CHAPTER 2 REVIEW OF LITERATURE

CHAPTER 2 REVIEW OF LITERATURE CHAPTER 2 – REVIEW OF LITERATURE Stevia rebaudiana Bertoni, a native plant of Paraguay, is non-caloric, natural, med...
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CHAPTER 2

REVIEW OF LITERATURE

CHAPTER 2 – REVIEW OF LITERATURE Stevia rebaudiana Bertoni, a native plant of Paraguay, is non-caloric, natural, medicinal and economical plant. Steviosides, a white crystalline compound isolated from Stevia is 250-300 times sweeter than table sugar. Stevia rebaudiana is used for the treatment of various conditions such as cancer (Yasukawa et al., 2002), diabetes (Lailerd et al., 2004), obesity, cavities, hypertension (Dyrskog et al., 2005), fatigue, depression and yeast infection. It possess hypoglycemic, hypotensive, vasodilating, taste improving, sweetening, antifungal, antiviral, anti-inflammatory, antibacterial properties and increases urination function of the body. It has been found to be nontoxic, non-addicitive, non-carcinogenic, non-mutagenic and is devoid of genotoxic effect. It does not affect blood sugar level hence safe for diabetes (Strauss, 1995).

Researchers in Denmark published a study, which demonstrates the in vitro hypoglycemic action of steviosides and steviol to stimulate insulin secretion via a direct action of beta cells (Jeppensen et al., 2000). Oviedo (1971) reported a 35.2% fall in the blood sugar level, 6-8 hours following the ingestion of Stevia leaf extract. These kinds of results have led physicians in Paraguay to prescribe Stevia leaf tea in the treatments of diabetes (Soejarto et al., 1983). Similarly, in Brazil, Stevia tea and Stevia capsules are officially approved for sale for the treatments of diabetes (Kinghorn and Soejarto, 1985). In fact, herbalists in Brazil have been recommended Stevia to regulate blood sugar levels for at least 40 years (Richard, 1996).

The plant is used to treat high blood pressure; there is hypotensive effect on both systolic and diastolic blood pressure, which is dose-dependent (Chan et al., 1998). A good deal of experimental work has been done on the effects of Stevia and steviosides on cardiovascular functioning in man and animals. It is

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always a slight lowering in arterial blood pressure at low or normal doses, changing to slight rise in arterial pressure at very high doses (Machado et al., 1986). Researches clearly shows that Streptococcus mutans, Pseudomonas aeruginos, Proteus vulgaris and other microbes do not thrive in the presence of the non-nutritive Stevia constituents. This fact, combined with the naturally sweet flavour of the herb, makes it a suitable ingredient for mouthwash and for the toothpastes.

Stevia extract shows anticandidal and vasodilator properties. In Brazil, Stevia has been used as a digestive aid that help positively influences the health of the pancreas (Gates, 1996). In 1968, Planas and Kuc claimed that certain tribes of Indians in Paraguay used Stevia as a contraceptive.

There is health as well as political controversy about the plant Stevia and its extract stevioside. In 1985, a study conducted by Pezzuto et al., reported that steviol (a breakdown product of stevioside and rebaudioside) is a mutagen in the presence of a liver extract of pretreated rats. But Procinska et al., 1991 criticized that the data was mishandled in such a way that even distilled water would appear mutagen. In 1991 at the request of anonymous complaint, the United States Food and Drug Administration (FDA) labeled Stevia as an “unsafe food additive” and restrict its import. The FDA’s stated reason was “toxicological information on Stevia is inadequate to demonstrate its safety”. The FDA requires proof of safety before recognizing a food additive as safe. A similar burden of proof is required for the FDA to ban a substance label it unsafe. Nevertheless, Stevia remained banned until Dietry Supplement Health and Education Act forced the FDA in 1995 to revise its stance to permit Stevia to be used as a dietry supplement, although not as a food additive (McCaleb, 1997).

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Genus Stevia (Synonyms: Eupatorium rebaudianum) belongs to family Asteraceae includes about 150 species; the important species are Stevia eupatoria, S. ovata, S. plummerae, S. rebaudiana, S. salicifolia and S. serrata.

VARIETY There are 3 varieties developed by Sun fruits Pvt. Ltd. Pune viz

SRB 123: It can be harvested 5 times in a year. The glycosides content is in between 9-12%, found suitable for South India. SRB 512: It is suitable for Northern India and glycosides content is in between 9-12%. SRB 128: It is suitable for Northern India with higher stevioside content up to 21%.

HISTORY For centuries, the Guarani tribes of Paraguay and Brazil used Stevia species, primarily S. rebaudiana which they called Ka`he`e^ - “Sweet herb”, as a sweetener in yerba mate and medicinal teas for treating heat burn and other aliments. The leaves of Stevia plant have 30-45 times the sweetness of sucrose i.e. ordinary table sugar.

