NORMAL AND REVERSED PHASE THIN LAYER CHROMATOGRAPHY OF SELECTED 2,5-ANHYDROALDOHEXOSE ETHYLENE ACETAL DERIVATIVES

J. LIQ. CHROM. & REL. TECHNOL., 22(10), 1473–1491 (1999) NORMAL AND REVERSED PHASE THIN LAYER CHROMATOGRAPHY OF SELECTED 2,5-ANHYDROALDOHEXOSE ETHYLE...
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J. LIQ. CHROM. & REL. TECHNOL., 22(10), 1473–1491 (1999)

NORMAL AND REVERSED PHASE THIN LAYER CHROMATOGRAPHY OF SELECTED 2,5-ANHYDROALDOHEXOSE ETHYLENE ACETAL DERIVATIVES S. M. Petrovi,1 D. Popovi,1 M. Popsavin,2 V. Popsavin2 1

University of Novi Sad Faculty of Technology Department of Analytical Chemistry Bul. Cara Lazara 1 21000 Novi Sad, Yugoslavia 2

University of Novi Sad Faculty of Sciences Institute of Chemistry Trg D. Obradovia 3 21000 Novi Sad, Yugoslavia ABSTRACT The chromatographic behavior of 31 samples of variously substituted 2,5-anhydroaldohexose ethylene acetal derivatives has been studied on silica gel and C-18 modified silica gel layers with, respectively, binary non-aqueous and aqueous mobile phases. The slopes and intercepts of the linear relationships between the retention constant (RM) and the logarithm of the volume fraction of the diluent in non-aqueous mobile phase, as well as of the volume fraction of organic component in aqueousorganic mobile phase, have been calculated and are discussed in relation to solute and mobile and stationary phase characteristics. The retention and relative retention of compounds depend largely on the retention behavior of their substituents. 1473 Copyright © 1999 by Marcel Dekker, Inc.

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1474

PETROVIj ET AL. INTRODUCTION

Variously substituted monosaccharide derivatives, frequently used as key intermediates in the synthesis of certain biomolecules,1 are convenient for studying the relationship between the molecular structure of a compound and its chromatographic properties. In a previous paper,2 the normal phase chromatographic behavior of selected 1,2-O-isoprpylidene derivatives of aldohexoses and 1,2-Ocyclohexylidene derivatives of aldopentoses was studied on silica gel thin layers. Significant and distinct effects of the types of compounds and the type, number, and position of substituents in a molecule on retention were observed. In this work, we have studied selected 2,5-anhydroaldohexose ethylene acetal derivatives of D-gluco, D-alo, D-gulo and L-ido series possessing a variety of substituents. The selected compounds have conveniently been used as key intermediates in the synthesis of C-nucleosides,3 as well as of (+)-muscarine and its analogs.4-6 The retention behavior of selected compounds has been studied by normal and reversed phase thin layer chromatography (TLC) using silica gel and C18 modified silica gel layers, and, respectively, non-aqueous and aqueous mobile phases.

EXPERIMENTAL TLC was performed on 10 x 10 cm HPTLC plates pre-coated with silica gel 60 or C18 modified silica gel (Merck, Darmstadt, Germany). The samples were dissolved in chloroform (2 mg mL-1) and 1-µL volumes of the solutions were applied to the chromatoplate with a micropipette. The binary mobile phases in normal phase chromatography were cyclohexane (Cx) or toluene (Tl) mixed with ethyl acetate (EtAc), acetone (An), dioxane (Dx), or tetrahydrofuran (THF) in various proportions of mixture components. In reversed phase chromatography, methanol (MeOH) or acetonitrile (ACN) were mixed in various proportions with water (H2O). Spots were detected by spraying with a 50% aqueous solution of sulfuric acid, followed by heating at 120oC for 10-15 min. Rf values are averages from at least three chromatograms developed for each solute-mobile phase combination. RM values were calculated by use of the formula RM = log(1/Rf - 1). The structures of the compounds are given in Fig. 1.

Figure 1. Structural formulas of the compounds examined.

2,5-ANHYDROALDOHEXOSE ETHYLENE ACETALS 1475

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PETROVIj ET AL.

Figure 2. Plots of RM vs. log ϕ (eqn. 1) (a) and RM vs. log ϕD (eqn. 2) (b) for mobile phase Cx-THF. Designation of solutes is as in Fig. 1.

2,5-ANHYDROALDOHEXOSE ETHYLENE ACETALS

1477

Figure 3. Plot of RMDo vs. nD for mobile phase Cx-Dx.

