Spectrophotometric, Fourier transform infrared spectroscopic and theoretical studies of the charge-transfer complexes between methyldopa [(S)-2 amino-3-(3,4-dihydroxyphenyl)-2-methyl propanoic acid] and the acceptors (chloranilic acid, o-chloranil and dichlorodicyanobenzoquinone) in acetonitrile and their thermodynamic properties.

Methyldopa is a much used antihypertensive drug. It is the subject matter of study mostly for the determination and estimation of methyldopa in pharmaceutical properties. These considerations led us to study the charge-transfer interactions between methyldopa, a centrally acting antihypertensive agent of limited use with the known acceptors like o-chloranil (o-ClN), chloranilic acid (ClA) and dichlorodicyanobenzoquinone (DDQ). Methyldopa (MDP) formed beautifully colored complexes (having absorption maxima at 581 nm and 368 nm; 519 nm; 583.5 nm, 547 nm and 346 nm, respectively) with the acceptors mentioned before. The physico-chemical properties of the complexes were studied using UV-visible spectrophotometry and FTIR measurements. The composition, the accurate association constants and thermodynamics of the complexes were determined spectrophotometrically. Attempts were made to interpret the thermodynamics of complexes in terms of I(D)(V), E(A)(V) and hν(CT). Solid CT complexes between MDP+o-ClN, MDP+ClA and MDP+DDQ were prepared and FTIR spectra of the complexes were studied. The energies hν(CT) of the charge-transfer complexes and vertical ionization potential I(D)(V) of methyldopa were compared with the theoretical values of hν(CT) obtained from HOMO and LUMO of the donors and acceptors calculated using Density Function Theory utilizing different basis sets. The agreement between the results can be regarded to be reasonable. Oscillator strengths and dipole strengths of the complexes were determined theoretically and experimentally and the limitations of the calculations were outlined.

Spectrophotometric studies on the thermodynamic properties of charge-transfer complexes between m-DNB (1,3-dinitrobenzene) with aliphatic amines in DMSO and determination of the vertical electron affinity of m-DNB.

1,3-Dinitrobenzene formed colored 1:1 complexes with aliphatic amines (chromogenic agents) like isopropylamine,ethylenediamine, tetraethylenepentamine and bis(3-aminopropyl)amine in DMSO having absorption maxima at 563 nm, 584 nm, 580.5 nm and 555 nm respectively. The complexes were stable for more than 24 h. The accurate association constants KAD and other thermodynamic parameters were determined with D and A usually in stoichiometric ratios. But in case of m-DNB and bis(3-aminopropyl)amine, the association constants KAD and the thermodynamic parameters were also determined using Benesi-Hildebrand equation to show the variations of KAD under different conditions. ΔG° values were found to be negative in all cases resulting from exothermic enthalpy changes and favourable entropy changes. The energies of transition for the CT complexes hνCT found experimentally were considerably different from the energies of transition (from HOMO of donor to LUMO of acceptor) calculated using AM1 but the differences were considerably reduced using DFT calculations. The vertical electron affinity of m-DNB was calculated using the method suggested by Mulliken. However, no FTIR measurements of the complexes could be made due to experimental limitations.

A preliminary investigation into the use of FTIR microscopy as a probe for the identification of bullet entrance holes and the distance of firing.

FTIR spectroscopy has been found to be a valuable probe for the analysis of the distribution of organic components such as nitroglycerine in gunshot residues deposited on and around the bullet entrance hole of the clothing of the victim in short range firearm discharges. The method can be utilised for the detection and estimation of organic gunshot residues (OGSR) on the hands and clothing of the shooter. The method is specific and sensitive and is likely to be free from interference from, for example, environmental pollution. The method shows potential to be utilised as a suitable alternative to the widely used SEM-EDX analysis of the total contents of lead, barium and antimony present in the gunshot deposits around the bullet entrance hole. The method was tested in the laboratory and the results were validated by Gas Chromatography-Mass Spectrometry (GC-MS).

Absorption spectroscopic and FTIR studies on EDA complexes between TNT (2,4,6-trinitrotoluene) with amines in DMSO and determination of the vertical electron affinity of TNT.