The Swiss botanist Moises Santiago Bertoni first described the plant and the sweet taste in detail (Bertoni, 1899) and named the plant in honor of Paraguayan chemist Dr. Rebaudi. Two French chemists isolated the glycosides that give Stevia its sweet taste (Bridel and Lavielle, 1931). The compounds were named steviosides and rebaudiosides and are 250-300 times sweet than sucrose (Brandle and Jim, 2004).

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In the early 1970`s, Japan began cultivating Stevia as an alternative to artificial sweetener such as cyclamate and saccharin, which are suspected to be carcinogen. The plant’s leaves, the aqueous extract of the leaves, and purified stevioside are used as sweetener. The Japanese firm Morita Kagaku Kogyo Co. Ltd produced the first commercial Stevia sweetener in Japan in 1971. Stevia is grown commercially in many parts of Brazil, Japan, Korea, Taiwan, Isreal, Thailand, China and South East Asian countries. Stevia products are used commercially extensively in Japan, using locally and imported dried leaves, where they make up over 40% of non-sucrose sweetener and 5-6% of total sweetener market. China is the main producer of Stevia products.

PLANT DESCRIPTION Stevia rebaudiana Bertoni, commonly called as Sweet leaf, Sweet herb, honey leaf, sweet herb of Paraguay is a small, shrubby, herbaceous, perennial plant growing up to 1 m in cultivation (Brandle and Starratt, 1998) but plant height up to 1.8m and 20 branches per plant on more fertile soils had been observed. Stevia grows naturally on infertile, sandy soils with shallow water tables (Lester, 1999).

The natural climate is semi humid and subtropical with temperature extremes from 21-420C, averaging 240C with an annual rainfall of 1375 mm (Brandle and Starratt, 1998).

Stevia is a member of family Asteraceae. The plant has an extensive tap root system with brittle stems producing sessile, oppositely branched, lanceolate to oblanceolate leaves (Robinson, 1930). The leaves are bright green in colour, having distinct characteristics odour and sweet taste. The leaves are cuneate in shape with glabrous surface (Soni et al., 2008). Stevia is a short

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LIST

OF

COUNTRIES

WHERE

STEVIA

IS

GROWN

AND

RESEARCHED Country/ Location South AmericaParaguay Uruguay/Brazil

Commercial Research production ++ ++

Non-Agric Research ++

Approved for use ++

++

++

++

Central America

+

Mexico

+

++

+ +

+

USA

+

++

Canada

+

+

+

China

++

++

++

+++

Vietnam

+

++

+

+

Taiwan

+

++

+

++

Japan

+

++

++

+++

South Korea

+

++

++

++

Thailand

+

+

+

+

Malaysia

+

+

+

Indonesia

+

India

+

Georgia

+

+

+ +

+

Russia

+

++

++

Ukraine/Moldova

+

+

+

Spain

+

+

Italy

+

+

United Kingdom

+

+

Germany

+

+

Sweden

+

+

Commercial production excludes small quantities grown for domestic use.

Redrawn form - A new rural industry- Stevia- to replace imported chemical sweeteners (Midmore and Andrew, 2002).

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Figure 1:Stevia rebaudiana leaves Figure 2: Stevia rebaudiana flower

Figure 3:Stevia rebaudiana stem

Figure 4: Stevia rebaudiana plant

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day plant with a critical day length of 13 h (Zaidan et al., 1980) and remain vegetative in the spring through early summer and flower in late summer (Shock, 1982). Long day conditions increase internode length, leaf area and stevioside content (Metivier and Viana, 1979) whereas shade can reduce total growth, delay flowering and reduces the rate of flowering (Slamet and Tahardi, 1988). The plant can initiate flowering after a minimum of four true leaves have been produced (Carneiro, 1990). Flowering occurs at 54-104 days depending upon the day length sensitivity of the cultivars (Lester, 1999). The flowers are small (7-15mm), white and arranged in irregular cyme (Robinson, 1930). The amount of selfing between complete diallel cross with 8 parents ranged between 0-0.5%, while out crossing ranged between 0.768.7%, indicating that some self-incompatibility system is operating (Katayama et al., 1976).

Oddone (1997) also considers Stevia to be self-incompatible and insect pollinated. The seed is an achene with a feathery pappus (Brandle and Starratt, 1998). Seed are about 3 mm in length (Goettemoeller and Ching, 1999) and very light in weight. 1000 seeds weight usually ranges between 0.15-0.30 g (Carneiro, 1990). Seed viability and yield are affected by growing condition during pollination and filling. Extensive rainfall during pollination can affect both seed yield and viability (Shuping and Shizhen, 1995). Shock (1982), Duke (1993) and Carneiro et al., (1997) all mention poor production of viable seeds.

Stevia flowers need to be fertilized by pollen of another plant to produce viable seeds (Oddone, 1997). Fertile seeds are usually dark coloured whereas infertile seeds are usually pale (Felippe, 1978).