RESULTS AND DISCUSSION

Normal Phase Chromatography (NPC) The relationship between chemical structures of the compounds (Fig. 1) and their retention behavior in NPC has been studied using the well-known equation7,8 RM = RMo - n log ϕ

(1)

where ϕ denotes the volume fraction of the polar constituent of a binary mobile phase, RMo is an extrapolated RM value in pure polar solvent, and n is a constant. The numerical data of the constants n and RMo for each compound and mobile phase tested are presented in Table 1 for cyclohexane and in Table 2 for toluene as the diluent. The data in Tables 1 and 2 show that slope n values of Eq. 1 mainly follow the polarities of the compounds, i.e., the n value decreases with the decrease of the compound retention.

n 4.70 4.60 4.03 2.99 3.16 2.95 3.50 3.18 4.23 2.52 4.90 2.99 3.24 3.13 3.02 2.47 4.22 2.73 2.48 2.52 4.76 4.68 3.83 3.77

Cpd. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24

-0.538 -0.511 -0.715 -0.899 -1.028 -0.672 -0.816 -0.925 -0.645 -1.122 -0.532 -0.995 -1.309 -1.115 -1.334 -1.217 -0.687 -0.828 -1.163 -1.183 -0.660 -0.617 -0.616 -0.755

Cx-EtAc ϕ = 0.35 - 0.55 RM0 .9963 .9972 .9983 .9991 .9989 .9947 .9986 .9987 .9975 .9946 .9932 .9924 .9984 .9990 .9983 .9979 .9975 .9900 .9975 .9967 .9949 .9931 .9953 .9977

r 3.93 3.67 3.18 2.34 2.46 2.86 3.10 2.49 3.17 1.96 4.07 3.19 2.95 3.08 2.27 2.06 3.75 2.90 2.18 2.09 4.02 3.81 3.29 3.17

n -0.996 -0.997 -1.038 -0.971 -1.029 -0.752 -1.094 -0.989 -0.932 -1.065 -0.964 -1.340 -1.246 -1.276 -1.111 -1.169 -1.049 -1.167 -1.161 -1.084 -1.068 -1.095 -0.798 -0.876

Cx-An ϕ = 0.30 - 0.45 RM0 .9985 .9994 .9996 .9982 .9958 .9999 .9976 .9984 .9993 .9997 .9942 .9976 .9960 .9976 .9991 .9991 .9925 .9935 .9736 .9685 .9999 .9998 .9992 .9992

r 5.26 5.14 4.32 3.67 3.48 3.62 4.18 3.49 3.92 2.07 5.47 3.70 2.72 3.39 3.01 3.24 4.22 3.42 3.18 2.70 5.36 5.10 4.40 4.07

-1.248 -1.245 -1.295 -.1417 -1.413 -1.107 -1.326 -1.366 -1.039 -1.126 1.210 -1.605 -1.120 -1.348 -1.459 -1.666 -1.116 -1.351 -1.615 -1.429 -1.306 -1.264 -1.251 -1.229

Cx-Dx ϕ = 0.30 - 0.45 n RM0 .9968 .9993 .9974 .9910 .9962 .9982 .9983 .9941 .9987 .9990 .9965 .9951 .9992 .9971 .9993 .9959 .9959 .9910 .9932 .9858 .9941 .9932 .9976 .9995

r

Constants n and RM0 of Eq. 1 for Eluents Containing Cyclohexane*

Table 1

4.81 4.60 4.01 3.00 2.85 3.29 3.68 2.95 3.34 2.52 4.85 2.80 2.93 3.02 2.78 2.67 3.48 2.64 2.21 2.23 4.46 4.51 4.15 3.89

n

-1.047 -0.904 -1.085 -1.188 -1.054 -0.717 -1.055 -0.987 -0.667 -1.366 -0.891 -1.187 -1.292 -1.169 -1.323 -1.549 -0.795 -1.064 -1.312 -1.299 -0.985 -0.950 -1.110 -1.096

Cx-THF ϕ = 0.30 - 0.45 RM0

.9993 .9949 .9996 .9984 .9938 .9964 .9996 .9993 .9999 .9984 .9943 .9990 .9999 .9993 .9928 .99785 .9949 .9985 .9874 .9906 .9980 .9960 .9964 .9979

r

1478 PETROVIj ET AL.

25 4.44 -0.612 26 2.66 -0.852 27 2.53 -0.864 28 2.94 -1.049 29 3.88 -1.023 30 2.82 -1.105 31 3.02 -0.889 __________________ * r = correlation coefficient.

