SULFUR BONDED UNSYMMETRICAL BOROLE COMPLEXES: SYNTHETIC, SPECTROSCOPIC AND BIOCIDAL ASPECTS

Taruna Pandey1, V. P. Singh2 and R. V. Singh1*

1 Department of Chemistry, University of Rajasthan, Jaipur-302004, India 2 DSME, S.C.E.R.T., Varun Marg, Defence Colony, New Delhi-110024, India

ABSTRACT Synthesis, characterization and biological activities of some of the unsymmetrical borole complexes containing B-S and B<-N bonds are described. The sulfur containing conjugated bases were prepared by the condensation of caibonyl compounds with thiosemicarbazide. The resulting complexes have been characterized by elemental analysis, molecular weight determinations and spectral studies including IR, Ή NMR, "Β NMR and ,3C NMR. The spectroscopic results showed that the conjugated bases behave in a bidentate fashion, whereas, the complexes display a tetracoordinated environment around the atom as steriochemically active lone pair is also included in the coordinated sphere. The conjugated bases and their respective boron complexes have been screened for their antifungal and antibacterial properties.

INTRODUCTION Recently, few references on bidentate ligands coordinated to boron through the azomethine and the thiolo sulfur have been published in the literature1·2. These compounds are interesting in many respects. Some have got a number of applications in industry, biology and agriculture. Industrial applications of coordination compounds of boron have remained fairly limited. Boron analogues of carboxylic acids and peptides have been shown to possess interesting biological activities in particular as serine protease inhibitors3. Use of boron-nitrogen compound in neutron capture therapy of brain tumors have also been investigated4. Several organoboranes find promising applications in the synthesis of insect pheromones5. Metalloboranes are widely used in industry and medicine as plastic stabilizers, polymerization accelerators, lubricants and bactericides. Some boron complexes containing nitrogen and sulfur have been shown to exhibit antifungal and antibacterial properties6. Several chelates of boron have been tested for potential pharmaceutical applications and some of them have been introduced in to practice7·8. Thiosemicarbazides and conjugated bases have attracted much attention due to their various biological activities9. It was found that some drugs have increased activity when administered as metal complexes10 and a number of metal chelates were found to inhibit tumor growth". In the treatment of cancer the active species is a metal chelate of conjugated base12. Following the work of Mashima13, coordination chemists have taken greater interest in thiosemicarbazides and their azomethines as potential ligands. Boron complexes of these ligands have been found to possess conspicuous biocidal activity14. It is however interesting that the biological activity gets enhanced on undergoing complexation with the metal ions15"16. In view of the above and the success achieved earlier in the synthesis and characterization of Schiff base complexes of boron, it was considered of interest to synthesize a wide variety of Schiff base derivatives of boron by the reactions of unsymmetrical borole and the Schiff bases having nitrogen and sulfur as coordination sites. The conjugated bases used during these investigations are: (i) 2-[ 1 -(2-Naphthenyl)ethylidene]hydrazine carbothioamide (L,H) (ii) 2-[l-(2-Pyridinyl)ethylidene]hydrazine carbothioamide (L2H) (iii) 2-[l-(2-Thienyl)ethylidene]hydrazine carbothioamide (L}H) RESULTS AND DISCUSSION The 1:1 molar reactionso f unsymmetrical borole, 2-isopropoxy-4-methyl-l ,3,2-dioxaborolane B(C3H602) (OPi*) and 2 -isopropoxy-4-methyl-1,3,2 -dioxaborinane B(C4H,0.,) (OPf) with 2-[l-(2-thienyl)ethylidene]hydrazine carbothioamide, 2-[l-(2-pyridinyl)ethylidene] hydrazine carbothioamide and 2-[l-(2-naphthenyl)ethylidene] hydrazine carbothioamide can be represented by the following equations :

185 Vol. 21, No. 4, 1998 Sulfur Bonded Unsymmetrical Borole Complexes: Synthetic, Spectroscopic and Biocidal Aspects

1 [B(C3H602) (OPr )] + LnH • [B(C3H605) (Ln)]+ΡΛ>Η

Benzene [Bi^HgOj) (OPr*)] + LH — > [Β^Η,Ο,) (L)] + PrOH

Where, LnH represents the conjugated base molecule and η = 1, 2 or 3. The resulting coloured solids are soluble in MeOH, DMF and DMSO. The method used for the preparation and isolation of the resulting complexes give materials of good purity as supported by their analysis. These are monomelic, non-electrolytes, stable and resistant to hydrolysis.