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PROPAGATION Stevia plant can be propagated by seed or by vegetative methods. The crop could be transplanted in February or March and seed collected in late summer (Katayama et al., 1976). Life span of the crop is reported to 7-8 years and herb yield increases up to 4 years (Columbus, 1997).

Tamura et al., 1984 showed no significant difference in growth or in glycosides content in plants propagated by seed, by tissue culture methods or by vegetative methods.

SEED PROPAGATION Establishment of plants from seed is generally unsuccessful as seed germination is very poor and less than 50% (Miyazaki and Watanabe, 1974). Seed germinate within 7-10 days and seedling growth is very slow. Germination requires at least 200C and often more than 250C. Light generally increase germination (Tanaka, 1985). Timing of flowering, seed harvest and pollination methods play an important role, rain at flowering can also reduce seed setting. Shade can reduce total growth, delay flowering and reduce the rate of flowering (Slamet and Tahardi, 1988).

VEGETATIVE PROPAGATION As establishment of plants from seed is difficult, several researchers studied alternative propagation method of the plant as vegetative propagation by stem cuttings (Chalapathi et al., 2001), in vitro propagation (Uddin et al., 2006), clonal propagation (Tamura et al., 1984) etc. Cuttings of new stems and shoots can be propagated successfully (Lee et al., 1979) and rooting of cuttings can be stimulated (but not always), by use of growth regulators (Bondarev et al., 1998). The size (numbers of leaves) of cuttings and day length can influence rooting and growth (Zubenko et al., 1991). Cuttings

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taken in late winter rooted better than those taken at other times (Carvalho et al., 1995).

The location on the plant from which cuttings are taken can also affect growth and rooting, cuttings from the top half and with four internodes performing best (Tirtoboma, 1988). For vegetative propagation in Stevia, length of cutting also influences. According to Chalapathi et al., 2001 use of 15 cm cutting gave higher sprout percentage than 7.5 cm cutting.

For tissue cultured propagation many different parts of the plant can be used successfully-leaves, auxillary shoots, root neck sprouts, shoot primodia, internodal explants (Akita, 1994). Usually propagation is done by stem cuttings which root easily.

Stevia has been successfully taken to a wide range of climatic locations around the world and apparently grown successful, although often by using vegetative propagation methods and seedling establishment in a green house before planting in the field.

CHEMICAL COMPOSITION A number of plant chemicals have been investigated from Stevia. Stevia come in attention due to its natural, non-caloric sweetener commonly called as glycosides. The two main glycosides are stevioside, 5-10% of the dry weight of the leaves and rebaudioside A (R-A), 2-4 % of the dry weight of the leaves. There are also related compounds including rebaudioside (1-2%) and dulcosides A and C, as well as minor glycosides, including flavonoid glycosides, coumarins, cinnamic acid, phenyl propanoids and some essential oils (Dzyuba, 1998). In different plant parts natural sweetener is present in highest amount in the leaves (Bondarev et al., 1998) and the major sweetener in leaves is steviosides. Yield of sweetening compound in leaf tissue can vary

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according to method of propagation, day length (Metivier and Viana, 1979) and agronomic practice (Shock, 1982).

The structure and biosynthetic pathways for glycosides has been study by many worker (Strauss, 1995; Brandle and Starratt, 1998). C Nuclear Magnetic Resonance (C-NMR) spectrometry has been used to determine the chemical structure of glycosides (Tanaka, 1985). The biosynthesis is linked to the biosynthesis of gibberellin growth regulators; Stevia thus may be a potential source of these compounds. Hershenhorn et al., 1997 reported plant growth regulator derived from stevioside. Biosynthesis of sweet compound takes place in green tissue (chloroplasts) and so they are in the leaves and green tips of the plants. As stem matures and lose colour any steviosides present dissipate (Shaffert and Chebotar, 1994).

The sweetening effect of these compounds is purely by taste; they are undigested and the body absorbed no part of the chemical. They are therefore of no nutritional value (Hutapea et al., 1997).

There are suggestions that the stems and the flowers could contain some of the compounds, which may give Stevia some of its other properties, especially flavour enhancers (flavanoids), odour enhancers and organoleptic substances (Darise, 1983). Enzyme modification of stevioside produces a flavonoid rebaudoside (Ikan, 1993).

Over 100 phytochemicals have been discovered in Stevia since. It is rich in terpenes and flavonoids. The main plant chemicals in Stevia include: apigenin, austroinulin, avicularin, beta-sitosterol, caffeic acid, campesterol, caryophyllene, centaureidin, chlorogenic acid, chlorophyll, cosnosiin, cynaroside, daucosterol, diterpene glycosides, dulcosides A-B, foeniculin, formic acid, gibberellic acid, gibberellin, indole-3-acetonitrile, isoquercitrin,

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isosteviol, jhanol, kaempterol, kaurene, lupeol, luteolin, polystachoside, quercetin, quercitrin, scopoletin, sterebin A-H, steviol, steviolbioside, steviolmonoside, stigmesterol, umbelliferone and xanthophylls (Taylor, 2005).