.9866 .9990 .9970 .9981 .9992 .9955 .9995

3.90 2.58 2.44 2.80 2.92 2.62 2.93

-0.991 -1.196 -1.056 -1.203 -0.961 -1.206 -1.158

.9987 .9999 .9943 .9866 .9998 .9995 .9971

4.04 2.94 3.08 3.39 3.36 3.46 3.29

-1.043 -1.361 -1.361 -1.397 -1.118 -1.525 -1.362

.9994 .9983 .9965 .9971 .9982 .9995 .9996

3.76 2.54 2.49 2.66 2.92 2.54 2.31

-0.774 -1.112 -1.100 -1.115 -0.775 -1.115 -1.044

.9971 .9966 .9964 .9983 .9991 .9974 .9959

2,5-ANHYDROALDOHEXOSE ETHYLENE ACETALS 1479

n 2.67 2.27 2.24 1.91 1.60 2.49 1.94 1.70 2.60 1.22 2.91 2.01 1.62 1.52 1.72 1.10 2.65 1.70 1.11 1.11 2.77 2.52 2.08 2.08

Cpd. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24

-0.111 -0.205 -0.641 -0.795 -0.897 -0.223 -0.645 -0.867 -0.145 -1.157 -0.065 -0.903 -1.096 -0.931 -1.097 -1.098 -0.377 -0.671 -1.102 -1.195 -0.317 -0.249 -0.599 -0.639

Tl-EtAc ϕ = 0.30 - 0.50 RM0 .9999 .9996 .9991 .9986 .9979 .9992 .9998 .9997 .9992 .9999 .9994 .9951 .9903 .9905 .9980 .9998 .9999 .9969 .9998 .9998 .9922 .9957 .9966 .9981

r 2.83 2.28 1.87 1.46 1.45 1.93 1.49 1.39 2.50 0.92 3.39 1.38 1.40 1.52 1.47 1.02 2.22 1.43 1.02 0.94 2.61 2.50 1.55 1.60

n -0.879 -0.811 -1.096 -1.091 -1.205 -0.689 -1.016 -1.235 -0.698 -1.315 -0.447 -1.053 -1.374 -1.334 -1.346 -1.218 -0.689 -0.999 -1-187 -1.210 -0.832 -1.070 -0.869 -0.985

Tl-An ϕ = 0.25 - 0.45 RM0 .9983 .9957 .9996 .9954 .9995 .9942 .9989 .9947 .9979 .9998 .9938 .9952 .9999 .9982 .9957 .9973 .9994 .9957 .9951 .9951 .9984 .9939 .9980 .9980

r 3.05 2.20 1.69 1.58 1.49 2.40 1.51 1.49 2.80 1.20 4.00 1.61 1.53 1.60 1.38 1.34 2.53 1.80 1.22 1.28 2.72 2.29 2.09 1.73

-0.966 -0.885 -0.973 -1.214 -1.308 -0.910 -0.956 -1.261 -0.850 -1.382 -0.706 -1.232 -1.392 -1.334 -1.270 -1.506 -0.772 -1.154 -1.466 -1.348 -0.975 -1.001 -1.149 -0.992

Tl-Dx ϕ = 0.25 - 0.45 n RM0 .9994 .9997 .9940 .9995 .9964 .9999 .9961 .9944 .9991 .9997 .9985 .9947 .9984 .9993 .9869 .9845 .9868 .9781 .9999 .9915 .9926 .9939 .9999 .9962

r

Constants n and RM0 of Eq. 1 for Eluents Containing Toluene*

Table 2

2.97 2.32 1.81 1.64 1.69 2.42 1.62 1.48 2.62 1.38 3.49 1.70 1.43 1.62 1.43 1.38 2.54 1.93 1.19 1.34 2.57 2.51 1.78 1.67

n

-0.963 -0.843 -0.904 -1.130 -1.260 -0.854 -0.920 -1.098 -0.750 -1.581 -0.589 -1.190 -1.393 -1.297 -1.240 -1.581 -0.723 -1.303 -1.392 -1.568 -0.982 -0.979 -0.996 -0.959

Tl-THF ϕ = 0.25 - 0.45 RM0

.9972 .9999 .9973 .9916 .9929 .9998 .9997 .9995 .9978 .9950 .9989 .9918 .9998 .9999 .9993 .9950 .9999 .9999 .9474 .9918 .9976 .9940 .9921 .9897

r

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25 2.55 -0.002 26 1.51 -0.794 27 1.47 -0.681 28 1.83 -0.938 29 2.32 -0.678 30 1.55 -1.076 31 1.87 -1.125 __________________ * r = correlation coefficient.

