Synthesis and Characterisation of Some New Aluminium Derivatives of Schiff Bases Containing N, O and S Donor Atoms and the Anti Fertility Activity of the Derivative Al[SC6H4N:C(CH3)CH2COCH3]3

Some new compounds of aluminium having the general formula AI[SC6H4N:C(R) CH2 C(O)R’]3 where R = CH3, R' = CH3 (1); R' = CH3, R' = C6H5 (2); R = CF3, R’ = -C = CH - CH = CHS (3); R = CF3, R’ = C6H5 (4) have been synthesised by the reactions of Al(OPri)3 and the corresponding ligands in 1:3 molar ratios in benzene. Elemental and spectroscopic (IR, 1H, 13C, and 27AI NMR) characterisation of these monomeric compounds reveals monofunctional bidentate behaviour of ligand moiety and the octahedral geometry around aluminium atom. Compound (1), AI[SC6H4N:C(CH3)CH2COCH3], has been tested for its antifertility activity in male albino rats. The oral administration of this compound at the dose level 6.5 rag/rat/day reduced the weights of testes and epididymides. Significant decrease in sperm motility as well as sperm density resulted in the reduction of male fertility by 100%. Production of primary spermatocytes (preleptotene and pachytene), secondary spermatocytes and step-19 spermatids declined by 56.10%, 44.42 %, 63.35 % and 64.57 % respectively. These results indicate that the administration of compound (1) in male rats brought about an interference with spermatogenesis which ultimately caused infertility.


INTRODUCTION
Organic compounds containing-NC6H4S-unit are well known for their significant biological activities /l/. Phenothiazines significantly affect the hypothalamous pituitary gonadal axis, thus resulting in a delay in ovulation and menstruation in women/1/. In general biological activity of such type of compounds enhances considerably on complexation with metal atom/2/. Some organo-antimony(IIl) and -bismuth(Ill) compounds containing-NC6HaS-unit have been reported to show antifertility /3/ and antitumor /4/ activity, respectively.

Bases Containing N, 0 and S Donor Atoms and the Anti Fertility
Benzothiazolines constitute an important class of-NC6H4S-containing ligands. The metal derivatives of this class of ligands have been extensively pursued to investigate the chelating properties of benzothiazolines.
Although mono-and bis-aluminium derivatives/5/of benzothiazolines derived from 13-diketones and 2-aminothiophenol in which the ligands behave as bifunctional tridentate /5,6/ moiety have been reported, no tris-aluminium derivatives of these ligands appear to have been synthesized so far. In view of the above, it has been considered worthwhile to synthesize a new series of tris-aluminum derivatives of benzothiazolines, and to investigate their physico-chemical and structural features as well as the antifertility activity of one of the representative compounds in male albino rats. EXPERIMENTAL Moisture was carefully excluded throughout the experiments. Al(OPri)3 was prepared by literature method /7/. Solvents (E. Merck) were dried by standard procedures/8/. Isopropanol in the azeotrope was estimated oxidimetrically/9/, using 1N KzCr207 solution in 12.5 % H2SO4. Aluminium was estimated gravimetrically as oxinate/9/. Nitrogen and sulphur w.ere determined by Kjeldahl and Messenger methods/9/respectively. 2-Aminothiophenol and [3-diketones were distilled prior to use. The benzothiazolines /10/ RC(NH C6H4S)CH2C(O)R' [where R CH3, R '= CH3(LIH); R CH, R '= C6Hs(L:ZH), R CF3, R' 2-C4H.S (L3H); R CF3, and R' C6Hs(L4H)] have been prepared by the equimolar condensation reactions of 2aminothiophenol and substituted 13-diketones in refluxing benzene. Liberated water was removed azeotropically with benzene. IR spectra of the ligands and their complexes have been recorded on a Nicolet DX FT IR Spectrophotometer using CsI cell. IH, 3C and -7A1 NMR spectra were recorded on a JEOL-FX-90Q (90 MHz) or Bruker DPX 300 MHz Spectrometer either in CDCI3 or C6H6 solution. Molecular weights of these compounds were determined on a Knauer Vapour Pressure Osmometer in benzene solution.
Since all the complexes have been synthesised by the same method, for the sake of brevity, synthesis of one representative compound is given in detail. Synthetic and analytical results of the other compounds are summarised in Table I. SYNTHESIS OF COMPOUND (1) About 25 ml benzene solution of Al(OPri)3 (0.56 g, 2.74 mM) was added to the benzene solution (-20 ml) of the ligand [CH3C(NHC6H4S)CHzCOCH3] (1.71 g, 8.25 mM) and the solution was refluxed on a fractionating column for-4 h. The isopropanol formed in the reaction was fractionated out azeotropically with benzene. Progress as well as the completion of the reaction was checked by the estimation of the isopropanol in the azeotrope by the oxidimetric method/9/. After the completion of the reaction, the excess of the solvent was removed under reduced pressure. For the purification, this compound was dissolved in a small amount of benzene 15 ml and then the petroleum ether (40-60 C) was added slowly until a viscous compound began to separate. The solution was kept overnight at about-10 C. The viscous compound, left after decanting off the solvent, was dried under reduced pressure. Yield, 87% (1.54 g). Male albino rats of Wistar strain weighing 150-180 g obtained from Jamia Hamdard, Hamdard University, New Delhi, were used. Animals were kept in plastic cages with proper aeration and temperature, feeding on a standard rat pellet diet (Hindustan Lever Limited) and water ad libitum, After 55 days of drug administration the mating exposure test was performed in control as well as in treated groups. Rats of both groups were cohabitated with normal adult proestrous females for fertility test in the ratio of 2. Successful mating was confirmed the following morning by presence of sperms in the vaginal smear. The inseminated females were separated and number of litters delivered were recorded/11/. Proven fertile male rats were taken and divided into two groups of 10 rats each Group Rats received 0.5 ml olive oil for 60 days, Group II--> Rats treated with compound (1) dispersed in 0.5 ml olive oil, 6.5 mg/rat/day for 60 days.
On day 61 st animals of both groups were sacrificed using ether anesthesia and their reproductive organs All the above derivatives are either coloured viscous liquids or solids and highly soluble in organic solvents like benzene, chloroform etc. These derivatives are found to be monomeric in benzene solution.
Spectroscopic studies Infrared spectra The absence of vSH mode (-2540 cm1) and the presence of vNH mode in the range 3230-3340 cm 1