Figure 5: Central structure of steviosides and related compounds.

CULTURAL PRACTICES FERTILIZERS Fertilizer requirement are moderate for Stevia, partially due to its adapation to poor quality soils. Fertilizer trials show yield reduction at high rates of fertilizers (Goenadi, 1985). Stevia plant consist of 1.4% N, 8.3% P and 2.4% K (Katayama et al., 1976). The significant response in terms of growth and yield was up to 40:20:30 Kg NPK/ha and further increase in fertilizer dose to 60:30:45 Kg NPK/ha caused marginal increase in growth and yield attributes. The dry leaf yield was higher with 40:20:30 Kg NPK/ha. The uptake of N, P and K increases with increase in levels of fertilizers (Chalapathi et al., 1999).

PLANT DENSITIES Most cultural systems involve transplanted seedling and so densities less than the optimum is usually recommended. Densities of 80-100,000 plants per

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Compound name

R1

1. Steviol

H

R2 H -Glc- -Glc

(2 -

-Glc- -Glc

-

-Glc- -Glc

(2 (2 | -Glc (3 -Glc- -Glc (2

-

| -Glc (3 -Glc- -Rha

(2 (Dulcoside B) 7.

9. Rebaudioside F (2

10. Dulcoside A (2

-Glc- -Glc (2

-Glc- -Glc (2 -

-

| -Glc (3 -Glc- -Glc (2 | -Glc (3 -Glc- -Glc (2 -Glc- -Xyl | -Glc (3 -Glc- -Rha

Figure 6: Structure of stevioside and related compounds. In -Glc- -Glc (2->1). In rebaudioside A, B, C, D, E, and F group R2 an additional sugar Glc is substitute for by- -Xyl. (Redrawn from J.M.C. Genus, (2003))

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hectare on row spacing of 45-60 cm are generally recommended (Donalisio et al., 1982).

IRRIGATION Supplementary irrigation is generally assumed to be essential to avoid any water stress on plants unless the growing area has reliable rainfall throughout most of the year. Spray irrigation has been suggested (Columbus, 1997) but this would encourage leaf disease and reduce seed set. Frequent irrigation is required to maintain soil moisture above plant wilting, which suggested being as high as 80% of the field capacity (Oddone, 1997). Any moisture stress can reduce leaf production (Goenadi, 1983).

DISEASES Two fungal disease, Septoria steviae and Sclerotinia sclerotiorum, have been reported in Stevia grown in Canada (Lovering and Reeleder, 1996; Chang et al., 1997). Depressed, angular, shiny olive gray lesions, sometimes surrounded by a chlorotic halo, characterized Septoria disease. Scleotinia disease was characterized by brown lesions on the stem, near the soil line, followed by wilting and eventually by the complete collapse of affected individuals. No means of controlling this disease have yet been published. Since Stevia is very slow to established and does not compete well with weeds, herbicides or other means, will be essential to control weed growth to produce ample yield and a clean crop. The herbicide trifluralin appears to be well tolerated by Stevia (Katayama et al., 1976).

HARVESTING Time of harvesting depends on land type, variety and growing season. In colder climates, only one harvest per planting is possible. Under these circumstances the harvesting should be done when the yield is greatest and it

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can be at the onset of cold whether (Columbus, 1997), early harvesting will reduce total yield (Shuping and Shizhen, 1995). Harvesting should be done before flowering at flower bud appearance as the stevioside content of the leaves fall when flowering commences (Bian, 1981). Cutting the whole plant 5-10 cm above the ground effects harvesting. In warmer climates, where some plant growth is possible for most of the year, more than one harvest per year is normal. In Paraguay/Brazil three harvest a year are normal (Donalisio et al., 1982). In India four or five harvest a year are possible and in Indonesia up to seven harvests have been possible per year (Basuki and Sumaryono, 1990).

BIOCHEMICAL ASPECTS OF DISTILLERY SPENT WASH The quality of distillery spent wash fluctuates significantly as it depends on various processes that takes place within the distillery and the organic components in the spent wash. World bank reported that untreated distillery spent wash typically contained suspended solids, high biological and chemical oxygen demand, nitrogen, potassium and phosphorus. However, all the organic materials are not dissolved in the spent wash hence some organic material remains as particulate. If land with suitable topography, soil characteristics and drainage is available, distillery spent wash can be put to good use as both source of irrigation water and plant nutrients. In most of the areas, water scarcity has forced the farmers to use this spent wash as a substitute of irrigation water (Mittal and Tawari, 2008).