IR Spectra The comparison of the infrared spectra of the conjugated bases and their complexes suggests that conjugated bases are bidentate with the thiolo-sulfur and azomethine nitrogen as the coordinated sites. Strong bands in the region 3300-3100 cm"1 in the conjugated bases disappear in the corresponding borole complexes, indicating the deprotonation of the -NH functional group. The conjugated bases exhibit a sharp band at 1610-1590 cm'1 due to stretching mode of azomethine group. The band shifts to the higher frequency region (ca. 15 cm1) on complexation which is due to an increase in the bond order17 as a result of (B<-N) bond formation. An indicative evidence for the support of the structure has been noted by the new strong and sharp peaks appearing at 1360, 875 and 1550 cm1 assigned to ν (B-O)18 v(B-S)19 and v(B<-N)J0 modes, respectively.

Ή NMR Spectra Table I: Ή and "B NMR Spectral Data δ, ppm/(JHH) of Conjugated Bases and their Corresponding Unsymmetrical Borole Complexes

U Compound -NH -NH2 Aromatic Protons* -CH, B

0») (b.) 1 3 4 5 6 7 (·) (»)

L)H 10.65 2.90 8.93(s) 8.0-8.32(m) 7.84-7.55(m) 1.91 .

BCCJHIOZ) (Ll) - 2.86 8.96(s) 8.72-8.16(m) 8.08-7.76(m) 2.16 1.06

B^HGOJ) (Ll) - 2.88 8.98(*) 8.56-8.0(m) 7.92-7.60(m) 2.20 0.8

LjH 10.64 2.84 - 8.24(d) 8.88(dd) 7.48(dd) 8.92(d) 1.82 - (7.1Hz) (7.1Hz)(7.8Hz) (7.8Hz)(7.5Hz) (7.5Hz)

BiCjHeOz) (L2) - 2.72 - 8.32(d) 8.96(dd) 7.52(dd) 9.04(d) 1.94 4.86 (7.2Hz) (7.2Hz)(7.9Hz) (7.9HzX7.5Hz) (7.5Hz)

BiC^gOj) (Lj) - 2.80 8.40(d) 9.04(dd) 7.60(dd) 1.91 4.07

(7.1Hz) (7.1Hz)(7.9Hz) (7.9HzX7.4Hz) I

L3H 10.68 2.81 - 7.16(d) 7.60(dd) 8.68(d) - 1.68 - (7.4Hz) (7.4HzX7.3Hz) (7.3Hz)

BfCjHiOj) (Lj) - 2.72 - 7.68(d) 8.16(dd) 8.72(d) - 1.88 6.51 (7.3Hz) (7.3HzX7.4Hz) (7.4Hz)

WWW (Lj) - 2.78 - 7.42(d) 7.76(dd) 8.82(d) - 1.80 7.6 (7.2Hz) (7.2HzX7.4Hz) (7.4Hz)

•Aromatic protons of first three compounds are observed in two group of multiplets (3,4,5,8 & 6,7). The proton magnetic resonance spectral data of hydrazine carbothioamides of [l-(2-pyridinyl) ethanone], [l-(2-naphthenyl)ethanone] and [l-(2-thienyl)ethanone] and their corresponding unsymmetrical borole complexes have been recorded in DMSO-d6. The chemical shift values relative to the TMS peak are listed in Table I. is® Φ Φ

186 Taruna Paney, V.P. Singh and R. V. Singh Main Group Metal Chemistry

The Ή NMR spectra of each synthesized conjugated bases exhibit a singlet in the region δ (10.64- 10.68) due to -NH proton which disappears in the spectra of complexes suggesting the coordination of nitrogen as well as sulfur to the boron atom. However, in the 1:1 complexes of unsymmetrical borole the signal due to -NH2 protons observed nearly in the same region δ (2.72-2.90) ppm showing that the deprotonation does not take place and the bonding may take place only through the nitrogen and sulfur atom of the conjugated bases. In the spectra of the complexes the deshielding in the position of CHj-C=N protons further substantiates the coordination of azomethine nitrogen to the boron atom. The downfield shift indicates the delocalisation of the electronic charge within the chelate ring and thereby the stabilization.

"C NMR Spectra I3 The C NMR spectral data of L,H, L2H, LjH and their unsymmetrical borole complexes show marked shifts in the positions of thiolo and azomethine attached to the participating groups clearly showing the complexation of boron through nitrogen and sulfur atoms (Table II). Table Π: 13C NMR Spectra Data (δ ppm) of Conjugated Bases and their Corresponding Unsymmetrical Borole Complexes.