H NMR Spectra
A comparative study of the H NMR spectra of aluminum derivatives (Table-ll  )-7Al NMR spectra of these complexes (1-4) ( Table-II  It is clear from the spectral data that these ligands behave as monofunctional bidentate moiety in these derivatives. These results are in contrast to the earlier reports in which these ligands are reported to behave as bifunctional tridentate /5,6/. The monofunctional nature of the ligands in these aluminium complexes is confirmed as there is no evidence of any enolisation in these ligands and their bidentate behaviour may be easily established on the bases of maximum coordination number of aluminium, which is six, and the monofunctional nature of the ligands indicates that the enolisation of these ligands does not take place during the complexation, which is also clear from the spectroscopic evidences. Antifertility activity of compound AI[SC6H4N:C(CHa)CH2COCH3]3 (1) The body weight of rats after administration of compound (I) did not alter significantly; however the weights of testes (P < 0.001), epididymis (P < 0.001), seminal vesicle (P < 0.01) and ventral prostate (P < 0.001) were reduced significantly (Table IV). The motility of the cauda epididymal sperm as well as concentration of sperm in testes and cauda epididymis was reduced significantly (P < 0.001). The fertility of compound (1) treated rats declined up to 100 % (Table V). Most of the cell types of seminiferous tubule showed significant reduction. The population of primary spermatocytes (preleptotene and pachytene), secondary spermatocytes and step-19 spermatids declined by 56.10 %, 44.42 %, 63.35 % and 64.54 % respectively (Table VI). 200 +/-11.5"* aValues are mean +/-SEM (n 10), ns, non significant * P < 0.01, ** P < 0.001 vs control (student's t-test) The compound (l) treatment brings about a depletion in protein content of testes (P < 0.001), cauda epididymis (P < 0.01), seminal vesicle (P < 0.01) and ventral prostate (P < 0.001). Sialic acid content of the testes (P < 0.001), cauda epididymis (P < 0.01), seminal vesicle (P < 0.01) and ventral prostate (P < 0.001) was depleted in treated rats. Testicular glycogen was decreased to a significant (P < 0.001) degree. Level of cholesterol in testes was elevated whereas seminal vesicular fructose decreased significantly (P < 0.001) in comparison to controls (Table VII). Values are mean +/-SEM (n 10) ** P < 0.001 vs control (student's t-test) ,Synthesis and Characterisation of Some New Aluminium Derivatives qf Schiff Bases Containing N, 0 and S Donor Atoms and the Anti Fertility The significant reduction in testes weight after the treatment with compound (I) can be attributed to the loss of germ cell/20/. It has been observed that the size of testes bears a direct relationship to the number of sperms produced/21/. The marked reduction in sperm count in our experiment also suggests a disturbed testicular and epididymal microenvironment. The treatment with compound (1) exerted a strong inhibitory effect on epididymis, seminal vesicle and ventral prostate. The decrease in weight of accessory sex organs indicates the atrophy of glandular tissue and reduction in secretory ability, thus reflecting the decreased level of testosterone/22/. It has been reported earlier that there is a high correlation of reproductive capacity with the mass of accessory sex organs/23/. Decreased seminiferous tubular diameter reflects tubular shrinkage which may occur due to cell death or sloughing of epithelial cells/24/. Sperm motility and density directly correlate with fertility and any change in them may cause infertility/25/. It is also evident that testicular function would be altered by reduced protein content/26/. Reduction of the protein content of the testes is due to loss of germ cell as well as the depletion of membrane protein content/27/.  (1)  aValues are mean + SEM (n 10) ns, non significant ** P < 0.001 vs control (student's t-test) bValues in parentheses are percentage reduction in particular cell types.
The structural integrity of acrosomal membrane is dependent upon sialic acid and due to alteration in its content, the motility and fertilizing ability of sperms may also be effected /2 8, 29/. The significant decrease in glycogen content after the treatment with compound (1) possibly could be due to inhibition of glycolysis during spermatogenesis as has been observed by Bone et al. /30/ in ornidazole treated rats. Glycolytic metabolism of spermatozoa is also hampered by reducing level of fructose resulting in abnormal sperm function which may ultimately lead to complete male sterility/31/.