Irrigation with distillery wastewater seems to be an attractive agriculture practice, which not only augment crops yield but also provides a plausible solution for the land disposable of the spent wash. The spent wash contained N, P, K, Ca, Mg and S and thus valued as a fertilizer when applied to soil through irrigation with water (Samuel, 1986). Application of distillery spent

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wash should be done after proper dilution (1:10 to 1:50) with irrigation water or by pre-plant application (40-60 days before planting) (Baskar et al., 2003).

Kaushik and Khan, (2008) called spent wash as liquid gold for agriculture because of its valuable macro and micronutrients. The spent wash contains an excess of various form of cations and anions, which are injurious to plant growth and these constituents should be reduced to beneficial level by diluting spent wash, which can be used as a substitute for chemical fertilizer (Sahai et al., 1983). The diluted spent wash irrigation improved the physical and chemical properties of the soil and further increased soil microflora (Kaushik et al., 2005). The distillery spent wash contained all necessary elements and bio fertilizer microbes (Rhizobia, Azospirilla, Azotobacter and Phosphobacteria) to support the growth of plants (Babu et al., 1996).

Because of high concentration of organic load, distillery spent wash is a potential source of renewable energy. Due to high organic contents, the wastewater can be subjected for the production of biogas, biocomposting and potash recovery (Singh, 2008).

The distillery spent wash purely of plant origin, contains a number of plant nutrient and other valuable elements, which can be used in agricultural practice for irrigation purpose after proper dilution. Disposable of the spenyt wash is often done by discharge it into water bodies, which causes various environmental problems. The utilization of distillery spent wash in agriculture would save cost on fertilizer, better crop productivity and facilitate reduction in pollution load on aquatic system. The spent wash contained an excess of various forms of cations and anions, which are injurious to plant growth and these constituents should be reduced to beneficial level by diluting the spent wash, which can be used as a substitute for chemical fertilizers (Selldurai et al., 2010).

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Chatterejee et al., 1976 described the recovery of potash fertilizers from distillery spent wash. Paul, 1976 studied the conversion of alcohol distillery waste into fertilizers and cattle feed. From the analysis of spent wash, the potash content were estimated approximately 14% of solids present in the spent wash.

Datar and Bhargava, 1984 studied the effects of temperature on BOD, COD and kinetics during aerobic digestion. The results of the laboratory studies on aerobic batch digestion of activated sludge over a wide range of digestion temperature (40-500C) showed that the digestion period of 9 to 12 days was found to be sufficient to achieve acceptable results from the consideration of reduction in biological and chemical oxygen demand. Nandan et al., 1987 studied the microbial conversion of distillery waste to bio energy and noted changes in COD values of distillery waste during microbial methanogenesis.

Patil et al., 1984 pointed out the rate of decomposition of spent wash and farmyard manure (FYM) in soil under laboratory conditions. The total quantity of CO2 evolved from spent wash amended soil was nearly 3 times that from FYM treatment. The rate of decomposition of spent wash and FYM followed first order kinetics during initial period of incubation. Bhardwaj, 1985 concluded that recycling of crop residues promotes nitrogen economy.

GROWTH ASPECTS

SEED GERMINATION Seed germination is an important event for the establishment of plant. In Stevia seed germination rate is very low. In wild conditions the plant is raised through seed but in cultivation the propagation of plant through seed is unsuccessful due to poor viability of seed. Goettemoeller and Ching (1999) have studied seed germination conditions. 23

A laboratory work was undertaken to assess the effect of different concentration of distillery spent wash like 0%, 25%, 50%, 75% and 100% on seed germination, speed of germination, peak value and germination value of three selected seeds i.e. wheat (Triticum aestivum), pea (Pisum sativum) and lady’s finger (Abelmoschus esculentus). Germination percentage decreases with increasing concentration of spent wash in all the treated seeds, where as the germination speed, peak value and germination value increases from control to 25% and 50% concentration and decreases from 50% to 75% and 75% to 100% spent wash (Pandey et al., 2007). The effect of different concentrations of distillery spent wash on germination of three multi-useful tree species: Acacia catechu, Dalbergia sissoo and Morus alba has been studied by Pandey and Sony, 1994. Low spent wash concentration enhanced the germination of all species. Higher spent wash concentrations (>10%) however inhibited germination percentage.

Also a number of researches have studied effect of distillery spent wash on seed germination of a number of plants such as sunflower (Tomer et al., 2002), blackgram (Misra and Pandey, 2002), bengal gram (Pandey and Neraliya, 2002), pigeon pea (Karande and Ghanvat, 1994), okra (Hari et al., 1994), filed crops (Singh and Raj, 1995), sugarbeet (Sharma et al., 2002), vegetables crops (Ramana et al., 2002), rice (Singh et al., 1985, Suresha and Puttaiah, 2006), cotton, pulses, millets and small millets (Sindhu et al., 2007) and green gram (Santiago and Bolan, 2006). All the above researches showed improvement in seed germination percentage at low concentration of distillery spent wash.