TNT (2,4,6-trinitrotoluene) formed deep red 1:1 CT complexes with chromogenic agents like isopropylamine, ethylenediamine, bis(3-aminopropyl)amine and tetraethylenepentamine in DMSO. The complexes were also observed in solvents like methanol, acetone, etc. when the amines were present in large excess. The isopropylamine, complex showed three absorption peaks (at 378, 532 and 629 nm) whereas higher amines showed four peaks (at 370, 463, 532 and 629 nm). The peak at 463 nm vanished rapidly. The peak of the complexes near 530 nm required about 8-10 min to develop and the complexes were stable for about an hour but the peak slowly shifted towards 500 nm and the complexes were found to be stable for more than 24 h. The evidence of complex formation was obtained from distinct spots in HPTLC plates and from the shifts in frequencies and formation of new peaks in FTIR spectra. The peaks near 460 nm (transient) and 530 nm may be due to Janovsky reaction but could not be established. The extinction coefficients of the complexes were determined directly which enabled the accurate determination of the association constants KDA with TNT and amines in stoichiometric ratios. The results were verified using iterative method. The quantification of TNT was made using epsilon value of the complex with ethylenediamine. The vertical electron affinity (EA) of TNT was calculated using the method suggested by Mulliken.

Qualitative and quantitative analysis of seized street drug samples and identification of source.

In this paper we describe the identification of constituents of the illicit drugs seized from different regions of eastern India by GC-MS. The constituents were identified to be heroin, acetyl morphine, morphine and acetyl codeine. Quantitative estimation of the constituents were made by GC-MS and HPTLC. In view of non-availability of the authentic samples of drugs of different origin, nothing positive can be said about the origin of illicit drug samples. The possibility of isotopic substitution, an important method for identification of source, was examined from the comparison of the intensity of different (ion) peaks 369 (heroin, m/z=369), 370, 371 and 372 using selective ion monitoring mode. No isotopic substitution in the constituents was observed. Attempts were made to identify the source of the illicit samples from heroin/acetylcodeine ratios in the way described in the literature.

Synthesis of chloro and bromo substituted 5-(indan-1'-yl)tetrazoles and 5-(indan-1'-yl)methyltetrazoles as possible analgesic agents.

Chloro and bromo substututed indanyl tetrazoles (compounds 5a, b) and indanyl methyltetrazoles (compounds 5c, 5d) have been synthesized from their respective acids through amide and nitrile routes, and characterized. The title compounds (5a, 5b, 5c and 5d) were subjected for their analgesic activity in the acetic acid induced writhing test on albino mice. The significant (p < 0.001) analgesic activity, exhibited by the compound 5b at a dose of 50 mg/kg body weight, was comparable to that of phenylbutazone and indometacin at a dose of 100 and 50 mg/kg body weight respectively. The effect of substitution at the benzenoid part of the indan nucleus and chain lenght on analgesic activity was in the following order: bromine > chlorine and tetrazole > methyltetrazole.

Purification and partial characterization of a haemorrhagin (VRH-1) from Vipera russelli russelli venom.

A haemorrhagic toxin (VRH-1) has been purified to homogeneity from Vipera russelli russelli venom by subjecting it to chromatography twice successively on CM-Sephadex C-50. It is a protein of mol. wt 22,000 and contains one mole of Mg2+. Intradermal administration of this haemorrhagin in mice resulted in severe lung haemorrhage but produced little haemorrhage in skin. This apparent organ preference led us to develop a new haemorrhage assay method utilizing dye diffusion from lung in vitro. Proteolytic activity of VRH-1 was demonstrated using dimethylcasein as substrate following quantitation by reaction with trinitrobenzoyl sulfonic acid. Both haemorrhagic and proteolytic activities of VRH-1 were inhibited by serine protease inhibitors like phenylmethyl sulfonyl fluoride and chymostatin, but metal chelators had no effect. Lung haemorrhage is unlikely to be a direct reflection of a high local concentration of VRH-1. The administration of supernatant generated by incubation of chopped liver from untreated mouse and VRH-1 (in subhaemorrhagic dose) results in severe lung haemorrhage. This raises the possibility that VRH-1 leads to the formation of intermediate(s) which causes the haemorrhage.

Prostaglandin biosynthesis inhibitory activity of some indian-1 acids in relation to their anti-inflammatory activity and ulcerogenic potency.

Continuation of our work towards development of some newer non-steroidal anti-inflammatory agents led us to some substituted indian-1-acids with low ulcerogenic liability. Prostaglandin biosynthesis inhibitory activity of these indian acids and their acid dissociation constants were evaluated in view of their activity profile.

Evaluation of some newer non-steroidal anti-inflammatory indan-1-acids in various biological systems.

In line of the effort towards development of some newer indanyl non-steroidal anti-inflammatory agents and providing comprehensive SAR among this class of compounds some significantly active derivatives with low ulcerogenic potential were identified. Dealing with various long chain and branched chain compounds among this series, 3-(5, 6-dimethoxy indan-1-yl) propionic acid, 2-(5, 6-dimethoxy indan-1-yl) propionic acid and 3-(6-methoxy indan-1-yl) propionic acid were observed to have encouraging biological activity. Screening in various animal models of inflammation suggests their longer duration of action and lower ulcerogenic liability.