.9922 .9920 .9969 .9999 .9999 .9951 .9939

2.56 1.22 1.27 1.24 2.01 1.17 1.66

-1.023 -1.177 -0.978 -0.963 -1.062 -1.133 -1.183

.9992 .9957 .9983 .9941 .9957 .9903 .9997

2.40 1.66 1.52 1.66 1.86 1.57 1.67

-0.850 -1.469 -1.180 -1.266 -0.963 -1.457 -1.116

.9932 .9978 .9960 .9940 .9986 .9994 .9971

2.49 1.83 1.83 1.86 2.10 1.45 1.77

-0.881 -1.293 -1.298 -1.378 -1.087 -1.359 -1.144

.9998 .9969 .9999 .9999 .9999 .9968 .9933

2,5-ANHYDROALDOHEXOSE ETHYLENE ACETALS 1481

-nD 5.83 5.70 4.90 3.66 3.88 4.90 4.35 3.90 5.25 3.13 6.08 3.70 3.98 3.86 3.71 3.06 5.23 3.39 3.08 3.15 5.90 5.80 4.71 4.64

Cpd. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24

2.657 2.613 1.998 1.119 1.110 2.013 1.565 1.227 2.232 0.593 2.800 1.035 0.980 1.026 0.723 0.459 2.179 1.029 0.525 0.534 2.573 2.561 1.975 1.798

Cx-EtAc RMD0 .9979 .9936 .9859 .9948 .9970 .9883 .9985 .9943 .9825 .9896 .9982 .9951 .9940 .9917 .9918 .9967 .9903 .9965 .9965 .9887 .9966 .9932 .9932 .9960

r 8.09 7.53 6.66 4.92 5.19 5.98 6.53 5.24 6.58 4.09 8.45 6.50 6.08 6.36 4.70 4.33 7.67 6.02 4.59 4.41 8.22 7.90 6.91 6.60

-nD 2.335 2.112 1.690 1.040 1.090 1.700 1.573 1.155 1.768 0.616 2.502 1.355 1.260 1.341 0.821 0.603 2.126 1.304 0.709 0.710 2.334 2.148 2.021 1.830

Cx-An RMD0 .9978 .9965 .9982 .9996 .9999 .9955 .9998 .9995 .9920 .9973 .9957 .9860 .9948 .9983 .9955 .9989 .9839 .9995 .9932 .9904 .9919 .9931 .9936 .9842

r 7.82 7.68 6.51 5.11 5.26 5.46 6.30 5.30 5.90 3.77 8.21 5.61 4.95 5.12 4.52 4.43 6.27 5.14 4.31 4.10 7.94 7.65 6.60 6.08

-nD 2.649 2.569 1.925 1.101 1.184 1.589 1.789 1.245 1.880 0.534 2.861 1.162 1.058 1.180 0.783 0.632 2.007 1.195 0.626 0.589 2.657 2.530 2.021 1.793

Cx-Dx RMD0 .9823 .9907 .9970 .9975 .9987 .9981 .9966 .9993 .9955 .9996 .9994 .9998 .9995 .9986 .9938 .9999 .9819 .9995 .9853 .9922 .9762 .9989 .9911 .9912

r

7.93 7.68 6.64 4.99 4.76 5.49 6.10 4.89 5.52 4.18 8.08 4.63 4.84 4.98 4.64 4.41 5.71 4.35 3.63 3.66 7.39 7.52 6.82 6.48

-nD

2.659 2.660 2.012 1.135 1.157 1.833 1.786 1.293 1.908 0.581 2.865 0.974 0.967 1.158 0.831 0.510 1.884 0.973 0.390 0.418 2.460 2.542 2.086 1.916

Cx-THF RMD0

r .9931 .9998 .9959 .9994 .9986 .9993 .9978 .9986 .9962 .9983 .9835 .9938 .9957 .9925 .9973 .9947 .9824 .9902 .9736 .9789 .9945 .9912 .9860 .9881

Constants nD and RMD0 of Eq. 2 and Average RMD0 Values for Eluents Containing Cyclohexane*

Table 3

2.575 2.544 1.906 1.099 1.135 1.783 1.678 1.230 1.947 0.581 2.757 1.132 1.066 1.176 0.790 0.551 2.049 1.116 0.563 0.563 2.506 2.445 2.026 1.834

Mean RMD0

1482 PETROVIj ET AL.

25 5.50 2.401 26 3.26 0.945 27 3.11 0.850 28 3.61 0.941 29 4.81 1.613 30 3.51 0.816 31 3.74 1.161 __________________ * r = correlation coefficient.