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SEEDLING GROWTH Establishment of plant is greatly effected by the seedling growth. For Stevia rebaudiana Bertoni two month old seedling of 5-7 leaf stage and 8-10 cm. height was suitable for transplant in the field (Megeji et al., 2005). Effect of different concentration of distillery spent wash on seedling growth of plants have been observed by a number of workers such as by Singh et al., 1985 (Rice), Pandey and Neraliya, 2002 (Bengal gram), Karande and Ghanvat, 1994 (Pigeon pea), Sharma et al., 2002 (Sugarbeet) and Suresha and Puttaiah, 2006 (Paddy).

BIOMASS After proper establishment of seedling, there is accumulation of organic matter by photosynthesis. Standing crop data of an ecosystem is an index of the ecological efficiency and flow of energy. A number of researchers have been studied about the effect of spent wash on biomass and productivity of different plants.

Chidankumar et al., 2009 cultivated some top vegetables (Creepers) such as bottle gourd, ash gourd, pumpkin, snake gourd, ridge gourd and bitter gourd and was made by irrigation with distillery spent wash of different proportions. The primary treated distillery spent wash, 50% and 33% spent wash were analyzed for additive plant nutrients such as N, P, K and other physical and chemical parameters. Experimental soil was tested for its chemical and physical parameters. The vegetables seeds were sowed in the prepared land and irrigated with raw water, 50% and 33% spent wash. It was found that the yield of top vegetables were high in 33% spent wash than 50% spent wash and raw water irrigation.

Tripathi et al., 2007 show that dilutions of distillery spent wash with irrigation water significantly affected the grain yield as well as biomass

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production in rice and wheat. The pre-sown application of diluted distillery spent wash at the rate 20 m3/ha was optimum for building up soil fertility and increasing the yield of rice and wheat.

A pot experiment was conducted to study the effect of irrigation with tube well water (S0) as well as with spent wash diluted for 25 (S25), 50 (S50) and 100 (S100) times on yield of and nutrient uptake by sugarcane grown. The results revealed that application of spent wash diluted at any level increased the sugarcane yield and nutrient uptake significantly over S0. Less diluted spent wash gave higher yield of biomass than the concentrated one (Zalawadia et al., 1997).

Kaushik and Khan, 2008 used distillery spent spent wash as ferti-irrigation i.e. application of distillery spent wash through irrigation water after proper dilution to Saccharum officinarum (COS 94257). Dilution levels of 1:3 and 1:4 shows significant results of GRI, cane girth, number of clump, milliable cane, Kg brix, pole in cane, purity coefficient and yield of sugarcane. Treated soil revealed that the available potassium, nitrogen and phosphorus increased. A number of researches have shown their results to demonstrate the effect of distillery spent wash on plant growth and crop productivity as in rice and wheat (Tripathi et al., 2007), groundnut (Ramana et al., 2001), C3 and C4 plants (Babu et al., 1996), and sugarcane (Singh et al., 2007; Zalawadia et al., 1997, Diangan et al., 2008). Diluted spent wash increases the growth of shoot length, leaf number per plant, leaf area and chlorophyll content of pea (Rani and Srivastava, 1990).

Yield of a number of plants have been reported to increase by the application of diluted or low concentration of spent wash. Such report have been stated by Chandraju and Basavaraju 2007 (leaves vegetables), Chandra et al., 2004 (Phaseolus aureus), Salawadia and Raman, 1994 (sorghum), Tripathi et al.,

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2007 (rice and wheat), Baskar et al., 2003 (soyabean, sugarbeet, potatoes, vegetables, forage crops, tree crops), Sukanya and Meli, 2004 (maize), Babu et al., 1996 (C3 and C4 plants), Pathak et al., 1999 (wheat and rice), Avitha et al., 2008 (sunflower), Ramana et al., 2001, Singhandhupe et al., 2009 (groundnut), Sindhu et al., 2007 (cotton, pulses, millets and small millets), Subramani et al., 1995 (Vigna radiata), Hari et al., 1994 (okra) etc.

GROWTH ANALYSIS Growth analysis is a method to follow the dynamics of photosynthesis production measured by the production of dry matter. Growths being define as net result of metabolism in plants. The primary values are assessed in growing plant material at certain time intervals. Growth analysis can also be used to investigate ecological phenomena such as the success of a species, genetic differences in yielding capacity and effects of agricultural treatment on crop growth.

Stefanini and Rodrigues, 2003 studied growth parameters of Stevia rebaudiana Bertoni such as NAR, RGR, and LAGR under the effect of gibberellic acid. Variability of the results does not allow to define a pattern of the influence of the GA3 on the RGR. Whereas Chengguo et al., 2009 studied the net photosynthetic rate of Stevia rebaudiana in autumn.