Detection and distribution of scorpion (Heterometrus bengalensis) venom in rabbit tissues by enzyme-linked immunosorbent assay (ELISA).

ELISA was carried out to detect distribution of scorpion venom in experimental animal tissues. The venom content of different tissues was in the order, liver greater than kidney greater than spleen greater than lung greater than heart greater than diaphragm greater than brain. Tissue distribution of venom antigen in the envenomental subject by ELISA will provide a better approach for serotherapy.

Lethal and haemorrhagic activity of Russell's Viper venom--neutralization by polyvalent, monovalent and toxoid antiserum.

Lethal and haemorrhagic activity of Russell's Viper venom was compared against polyvalent bivalent commercial antiserum and monovalent antiserum raised in rabbit. Formaldehyde-detoxified venom offered 7-fold protection against lethal activity and 12.5-fold against haemorrhagic activity of the venom. Whole venom and formaldehyde-detoxified venom along with Freund's complete adjuvant, injected in rabbits produced high titre antiserum. Amongst all the six antiserum tested, the monovalent antiserum raised in rabbit, showed maximum precipitating bands in immunogeldiffusion and immunoelectrophoresis. The toxoid-antiserum offered maximum protection against the venom-induced lethality and the monovalent antiserum offered maximum protection against haemorrhagic activity.

Histamine, 5-HT & hyaluronidase in the venom of the scorpion Lychas laevifrons (Pock).

Factors involved in the pathophysiological changes such as severe pain, burning sensation, redness, swelling and edema in case of the scorpion L. laevifrons were investigated. The presence of pain-producing autacoids histamine 2.1 +/- 0.18 micrograms/mg and 5-HT 0.23 +/- 0.1 micrograms/ml was confirmed by thin layer chromatography and bioassay. Histamine releasing substance was detected in vitro in the chopped guineapig lung. Venom also contained hyaluronidase 5 x 10(-4) N-acetyl-D-glucosamine released/h/mg, which facilitates spread of the toxic principles in the tissues. It is concluded that histamine, 5-HT, histamine-releasing factor and hyaluronidase are partly involved in the pathophysiological changes induced by the venom. It is suggested that mepyramine and cyproheptadine may prove useful in the management of scorpion envenomation.

Isolation, partial purification and pharmacodynamics of a slow contractile substance in the venom sac extract of the wasp Vespa cincta Fabr.

The venom of V. cincta contains acetylcholine (ACh), histamine and 5-hydroxytryptamine (5-HT). Blockers of these agonists did not block completely the hypotensive and smooth muscle contractile activity of venom. On smooth muscle, there was a residual slow contraction. The active substance which produced this slow contraction was separated by solvent extraction, gel filtration and TLC. The purified material (which has been provisionally designated "Vecikinin") lowered cat, rat and guinea pig blood pressure, increased amplitude of cardiac contraction, and increased capillary permeability. Vecikinin contracted several smooth muscle preparations (rat uterus, rat ascending colon, guinea pig ileum, guinea pig colon and rat ileum), while relaxing rat duodenum. Its contractile activity was not lost on boiling, but acid or alkali-boiling reduced its contractile activity. It was inactivated on incubation with chymotrypsin and carboxypeptidase but not with trypsin, pepsin or leucine aminopeptidase. It is a peptide, appears to be of low molecular weight, and could be distinguished from substance P, angiotensin, bradykinin and hornet or wasp kinin.

Isolation and purification of a lethal scorpion toxin with neuromuscular blocking activity.

Toxin-L a lethal neuromuscular blocking agent was isolated from the venom of the scorpion, Lychas laevifrons (Pocock), by the CM-cellulose ion-exchange chromatography. It was a homogenous, thermolabile and low molecular weight protein. The toxin produced irreversible blockade of indirect stimulation induced twitch responses on innervated rat phrenic nerve-diaphragm and chick biventer cervicis preparation. The toxin did not produce any contractile response on toad rectus abdominis muscle preparation. On chronically denervated rat diaphragm, the toxin failed to alter the responses induced by direct stimulation, exogenous acetylcholine, potassium chloride and caffeine. Acetylcholine and carbachol induced contractions on isolated chick biventer cervicis remained unaltered by the toxin. Neostigmine failed to alter toxin induced neuromuscular blockade on innervated rat diaphragm. The toxin released a significant amount of acetylcholine from innervated rat diaphragm. It may be concluded that the toxin acts presynaptically through the release of acetylcholine, thereby producing neuromuscular blockade.