.9985 .9952 .9949 .9957 .9927 .9960 .9909

7.97 5.39 5.06 5.83 5.89 5.51 5.97

2.308 1.013 1.020 1.185 1.503 1.048 1.315

.9906 .9950 .9992 .9974 .9966 .9983 .9866

6.06 4.39 4.63 5.05 5.04 5.20 4.92

1.961 0.812 0.931 1.115 1.380 1.049 1.080

.9922 .9885 .9939 .9874 .9927 .9952 .9906

6.21 4.20 4.11 4.39 4.85 4.20 3.83

2.124 0.849 0.819 0.934 1.485 0.845 0.742

.9912 .9900 .9916 .9940 .9979 .9934 .9938

2.199 0.897 0.905 1.044 1.495 0.940 1.075

2,5-ANHYDROALDOHEXOSE ETHYLENE ACETALS 1483

-nD 4.42 3.76 3.71 3.17 2.66 4.18 3.20 2.79 4.35 2.06 4.88 3.33 2.70 2.40 2.85 1.82 4.44 2.80 1.87 1.87 4.59 4.22 3.43 3.42

Cpd. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24

1.949 1.550 1.092 0.683 0.341 1.714 0.850 0.445 1.871 -0.200 2.327 0.651 0.160 0.230 0.231 -0.249 1.680 0.641 -0.237 -0.330 1.830 1.706 1.009 0.967

Tl-EtAc RMD0 .9968 .9940 .9963 .9989 .9987 .9975 .9942 .9983 .9992 .9987 .9957 .9907 .9907 .9983 .9979 .9956 .9974 .9999 .9985 .9985 .9990 .9991 .9931 .9998

r 5.94 4.78 3.39 3.07 3.05 4.02 3.09 2.92 5.25 1.94 7.13 2.86 2.97 3.20 3.08 2.14 4.73 2.99 2.15 1.97 5.50 5.25 3.25 3.33

-nD 1.546 1.140 0.402 0.162 0.039 0.961 0.275 -0.038 1.446 -0.525 2.462 0.151 -0.164 -0.024 -0.089 -0.374 1.235 0.224 -0.311 -0.407 1.410 1.074 0.459 0.379

Tl-An RMD0 .9952 .9992 .9948 .9857 .9967 .9910 .9989 .9809 .9881 .9941 .9996 .9958 .9965 .9884 .9830 .9997 .9978 .9989 .9999 .9999 .9975 .9986 .9998 .9986

r 6.41 5.19 3.94 3.72 3.53 5.05 3.55 3.51 5.74 3.43 7.26 3.71 3.56 2.91 2.86 2.47 5.32 3.78 2.48 2.70 4.95 4.81 3.80 3.58

-nD 1.650 1.083 0.528 0.200 0.028 1.150 0.394 -0.070 1.526 -0.368 2.500 0.194 -0.032 -0.054 -0.091 -0.426 1.398 0.387 -0.434 -0.249 1.203 0.961 0.526 0.482

Tl-Dx RMD0 .9974 .9987 .9847 .9944 .9995 .9958 .9999 .9998 .9909 .9978 .9933 .9856 .9999 .9994 .9892 .9947 .9994 .9992 .9940 .9992 .9996 .9997 .9970 .9960

r

6.25 5.46 3.81 3.45 3.00 5.70 3.35 3.11 6.13 2.89 7.33 3.63 3.02 3.30 3.00 2.89 5.33 4.06 2.51 2.83 5.39 5.27 3.74 3.51

-nD

1.584 1.231 0.650 0.277 -0.013 1.309 0.458 0.174 1.583 -0.401 2.401 0.280 -0.163 0.077 -0.016 -0.401 1.451 0.351 -0.369 -0.415 1.217 1.170 0.530 0.471

Tl-THF RMD0

Constants nD and RMD0 of Eq. 2 and Average RMD0 Values for Eluents Containing Toluene*

Table 4

.9972 .9964 .9999 .9999 .9985 .9985 .9906 .9922 .9901 .9999 .9920 .9994 .9931 .9954 .9908 .9999 .9951 .9960 .9665 .9985 .9999 .9998 .9813 .9997

r

1.682 1.251 0.668 0.331 0.099 1.284 0.494 0.128 1.607 -0.373 2.423 0.319 -0.050 0.057 0.009 -0.363 1.441 0.401 -0.338 -0.350 1.410 1.228 0.631 0.575

Mean RMD0

1484 PETROVIj ET AL.

25 4.19 2.000 26 2.49 0.372 27 2.42 0.452 28 2.99 0.470 29 3.88 1.120 30 2.55 0.120 31 3.13 0.326 __________________ * r = correlation coefficient.