ENERGETICS Living organism can use radiant and fixed form of energy for their metabolic activities. Former is the kinetic energy in the form of electromagnetic waves, whereas the later is the potential chemical energy bound up in various organic substances which can be broken down or reacted with something else in order to release their energy content. Autotrophs fix energy from carbon containing inorganic sources into organic molecules. Biomass can be expressed in terms of calories so that the relationship between the average

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standing state and the rate of energy flow can be established (Odum, 1963). Odum and Odum, 1955 have extensively studied natural communities such as forests, grasslands and aquatic system for their caloric contents. Phillipson, 1966 complied principles of energy flow in natural and man-made system. Lieth, 1968 has drawn energy flow on global level.

The caloric value of a plant is function of its genetic constitution, nutritive conditions and life history. Aslander, 1958 and Russel, 1961 reported relationship of nutrients with energy content and emphasized that nutrient deficiency reduces the energy trapping efficiency with consequent reduction in productivity.

MINERAL STANDING STATE All plants have certain basic requirements for mineral constituents from the soil for their proper growth. Out of seven major elements, nitrogen, phosphorus and potassium are needed in large amounts and thus form the major components of inorganic fertilizers. In most cases it is relatively easy to appreciate the requirement and utilization of elements by the plant. Nitrogen is a component of proteins and enzymes and many other cell content like chlorophyll, nucleic acid, nucleotides, coenzymes and vitamins. Phosphorus is an essential part of ATP molecule, nucleic acid, phosphorylated sugars and phospholipids. Potassium is often present in large amount but its function is not so readily seen as that of the other two minerals. The potassium concerns in osmo regulation is an essential co-factor for many enzyme systems.

Bradshaw, 1969 recognized that nitrogen deficiency is one of the major factors to limit the yield of agricultural produce. Addition of phosphorus and potassium fertilizer also stimulates growth in plants (Williams, 1946).

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Influence of fertilizers levels on growth, yield and uptake of Stevia was reported by Chalapathi et al., 1999. Das et al., 2008 studied the effect of biofertilizers on Stevia biomass and nutrients contents in the soil. Application of bio-fertilizer increase biomass of the plant. Das et al., 2006 studied effect of N, P and K on stevioside content in Stevia. They observed that application of NPK fertilizers enhances total biomass as well as steviosides content in Stevia plants.

Application of distillery spent wash as fertilizers has been demonstrated by Salawadia and Raman, 1994. A greenhouse trial conducted to study the effect of diluted waste water in sorghum crop indicated that 75% of the recommended fertilizer application with the distillery waste water the yield was at par with 100% recommended dose of fertilizer application with normal water irrigation with 75% fertilizer dose than with normal water irrigation with 100% fertilizer dose. After the crop harvest, the values of organic carbon, available N, P and K in soil were higher with the usage of spent wash water than with normal at the same level of fertilizer application. Marked increase in N, P and K contents of green gram crops was recorded due to spent wash treatment over the control (Santiago and Bolan, 2006).

Response of different concentration of distillery spent wash like 0%, 25%, 50%, 75% and 100% on crop plants i.e. pea and wheat were studied under field conditions. Distillery spent wash was serving as liquid fertilizer up to 50% spent wash concentration; more than 50% spent wash concentration it is posing inhibitory effect on the rested crops (Pandey et al., 2009).

A study of undertaken to understand the impact of distillery spent wash on the phosphorus content in wheat crop (Triticum aestivum Linn CV ‘HD 2285’). The experiment was organized in land in a completely randomized design had 3 replications with 25, 50 and 100% spent wash concentrations.

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Results showed that phosphorus content in leaves, stem, roots, inflorescence and grains of the wheat plant treated by 50% dilution value of spent wash was enhanced (Mittal and Tiwari, 2008). The successful use of distillery spent wash as a liquid fertilizer for augmenting crop productivity in C3 and C4 plants has been demonstrated by Babu et al., 1996.

NUTRIENT UPTAKE A field experiment conducted to observe plant nutrient contents, nutrient uptake and crop yield in sunflower. The study revealed that application of 1.5 times N through spent wash along with 1.5 times P through SSP and 1.5 times N through spent wash along with balance P through SSP gave significant superior yield and revealed higher N, P and K content in seed and stalk. Similarly total uptake is also highest with these treatments (Avitha et al., 2008).

A field investigation was carried out by Sukanya and Meli, 2004 to study the effect of conjuctive use of spent wash and water on maize yield and soil properties. Treatments included six dilution levels. The results indicated that dilution levels of 1:5 and 1:10 were found optimum to realize significantly higher grain yield of maize than other treatment combinations. The nutrient availability viz N, P, K, Zn, Cu, Fe and Mn contents in soil was significantly higher with an irrigation by undiluted spent wash.

Low concentration of distillery spent wash increase the uptake and available nutrients (N, P and K) in plants. Such observations have been reported by a number of researches in many plants such as in leaves vegetables (Chandraju and Basavaraju, 2007), cabbagae and mint leaf (Chandraju et al., 2008), top vegetables, pulses, condiments and root vegetables (Basavaraju and Chandraju, 2008), sunflower (Avitha et al., 2008), cotton, millets and small millets (Sindhu et al., 2007), sugarcane (Zalawadia et al., 1997) etc.