Compound Chemical Shift Values Thiolo Azomethine Methyl Aromatic Carbons Carbon Carbon

L]H 178.99 147.84 13.92 C1>133.33;C2,135.11;C3,132.84; Gt, 130.25; C5,129.64; C6,127.64; C7,127.47; Cg,126.88; C9,123.57; C]0,124.06

B(C3H602XLJ) 170.87 139.77 13.99 C] ,135.50; C^135.09; C3,134.12; C4,131.22; C5,129.58, Q;, 127.49; C7,129.54; Cg, 126.90; C9,123.56; C10,126.02

B(C4H802XLI) 168.92 141.38 13.76 C],133.45; C2,135.22; C3,132.67; C4,133.54; C5,131.67; Qs, 127.52; C7,128.84; C8,127.71; C9,127.56; C10,125.13

LqH 179.19 156.20 11.33 C2,147.52; C3,123.04; C4,119.89; C5)135.30;C6,146.88

173.22 148.17 11.40 C2,148.08; C3,123.13; C4,121.45; C5,136.62; Qs, 149.81

B(C4H802XL2) 164.13 146.54 11.16 C2,147.46; C3,124.98; C4,119.95; C5,137.15; Cö,148.88

L3H 178.62 152.76 13.17 C2,140.31; C3,127.70; C4,121.45; C5,125.37

B(C3H6Q2XL3) 169.28 146.86 13.09 C2,141.35; C3,128.02; C4,121.98; C5,127.30

B(C4H802XL3) 172.52 145.64 13.23 C2,140.86; C3,129.96; C4,122.69; C5,126.64

187 Vol. 21, No. 4, 1998 Sulfur Bonded Unsymmetrical Borole Complexes: Synthetic, Spectroscopic and Biocidal Aspects

11Β NMR Spectra The "B NMR spectral data showing peaks at δ 1.06,0.8,4.86,4.07 6.51 and 7.6 ppm in ΡίΟ,Η,Ο^)] [EK^HA) (L,)], [B(C3H602)(L2)], PiC^HgOj) (L2)], [B(C3H602)(L,)] and [Β^Ο,) (L}), respectively,indicatin g the tetra coordinated state21 of boron in these unsymmetrical borole derivatives.

HjC O. /-c^ Ov /S^NH, ι ii L \y η H(i—θ/ Vt!* HC^ -—O ^» «-cogi-a-O H,C / \ R ^ H,C X R

Thus on the basis of above discussions it is clear that the conjugated bases by coordinating to boron atoms through the thiolo group and azomethine nitrogen behave as monobasic bidentate agents.

BIOCIDAL ACTIVITY The data of fungicidal and bactericidal activities of conjugated bases and their respective unsymmetrical borole complexes against pathogenic fungi and bacteria were recorded in Tables III and IV. The studies show that the concentrations reached level that are sufficient to inhibit and kill the pathogens. Further, the results achieved from biological activity have also been compared with the conventional fungicide, Bavistin and conventional bactericide Streptomycin. The superiority of the unsymmetrical borole complexes as compared to the conjugated bases can well be understood by considering the chelation theory". Table m: Fungicidal Screening Data of Compounds (percent growth inhibition after 96 hrs. at 25±2°C)

Compound Fusarium oxysporum Macrophomina phaseolina 25 50 100 200 25 50 100 200

L,H 17 32 54 58 18 34 45 52 Lp 18 30 55 61 17 33 47 52 L,H 21 34 56 60 19 35 45 50

B(C3H p2)(L,) 29 50 67 77 29 43 51 61

B(C3HS02)(L2) 31 62 78 80 31 45 53 62

®(CjH602) (LJ) 46 68 81 Μ 38 48 56 62 BCC^XL,) 65 74 83 86 45 51 63 71 Bcc^cgcL,) 68 77 85 88 47 53 60 69

B(C4H802)(L3) 73 79 86 90 49 59 68 79 Standard 82 86 100 100 80 82 100 100 Bavistin

188 Taruna Paney, VP. Singh and R. V. Singh Main Group Metal Chemistry

Table IV: Bactericidal Screening Data of Compounds (Diameter of inhibition zone (mm) after 24 hrs at 30±2°C.) Compound Pseudomonas Xanthomonas Escherichia Staphylococcus cepacicola (-) compcstris coli (-) aureus (+) 500 1000 500 1000 500 1000 500 1000 L,H 4 6 4 6 3 5 5 7 hp 3 5 4 5 3 5 4 7 L,H 5 10 5 6 4 6 5 8 B^H^XL.) 6 9 5 7 5 7 6 9