Studies on non-steroidal anti-inflammatory and related biological activities of 5-(indan-1'-yl)tetrazoles and their intermediates.

Anti-inflammatory, analgesic and anti-pyretic activities of three new 5-(Indan-1'-yl)tetrazoles and anti-inflammatory activity of corresponding carboxamides were compared to those of standard drugs, phenylbutazone and aspirin. The results indicated 5-(Indan-1'-yl)tetrazole as the most promising compound in chronic anti-inflammatory and anti-pyretic tests.

Isolation, purification and immunological evaluation of toxin Hb from scorpion Heterometrus bengalensis (C.L. Koch) venom.

Toxin-Hb, a lethal toxic antigenic protein, isolated from the venom of H. bengalensis by CM-cellulose ion-exchange chromatography was a heat labile basic protein with a molecular weight of 10 kDa. It produced irreversible blockade on the isolated rat phrenic nerve diaphragm and chick biventer cervicis. LD50 of toxin Hb was 0.48 mg/kg (iv) in mice. Antiserum was raised in mice by hyperimmunization against toxin Hb. Antitoxin Hb antiserum was immunologically potent as revealed by immunogel-diffusion and immunoelectrophoresis. Five fold protection against the lethal action of toxin Hb was achieved by the antiserum. It also effectively antagonised toxin Hb induced neuromuscular blockade on isolated rat phrenic nerve diaphragm and chick biventer cervicis preparations.

Pharmacology of scorpion (Lychas laevifrons Pock) venom with special reference to its neuromuscular activity.

L. laevifrons venom caused irreversible blockade of electrically induced twitch responses on phrenic nerve diaphragm and chick biventer cervicis preparation. The venom lowered cat blood pressure, caused a brief cardiac arrest and increased cutaneous capillary permeability. It contracted several smooth muscle preparations. The quick contraction produced on guinea pig ileum was partly antagonized by mepyramine and completely by methysergide. The residual slow contraction was antagonized by SC 19220, a prostaglandin blocker. Haemolysis was not produced by the venom on human RBC. LD50 of crude venom in mice was 13.8 mg/kg (iv).

Comparative detoxification and protection of scorpion (Heterometrus bengalensis) venom by toxoid antiserum.

Comparative detoxification of scorpion venom by using different chemical agents was investigated. Detoxification by formalin showed the best optimum detoxifying agent. The formalin treatment resulted in 2.3% protein loss with 6-fold detoxification. This formal toxoid was immunogenic in rabbit giving high neutralizing antibodies as revealed from indirect haemagglutination test. Toxoid antiserum protected mice against the lethal action of venom. It also effectively antagonized the smooth muscle contractile response of venom, and venom-induced neuromuscular paralysis. This toxoid antiserum also protected the venom-induced cardiac arrest. The possibility of using this formal toxoid for antisera production and immunization for therapeutic use needs to be explored.

Immunological cross reactivity & paraspecificity of the scorpion Heterometrus bengalensis antivenom.

Immunological cross-reactivity and paraspecificity of scorpion H. bengalensis antivenom were studied to find out the intergeneric therapeutic relationship between the venom of other scorpions in West Bengal Buthus tamulus, Lychas laevifrons and Heterometrus swammerdami. Of these scorpions, Buthus tamulus and Lychas laevifrons failed to show any cross reactivity. However, H. swammerdami venom showed cross-reactivity with H. bengalensis antiserum as revealed from immunogeldiffusion and immunoelectrophoresis. This antiserum protected H. swammerdami venom-induced lethality in mice, blocked the contractile response in smooth muscles and antagonised the venom-induced neuromuscular blockade in rat phrenic nerve diaphragm and chick biventer cervicis.

Pharmacological and immunological evaluation of scorpion (Heterometrus bengalensis) venom antiserum.

An antiserum was prepared for the first time against the venom of a common scorpion, H. bengalensis, by hyperimmunization of rabbit. This antiserum showed positive precipitin bands in immunogeldiffusion and immunoelectrophoresis. The serum showed a high titre value tested by indirect haemagglutination test. The antiserum developed in rabbit protected mice against the lethal action of the venom. Smooth muscle contractile response of venom on guinea pig ileum, and rat uterus was antagonized by the antiserum. This antiserum effectively antagonized the venom induced neuromuscular paralysis tested on rat phrenic nerve diaphragm and chick biventer cervices. Antiserum also protected the venom-induced cardiac arrest tested on isolated guineapig heart and auricle preparations.