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PROTEIN CONTENT Stevia leaves contained essential amino acids (arginin, lysine, histidine, phenyl alanine, leucine, methionine, valine, therionine, and isoleucine) as well as non-essential amino acids (aspartate, serine, glutamic, proline, glycine, alanine, cystine and tyrosine). Thus Stevia leaves could be used as a good source of essential amino acids, which could be factors to maintain a good health (Abou-Arab et al., 2010).

Tadhani and Subhash, 2006 reported that Stevia rebaudiana is a good source of protein and carbohydrate. The proximal analysis study shows low oil content in the leaves. Potassium, calcium, magnesium, phosphorus, sodium and sulphur nutritionally important were found in reasonable amount in the leaf. A number of researches have studied the effect of spent wash on protein content of a number of plants such as Bengal gram (Pandey and Neraliya, 2002) and Phaseolus aureus (Chandra et al., 2004).

OIL AND TOTALCHLOROPHYLLS In Stevia rebaudiana Bertoni the leaves contains the essential oil. The amount of oil is very low in the leaves and in oil there is a great amount of linolenic acid (Tadhani and Subhash, 2006). Fujita et al., 1977 examined the essential oil of Stevia rebaudiana plants collected in several zones of Japan

caryophyllene, trans- -

-

-cadinene, caryophyllene

oxidase, nerolidol and an unidentified alcohol and the monoterpenes: linalool, terpinen-4-

-terpineol. Martelli et al., 1985 identified 54

components of a steam distillate of dried plant leaves whereas Markovic et al., 2008 report 88 components from the essential oil of Stevia leaf. Cioni et al., 2006 also examined the composition of essential oil of five different Stevia rebaudiana genotypes.

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Chlorophyll is a major component of plants. With the help of chlorophyll plants utilize sun energy and convert it into chemical energy in the form of carbohydrate by the process called as photosynthesis. Therefore, chlorophyll is the key substance to determine the productivity of the crop.

Total chlorophyll content of leaves in Stevia was examined by Abou-Arab et al., 2010 A field experiment with groundnut as last crop was conducted by Ramana et al., 2001 to evaluate the manurial potential of 3 distillery spent wash: raw spent wash (RSW), bio methanated spent wash (BSW) and lagoon sludge (LS) vis-à-vis recommended fertilizers (NPK+ Farm yard manure (FYM)) and a control. It was found that all three distillery spent wash increased total chlorophyll content, crop growth rate (CGR), total dry matter, nutrient uptake (N, P and K) and finally seed yield compared to control. It was concluded that these distillery spent wash potential could supply nutrients, particularly potassium, nitrogen and sulphur to the crops and thus reduce the fertilizer requirement of the crop.

Effect of distillery spent wash on chlorophyll content of plants has been observed by Sharma et al., 2002 in sugar beet, Pandey and Neraliya, 2002 in Bengal gram, Chandra et al., 2004 in Phaseolus aureus and Singh et al., 2007 in sugarcane.

EFFECT

OF

DISTILLERY

SPENT

WASH

ON

SOIL

CHARACTERISTICS Impact of distillery spent wash irrigation on soil microflora of the pots used for growing Phaselous aureus L. was investigated by Chandra et al., 2004. The irrigation of pots by 1-10% distillery spent wash stimulated the growth of soil microflora and growth aspects such as shoot and root length, biomass, chlorophyll and protein content. 15-20% distillery spent wash had toxic

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effect on soil microflora as indicated by reduced number of bacteria, fungi and actinomycetes. Reduction in root, shoot length, biomass, chlorophyll and protein content of P. aureus were also observed when irrigated by 15-20% distillery spent wash. It has been concluded from present study that lower concentrations of the distillery spent wash has stimulated the growth of the plant however higher concentration had toxicity to test parameters.

The experiment conducted by Sindhu et al., 2007 revealed that though at higher doses (>250 m3/ha) spent wash application is found to crop growth and soil fertility, its use at lower doses (125 m3/ha) remarkably improves germination, growth and yield of dry land crops.

Application of spent wash improves the physical characteristics of soil as proved by Singhandhupe et al., 2009. Zalawadia et al., 1997 observe betterment in some physical characteristics of soil as application of spent wash in sugarcane field. Soil pH was not much affected with the application of spent wash as shown by Hati et al., 2005.

Available N, P and K content in soil ware estimated by a number of researchers when the soil was treated with the different concentration of spent wash. The observations indicate high mineral content in soil when treated with low concentration of spent wash. Such observations has been reported by Sindhu et al., 2007, Singhandhupe et al., 2009 and Baskar et al., 2003. Similar reports has been present by Chidankumar and Chandraju, 2008 in post harvest soil of pulses and Kaushik and Khan, 2008 in post harvest soil of sugarcane.

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