B(C4Hp2)(L2) 4 7 6 8 7 9 9 10 BiC^HgOj) (Lj) 8 10 7 7 7 8 8 13 B^H/^KL,) 5 8 9 11 9 14 12 17

B(C3H602)(L2) 10 19 8 12 10 13 13 15

B(C,H602)(L5) 10 14 10 14 11 13 11 16 Standard 2 3 3 5 17 18 15 17 (Streptomycin)

In fungicidal activity although the bioactivity increased on undergoing complexation, it has also been observed that sulfur containing compounds are more toxic23 than oxygen containging compounds. If fungitoxicity is dependent on one or more chemical reactions as it must undoubtedly be in most of the cases than any property of the fungitoxic molecule which would increase the rate of chemical reactions must perforce enhance fungitoxicity24.

EXPERIMENTAL Adequate care was taken to keep the unsymmetrical borole complexes. Chemicals and solvents used were dried and purified by standared methods and moisture was excluded from the glass appratus using fused calcium chloride drying tubes. The conjugated bases and unsymmetrical borole were prepared by the literature methods and all the manipulations were carried out under anhydrous conditions.

Preparation of Conjugated Bases The conjugated bases were prepared by the condensation of ketones with hydrazine carbothioamide in equimolar ratio in absolute alcohol. The contents were refluxed for 45 minutes, recrystallized from the same solvent and dried under reduced pressure. The physical properties of these moieties are as follows : Conjugated Bases, Colour and M.P. (°C) 2-[l-(2-Naphthenyl)ethylidene]hydrazine carbothioamide, White ciystalline solid, 144°C-

Synthesis of Unsymmetrical Borole Complexes The unsymmetrical monoisopropoxy borane was taken in benzene and an equimolar quantity of the conjugated bases (L,H, L2H and L3H) was added to it. The reaction mixture was refluxed for a suitable period and the liberated isopropanol was fractionated azeotropically with benzene. After the reaction was complete the excess of the solvent was removed under vacuum and the product dried at 40°C/0.1 mm for four hours. These were recrystallized from a 1:1 solution of benzene and alcohol. All the compounds were isolated as powdered solids. The details of these reactions and the analyses of the resulting products are recorded in Table V.

189 Vol. 21, No. 4, 1998 Sulfur Bonded Unsymmetrical Borole Complexes: Synthetic, Spectroscopic and Biocidal Aspects

Table V: Analyses and Physical Properties of Unsymmetrical Borole Complexes Product formed MP Analysis% MoLWt and Colour (°Q Ν S Β Found Found Found Found (Calcd) (Calcd.) (Calcd) (Calcd)

1. B(C4H802)(L,) 200-204 12.25 9.21 3.12 375 Yellow (12.31) (9.39) (3.16) (341)

2. EKC^CgiL,) 170-172 19.11 10.71 3.63 322 Lamon Yellow (19.17) (10.97) (3.70) (292)

3. BiC^CW 178-180 15.79 11.94 3.99 289 Cream (15.84) (12.09) (4.07) (265)

4. ΒίςΗρ,)^) 197-200 12.75 9.62 3.22 350 Yellow (12.84) (9.79) (3.30) (327)

5. B(C3H602)(L2) 158-162 20.02 11.45 3.80 298 Yellow (20.14) (11.52) (3.88) (278)

6. ^(CjHjOj) (Lj) 176-179 14.69 22.48 3.75 302 Off white (14.83) (22.64) (3.81) (283)

Analytical Methods and Physical Measurements Carbon and analyses were performed at the microanalytical Laboratory of the Department. Nitrogen and sulfur were estimated by Kjeldahl's and Messenger's methods, respectively. The molecular weights were determined by the Rast camphor method. IR spectra were recorded on a Perkin Elmer 577 Grating Spectrophotometer in the range 4000-200 cm1 as Nujol mulls using KBr optics. Ή NMR spectra were recorded in DMSO-d6. "C NMR spectra were recorded in distil DMSO, using TMS as the standard. "B spectra were scanned on a Bruker WH90 Spectrometer operating at 64.21 MHz at 31°C using BF3.EtjO as an external standard.