.9972 .9989 .9999 .9952 .9965 .9984 .9915

5.37 2.55 2.69 2.60 4.18 2.45 3.50

1.168 -0.135 0.118 0.097 0.654 -0.133 0.244

.9999 .9996 .9994 .9991 .9980 .9978 .9981

4.99 3.37 3.20 3.50 4.40 3.20 3.44

1.199 -0.044 0.124 0.161 0.700 -0.125 0.310

.9979 .9997 .9997 .9999 .9998 .9981 .9998

5.11 3.84 4.30 4.38 4.43 3.05 3.72

1.234 0.272 0.355 0.285 0.718 -0.115 0.374

.9971 .9999 .9963 .9981 .9952 .9882 .9950

1.425 0.116 0.257 0.253 0.789 -0.063 0.314

2,5-ANHYDROALDOHEXOSE ETHYLENE ACETALS 1485

1486

PETROVIj ET AL. Table 5

Slopes and Intercepts of the Linear Relationship Between nD and RMD0, and nD and n in NPC, and Between m and RMW0 in RPC* NPC

Slope

Intercept RMD0/nD

r

Slope

Intercept n/nD

r

Cx-EtAc Cx-An Cx-Dx Cx-THF Tl-EtAc Tl-An Tl-Dx Tl-THF

0.742 0.441 0.551 0.534 0.847 0.530 0.587 0.558

-1.702 -1.251 -1.633 -1.432 -1.927 -1.471 -1.859 -1.779

0.9802 0.9745 0.9829 0.9703 0.9632 0.9759 0.9790 0.9672

1.24 2.09 1.43 1.67 1.59 2.10 1.80 2.19

-0.021 0.194 0.276 -0.033 0.174 -0.014 0.556 -0.092

0.9997 0.9986 0.9779 0.9998 0.9759 0.9971 0.9611 0.9852

Average RMD0/nD Cx Tl RPC

0.544 0.638

-1.437 -1.813

Average n/nD 0.9759 0.9914

1.57 1.96

-0.092 0.049

0.9982 0.9952

RMW0/nD

MeOH0.977 -0.968 H2O ACN1.145 -1.667 H2O __________________ * r = correlation coefficient.

0.9977 0.9912

There is, however, no correlation between n and RMo values. For that reason, we have introduced into Eq. 1 the volume fraction of diluent, ϕD, instead of ϕ , so that Eq. 1 can be written as RM = RMDo - nD log ϕD

(2)

where RMDo is an extrapolated RM value at ϕD = 1, and nD is a constant with negative sign. The numerical data for the constants nD and RMDo are presented in Tables 3 and 4. It should be pointed out that Eq. 2 is valid for the narrow range of ϕD values, because there is no linear relation between log ϕ and log ϕD.

2,5-ANHYDROALDOHEXOSE ETHYLENE ACETALS

1487

Table 6 Constants m and RMW0 of Eq. 3 for Eluents Containing Methanol and Acetonitrile

Cpd. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16

MeOH-H2O ϕorg = 0.6-0.8 -m RMW0 2.01 2.13 3.01 5.13 5.92 2.61 3.39 5.56 2.20 6.65 1.91 5.33 6.12 5.94 7.22 6.45

1.040 1.190 1.937 4.094 4.730 1.583 2.335 4.568 1.099 5.457 0.803 4.135 4.870 4.736 6.086 5.460

ACN-H2O ϕorg = 0.5-0.7 -m RMW0

2.38 2.92 3.43 2.10 2.62 3.30

1.002 1.609 2.189 0.627 1.308 2.059

3.52

2.288

2.72 3.32 3.28 3.70 3.56

1.460 2.066 2.057 2.501 2.532

Cpd. 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31

MeOH-H2O ϕorg = 0.6-0.8 -m RMW0 2.32 3.13 6.51 5.51 1.97 2.09 3.63 3.85 2.49 4.94 4.43 4.95 3.27 5.19 3.03

1.255 2.285 5.329 4.797 0.990 1.149 2.484 2.707 1.371 3.752 3.254 3.950 2.165 4.190 2.093

