU, Vol. 2, No. 1 (January-June, 2011) 66-71

To determine the amount of 4-ADPA bound 4-ADPA containing secondary amine and to natural rubber molecule, the NR-bound-4- residual epoxide groups which caused ADPA was calculated from corresponding crystallization. standard curve of variation of absorbance ratio (A1596/A1376) with various quantities 100 of 4-ADPA (Figure 3). 90 80 Weight (%)

0.35

Absorbance ratio

0.3 0.25 0.2

NR

60 ENR-30

50

NR- bound- 4-ADPA

40

-

30

y = 0.0585x R2 = 0.9938

0.15

70

20

0.1

10

0.05

0

200

400

600

800

0 0

1

2

3

4

5

O

Temperature ( C)

Quantities of 4-ADPA (phr)

Figure 4. TGA thermograms of NR, ENRFigure 3. Standard curve of variation of 30 and NR- bound 4-ADPA (heating rate absorbance ratios with quantities of 4-ADPA. 10oC/min under O2 atmosphere).

35

Control Tensile Strength (MPa)

The amounts of 4-ADPA bound to natural rubber in NR-bound-4-ADPA product were found to be 1.00, 1.48, 2.61 and 3.95 phr at 3, 6, 12 and 24 hr, respectively. The NR-bound4-ADPA with 1 phr of 4-ADPA was then used for further ageing resistance investigation.

30

Aging

25 20 15

10 Thermal Property of NR-Bound-4-ADPA. 5 The decomposition temperature of NR, ENR0 30 and NR-bound-4-ADPA determining by NR ENR-30 NR-bound-4-ADPA TG analysis were 372, 398 and 405oC, Type of Rubber respectively. The TGA thermogram of samples was shown in Figure 4. That is, NRbound-4-ADPA exhibited higher thermal Figure 5. Tensile strength of NR, ENR-30 resistance than those of ENR-30 and NR. and NR-bound-4-ADPA vulcanizates before This is due to incorporation of secondary and after ageing. amine antioxidant in natural rubber molecule. Figure 6 demonstrates elongation at break of Ageing Property of NR Vulcanizates. Tensile NR, ENR-30 and NR-bound-4-ADPA strength of NR, ENR-30 and NR-bound-4- vulcanizates before and after ageing. The ADPA vulcanizates are shown in Figure 4. It elongation at break of NR- bound-4-ADPA can be seen that the highest tensile strength vulcanizates was lower compared with NR was observed in NR-bound-4-ADPA vucani- and ENR-30 vulcanizates. The lower zates. However, all cases of rubber elongation at break of NR-bound-4-ADPA vulcanizates, their tensile strength are vulcanizates is attributed to higher polarity decreased after ageing. The higher tensile which caused weakening and susceptible strength of NR-bound-4-ADPA vucanizates breakdown. might be due to higher polarity of NR-bound-

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

70

Natural Rubber Bound 4-Aminodiphenylamine Antioxidant for Rubber Formulation

E lo n g atio n at b reak(% )

800

Control

700

Aging

600 500 400 300 200 100 0 NR

ENR-30 Type of Rubber

NR-bound-4-ADPA

changes of tensile strength and elongation at break of NR-bound-4-ADPA vulcanizates were found to be lower than that of the ENR30 and NR vulcanizates, respectively, whereas the modulus change was higher. It was due to the influence of 4-ADPA leading to intermolecular crosslinking formation which caused the lower change in tensile strength and elongation at break. This might be attributed to intermolecular crosslinking change of modulus.

Change (% )

Figure 6. Elongation at break of NR, ENR- 5. Conclusions 30 and NR-bound-4-ADPA vulcanizates NR-bound-4-ADPA was successfully prepabefore and after ageing. red by reaction of ENR-30 with 4-ADPA in the presence of phenol as a catalyst in toluene at 80 oC. The NR-bound-4-ADPA with 1 phr 20 of 4-ADPA was used as antioxidant in rubber 10 formulations compared with the conventional TS EB 4-ADPA antioxidant in natural rubber 0 100% 200% 300% vulcanizates. The NR-bound-4-ADPA vulca-10 Modulus Modulus Modulus nizates showed an improved ageing -20 resistance in compared with vulcanizates -30 containing a conventional antioxidant. It was NR ENR-30 also found that the NR-bound-4-ADPA -40 NR-bound-4-ADPA showed superior oxidation resistance than -50 those the conventional antioxidant in NR and ENR-30. Figure 7. Changes of modulus, tensile strength and elongation at break of NR, ENRAcknowledgements 30 and NR-bound-4-ADPA vulcanizates after ageing. The authors thank the kind support of the Graduate School and Research Fund, Prince Changes in properties of 100%. 200% of Songkla University, Pattani Campus, and 300% modulus, tensile strength and Thailand. elongation at break of rubber vulcanizates are illustrated in Figure 7. It was found that

References [1]

[2]

Jayawardena, S., Reyx, D., Durand, D. & Pinazzi, C.P. (1984). Synthesis of macromolecular antioxidants by reac- [3] tion of aromatic amines with epoxidized polyisoprene, 3a Reaction of 4-anilinoaniline with epoxidized 1,4-polyisoprene. Macromol. Chem. 185, 2089-97. [4] Perera, M. C. S. (1990). Reaction of aromatic amines with epoxidized

natural rubber latex. J. Appl. Polym. Sci. 39(3), 749-58. Avirah, S. A., & Joseph, R. (1995). Studies on natural rubber bound paraphenylene diamine antioxidants in NBR. J. Appl. Polym. Sci. 57(12), 1511-24. Sulekha, P. B., Joseph, R., & Prathapan, S. J. (2001). Synthesis and characteriza-

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.

P. Klinpituksa et al., Sci. J. UBU, Vol. 2, No. 1 (January-June, 2011) 66-71

tion of chlorinated paraffin wax-bound para-phenylene diamine antioxidant and its application in natural rubber. [7] J. Appl. Polym. Sci., 81(9), 2183-9. [5] Sulekha, P. B., Joseph, R., & Manjooran, K. B. (2004). New oligomer-bound antioxidants in natural rubber/polybutadiene rubber and natural rubber/styrenebutadiene blends. J. Appl. Polym. Sci. [8] 93(1), 437-43. [6] Nakason, C., Sainumsai, W., Kaesaman, A., & Klinpituksa, P. (2001). Cure and physical properties of natural rubber and epoxidized natural rubber [9] compounds using various types of

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accelerators. Songklanakarin J. Sci. Technol. 23(3), 415-24. Burfield, D. R., Lim, K. -L., Law, K.S., & Ng, S. (1984). Analysis of epoxidized natural rubber : A comparative study of d.s.c., elementary analysis and direct titration methods. Polymer. 25, 995-8. Barra, G. M. O., Crespo, J. S., Bertolino, J. (1999). Maleic anhydride grafting on EPDM: Qualitative and quantitative determination. Br. Polymer J., 10, 31-4. Annual Book of ASTM D412-98a. (2000). Rubber, Natural and Synthetic General Test Method, Carbon Black.

Copyright © 2011 Faculty of Science, Ubon Ratchathani University. All Rights Reserved.