IN VITRO STUDY : Fungicidal Screening (Poisoned Food Technique) Potato-dextrose agar medium was prepared in flask and sterilized. Accurate amount of all the compounds were added after being dissolved in methanol so as to get certain final concentrations of 25, 50, 100 and 200 ppm. Aliquots of 15 ml medium was poured in 90 sterilized Petri plates. A culture of test fungus is grown on PDA for certain period (generally seven days) at the optimum temperature for growth. Small disc (0.7 cm) of the fungus culture is cut with sterile cork borer and transfered as aseptically in the centre of a Petri dish containing the medium with a certain amount of fungicide. Control set without compounds was also maintained and a standard fungicide (Bavistin) was taken. Plates were incubated for 4 days at 25±2°C and colony diameter was measured after the incubation period of growth. The percentage inhibition of growth was calculated by percentage growth inhibition = (C-T) C1 χ 100, where, C = growth in control, Τ = growth in treatment.

2. Bactericidal screening (Inhibition Zone Technique) Flat bottomed 90 mm Petri discs were used and nearly 15 ml of the beef extract medium (Peptone -5 g., beef extract 5 g., NaCl 8 g., agar-agar 20 g and 1000 ml distilled water) was pipetted into the Petri disc. Bacterial suspension was added in the medium and after some time bacterial growth was seen in the medium. The tested compounds were dissolved in the methanol of 500 and 1000 ppm concentrations. Paper discs of

190 Taruna Paney, V P. Singh and R. V. Singh Main Group Metal Chemistry

Whatman NO. 1 filter paper of 5 mm diameter were soaked in these solutions of varied concentrations. The discs were dried and placed on the medium with organism in Petri disc at suitable distances. These Petri discs were incubated for 48 hours at 25±2°C and zone of inhibition was measured in mm.

ACKNOWLEDGEMENT The authors are thankful to U.G.C., New Delhi for financial support.

REFERENCES 1. N. Fahmi, and R.V. Singh, Phosphorus, Sulfur and Silicon, 104 (1995), 53. 2. C. Saxena and R.V. Singh, Phosphorus, Sulfur and Silicon, 97 (1994), 17. 3. D.S. Matteson, Chem. Rev., 89 (1989), 535 and refrences cited therein. 4. C.A. Perks, A.J. Mill, G. Constantine, K.G. Harrison and J.A.B. Gibbson, British J. Radiol 61 (1988), 1115. 5. S. Singh, R. Dhillon and J. Singh, Indian J. Chem., 306 (1991,) 355. 6. V.P. Singh, R.V. Singh, Indian Nal. Acad. Sei. Lett, 12 (1989), 311. 7. W. Kliegal 'Boron in Biologie, Med. Pharm. Springer Berlin (1980). 8. W. Kliegel, Pharmazie, 27 (1972), 1. 9. H.G. Petring, H.H. Buskirk and G.E. Underwood, Cancer Res., 64 (1964), 367. 10. D.R. Williams. Chem. Rev., 72 (1972), 203. 11. F.P Owyer, E. Mayhew, E.M.F. Roe and A. Schilman, Br. J. Cancer, 19 (1965), 195. 12. J. A. Crim and H.G. Petering, Cancer Res., 27 (1967), 1278. H.G. Petering and G.J. Vangeissen, "The Biochemistry of Copper·" Academic New York, 197 (1966). 13. M. Mashima, Bull. Chem. Soc. Jpn., 37 (1967), 974. 14. L. Bhal, J.P. Tandon and S.K. Sinha, Curr. Sei., 53 (1984), 566. 15. D. Singh and R.V. Singh, J. Inorg. Biochem., 50 (1993), 227. 16. P.N. Saxena and A.K. Saxena, Appl. Organomet. Chem., 3 (1989), 349. 17. C. Saxena and R.V. Singh, Indian J. Chem., 32 A (1993), 154. 18. S.M. Tripathi and J.P. Tandon, J. Inorg. Nucl. Chem., 40 (1978), 983. 19. R.K. Sharma, R.V. Singh and J. P. Tandon Synth. React. Inorg. Met.-Org. Chem., 9 (1979), 519. 20. K.K. Chaturvedi, RV. Singh and J.P. Tandon, J. Prakt. Chem. 324 (1984), 817. 21. V.P. Singh, R.V. Singh and J.P. Tandon, Synth. React, Inorg. Met.-Org. Chem., 19 (1989), 669. 22. B.G. Tweedy, Phytopathology, 55 (1964), 910. 23. N. Wasi and H.B. Singh, Inorg. Chim. Acta, 151 (1988), 287. 24. J.G. Horsfall and S. Rich, "Fungitoxicity of Sulfur Bridged Compound", Phytopath., VI (1953), 1-4.

Received: September 25, 1997 - Accepted: October 14, 1997 - Accepted in revised camera-ready format: January 14, 1998

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