ACN-H2O ϕorg = 0.5-0.7 -m RMW0 2.23 3.53 3.30

0.907 2.512 2.230

2.33 2.21 2.15 3.03 2.51 2.66 2.20 3.06 2.25

1.087 0.979 0.707 1.687 1.218 1.413 0.832 1.942 1.003

For ϕ values used and the same retention data, good correlation coefficients of the linear regression of experimental RM values were obtained for both equations (Tables 1-4). Some correlation lines are given in Fig. 2. Constants nD and RMDo generally follow the retentivity of the compounds; therefore, there is good linear correlation between them (Fig. 3 and Table 5). Good linear relationship is also obtained between constants n and nD (Table 5). As constants nD and RMDo of a particular compound are similar for the same diluent and various eluting systems, their mean values (Tables 3 and 4) will be used for discussion of solutes retention behavior. The retention of all studied compounds remarkably depends on the type of substituents and on the presence of other substituents in a molecule, but less on their positions or orientations. For example, compounds 1, 2, 21, and 22 comigrated in mobile phases with cyclohexane, suggesting the same retentivity of hydroxy and mesyloxy functions. Likewise, compounds 4 and 5 were similarly retained, as were compounds 12, 13, and 14. Compounds 23 and 25, as well as compounds 29 and 31, were, however, clearly resolved. Compounds with hydroxy groups were more retained.

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PETROVIj ET AL.

The effect of other substituents in a molecule on retention is evident. As expected, replacement of hydroxyl hydrogen or mesyl function with any other substituent resulted in reduced retention and, consequently, in decreased n and nD values. Substitution of cyclohexane with toluene as the diluent generally resulted in increased compound mobility, especially for those compounds with mesyloxy groups. This is evident from the RMDo data in Table 4. Exchange of hydroxy group for mesyloxy results in significantly higher ∆RMDo data for mobile phases containing toluene as diluent than those containing cyclohexane. Compound pairs 1-2, 21-22, and 4-5 were not resolved in systems with cyclohexane while, in systems with toluene, these pairs of compounds were completely separated with higher retention of those compounds possessing hydroxy groups. Increased compound mobility in systems with toluene also resulted in lower constants n, nD, and RMDo in comparison with mobile phases containing cyclohexane. The behavior of compounds bearing benzoyl or benzyl functions was generally as expected. The benzyl group is less polar than benzoyl; however, both groups caused low retentivity of the corresponding compounds. Compounds 6, 9, and 17 were of the similar retentivity, as were compounds 4 and 18, suggesting the similar effect of the benzyloxy or azide function on retention. A fluorine atom (compound 20) showed the same effect on retention as did an azide function (compound 19).

Reversed Phase Chromatography (RPC) The retention data obtained in RPC were correlated with the volume fraction of organic component, ϕorg, in a binary aqueous mobile phase according to the equation RM = RMWo + m ϕorg

(3)

where RMWo is an extrapolated RM value in pure water, and m is a constant with negative sign. The numerical data for the constants m and RMWo are given in Table 6. Correlation coefficients of the linear regression of the experimental RM values varied from 0.9971-0.9999. For some compounds, the retention constants were not measured, due to their great mobility in mobile phases containing acetonitrile. There is good correlation between RMWo and absolute m values (Table 5). The slopes of the lines are about unity, indicating that the slope increments, ∆m, are approximately equal to the retention increments, ∆RMWo (Table 7). According to Eq. 3, RMWo values for the particular compound and various

4 and 5 1 and 2 12 and 13 21 and 22 17 and 18 1 and 3 4 and 6 18 and 19 17 and 19 9 and 10

3-OHÆ3-OMs 4-OHÆ4-OMs

3,6-OHÆ3,6-OBz 4,6-OHÆ4,6-OBz

3-OHÆ3-OBz 4-OHÆ4-OBz 6-OHÆ6-OBz

Compounds Compared

Converted Substituent 0.79 0.12 0.79 0.12 0.81 1.00 2.52 3.38 4.19 4.45

0.64 0.15 0.74 0.16 1.03 0.90 2.51 3.04 4.07 4.36

MeOH-H2O ∆m ∆RMW0

0.98 1.61

0.61

0.60

0.82 1.30

0.58

0.51

ACN-H2O ∆m ∆RMW0

3-N3Æ3-F 6-OMsÆ6-J

4-OHÆ4-N3 4-OBzÆ4-OBn

4-OHÆ4-Obn 3-OHÆ3-OTs 4-OHÆ4-OTs 4-OHÆ4-OTf

Converted Subsituent

1 and 7 29 and 30 21 and 23 12 and 15 21 and 24 12 and 19 5 and 8 3 and 7 19 and 20 21 and 28

Compounds Compared

1.38 1.92 1.66 1.89 1.88 1.18 0.36 0.38 1.00 2.98

1.30 2.03 1.49 1.95 1.72 1.19 0.16 0.40 0.53 2.96

MeOH-H2O ∆m ∆RMW0

∆m and ∆RMW0 Values for Conversion of Substituents in Compound Molecules

Table 7

0.81 0.13 0.24 0.23

0.98

0.86

1.05 0.13 0.31 0.28

1.04

1.11

ACN-H2O ∆m ∆RMW0

2,5-ANHYDROALDOHEXOSE ETHYLENE ACETALS 1489

1490

PETROVIj ET AL.

mobile phases should be equal at ϕorg = 0. There is, however, significant difference between the RMWo values calculated for the mobile phases used, suggesting that the relationship between RM and ϕorg is not linear for the wide range of ϕorg.9-11 The retention of the compounds depended on the hydrophobicity of substituents in a molecule. Compound 15, bearing three hydrophobic substituents, was most retained. Therefore, the effect of substituents on retention will be discussed using ∆m and ∆RMWo values presented in Table 7. It is evident from these data that, for the same substitution and various solute pairs, both increments are reasonably constant and independent of the substituent position or orientation. The presence of other substituents in a molecule affects retention and, consequently, relative retention. For example, the mesyloxy function is slightly more hydrophobic than hydroxy; however, for conversion of hydroxy to mesyloxy function, the presence of hydrophobic benzyloxy or benzoyloxy functions in a molecule (compound pairs 4-5 and 12-13 ) increases both increments in comparison with the presence of other mesyloxy groups (compound pairs 1-2 and 21-22). This is also observed for replacement of hydroxy with a benzoyloxy function. For compound pairs 4-6 and 18-19, both ∆m and ∆RMWo values are significantly higher than those for compound pairs 1-3 and 17-18. Hydrophobicities of benzyl, benzoyl, tosyl, or trifluoromethanesulfonyl functions are generally similar; azide is slightly more polar. Halogens, as electron-donating atoms, exhibit high hydrophobicity. Therefore, the retention difference between compounds 21 and 28 is about 3 (Table 7). The retention sequence of compounds is generally opposite in RPC compared to that in NPC, especially when particular solute series are considered. However, an exception is observed, i.e., compound 15 was more retained than compound 16 in both NPC and RPC. Also, in both NPC and RPC, compounds 16 and 19, which are of similar functionality, co-migrated; therefore the trifluoromethanesulfonyl function is responsible for higher retention of compound 15. The trifluoromethyl moiety is hydrophobic and affects retention in RPC, while the sulfonyl moiety determines retention of compound 15 in NPC.

REFERENCES 1. S. Hanessian, Total Synthesis of Natural Products: The Chiron Approach, Organic Chemistry Series, Pergamon Press, Oxford, 1983. 2. S. M. Petrovi, E. Lonœar, M. Popsavin, V. Popsavin, J. Planar Chromatogr., 10, 427-431 (1997).

2,5-ANHYDROALDOHEXOSE ETHYLENE ACETALS

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3. M. Popsavin, V. Popsavin, N. Vukojevi, J. Csanadi, D. Miljkovi, Carbohydr. Res., 260, 145-150 (1994). 4. V. Popsavin, O. Beri, M. Popsavin, J. Csanadi, D. Miljkovi, Carbohydr. Res., 269, 343-347 (1995). 5. V. Popsavin, O. Beri, M. Popsavin, J. Csanadi, S. Lajši, D. Miljkovi, Collect. Czech. Chem. Commun., 62, 809-815 (1997). 6. V. Popsavin, O. Beri, M. Popsavin, S. Lajši, D. Miljkovi, Carbohydr. Lett., 3, 1-8 (1998). 7. E. Soczewinski, Anal. Chem., 41, 179-182 (1969). 8. P. Jandera, J. Churaœek, J. Chromatogr., 91, 207-221 (1974). 9. P. J. Schoenmakers, H. A. H. Billiet, R. Tijsen, L. de Galan, J. Chromatogr., 149, 519-537 (1978). 10. P. J. Schoenmakers, H. A. H. Billiet, L. de Galan, J. Chromatogr., 185, 179195 (1979). 11. B. Peki, S. M. Petrovi, B. Slavica, J. Chromatogr., 268, 237-244 (1983).

Received May 27, 1998 Accepted July 10, 1998 Manuscript 4902-TLC

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