Characterization of novel natural cellulose fiber from Ficus macrocarpa bark for lightweight structural composite application and its effect on chemical treatment.

This study investigates Ficus Macrocarpa tree bark fibers (FMB) as a sustainable alternative reinforcement for polymer composites. The Industrial Revolution marked the evolution of polymer composites with synthetic material reinforcement, leading to environmental concerns. Natural fibers have recently gained prominence as efficient alternatives for polymer composites. Despite numerous natural fibers being considered, ensuring a sustainable raw material source remains crucial. In this study, fibers were extracted from FMB and subjected to alkali treatment to evaluate their impact on physical, chemical, and thermal properties. Initially, the extracted fibers measured 253.80 ± 15 µm in diameter, reduced to 223.27 ± 12 µm post-alkali treatment. Chemical analysis showed an increase in cellulose content to 59.7 wt%, a 23.34 % improvement over untreated fibers (48.4 wt%). The crystalline index for untreated and treated fibers measured 80.20 % and 84.75 %, respectively, with no noticeable changes in the cellulose phase. Additionally, the crystalline size increased to 3.21 nm. Thermogravimetric analysis demonstrated enhanced stability of treated fibers up to 378.87 °C, while the kinetic activation energy remained constant at 64.76 kJ/mol for both the treated and the untreated fibers. The alkali treatment further improved surface roughness to 39.26, confirmed by scanning electron microscopic images. These findings highlight the potential of cellulose fibers from Ficus Macrocarpa bark as a sustainable and environmentally friendly replacement for synthetic fibers in polymer composites. The enhanced physical properties and excellent thermal stability make them a promising choice for eco-conscious materials.

Generation Mean Analysis Reveals the Predominant Gene Effects for Grain Iron and Zinc Contents in Pearl Millet.

Pearl millet [Pennisetum glaucum (L.) R. Br.] is a climate-resilient dryland cereal that has been identified as a potential staple food crop that can contribute to alleviating micronutrient malnutrition, particularly with respect to grain iron (Fe) and zinc (Zn) contents, in Sub-Saharan Africa and India. In this regard, an understanding of the inheritance pattern of genes involved in Fe and Zn contents is vital for devising appropriate breeding methods to genetically enhance their levels in grains. In this study, we aimed to determine the genetic effects underlying such inheritance and their interactions based on the generation mean analyses. Four experimental crosses and their six generations (P1, P2, F1, BCP1, BCP2, and F2) were independently evaluated in a compact family block design in 2017 rainy and 2018 summer seasons. ANOVA revealed highly significant mean squares (p < 0.01) among different generations for grain Fe and Zn contents. Six-parameter generation mean analyses revealed a predominance of additive genetic effect and a significant (p < 0.05) additive × dominant interaction for the grain Fe content. The additive genetic effect for the grain Zn content was also highly significant (p < 0.01). However, interaction effects contributed minimally with respect to most of the crosses for the grain Zn content and hence we assume that a simple digenic inheritance pattern holds true for it. Furthermore, we established that narrow-sense heritability was high for the grain Fe content (>61.78%), whereas it was low to moderate for the grain Zn content (30.60-59.04%). The lack of superior parent heterosis coupled with non-significant inbreeding depression for Fe and Zn contents in grains further confirmed the predominance of an additive genetic effect. These findings will contribute to strategizing a comprehensive breeding method to exploit the available variability of grain Fe and Zn contents for the development of biofortified hybrids of pearl millet.

Ambient assisted living technologies for older adults with cognitive and physical impairments: a review.

The global number of people over the age of 60 years is expected to increase from 970 million to 2.1 billion in 2050 and 3.1 billion in 2100. About 80% of the aging population will be in the developing countries. Aging population may suffer from various physical, cognitive, and social problems, due to aging process such as impairment of physical related functions (decreased mobility and walking speed, falls, frailty, decreased walking speed, difficulties in basic, and instrumental activities of daily living), cognitive related functions (memory-related issues), sensory functions (hearing loss, cataracts and refractive errors, presbyopia, decreased vestibular function), behavioural and psychological disorders, social isolation issues, and poor quality of life. Over the period of the last few decades, emerging technologies such as internet of things (IoT), artificial intelligence (AI), sensors, cloud computing, wireless communication technologies, and assistive robotics have given the vision to develop various ambient or active assisted living (AAL) approaches for supporting an elderly people to live safely and independently in their living environment and participate in their daily and community activities, as well as supporting them to maintain their physical, mental health, and quality of their life. The aim of this paper is to review the use of Ambient or Active Assisted Living for older adults with physical, cognitive impairments, and their social participation.

Quantification, distribution and diversity of ESBL/AmpC-producing Escherichia coli on freshly slaughtered pig carcasses.

This study quantified cefotaxime-resistant E. coli (CREC) on nine different carcass areas of 104 freshly slaughtered pig carcasses. In 49% [95% confidence interval (95% CI): 29-69%] of the carcasses CREC could be isolated and enumerated (using Tryptone Bile Agar with X-Glucuronide supplemented with 1 mg/L cefotaxime). Proportions of positive samples varied between carcass areas from 1% [95% CI: 0-10%] (loin) to 23% [95% CI: 10-44%] (head). Maximum concentrations on positive samples ranged between -0.6 log10 CFU/cm2 (loin, elbow before evisceration) and 1.7 log10 CFU/cm2 (head). The head was significantly more frequently contaminated than the loin (p = 0.027) and ham (3% [95% CI: 1-15%]). The foreleg was significantly more frequently contaminated (20% [95% CI: 13-30%]) than the ham. Combination disk diffusion assays revealed that 81% of the CREC isolates were extended-spectrum beta-lactamases (ESBL) producers, 13% were AmpC cephalosporinases (AmpC) producers and 2% ESBL and AmpC co-producers. Genotyping denoted blaCTX-M-gr1 (63%) and blaTEM (40%) as most present antibiotic resistance genes. Multiple gene combinations in one isolate and multiple combinations of genotypes and phenotypes among isolates of one sample were observed. These quantitative data can be used for intervention strategies to lower human exposure to CREC.

High abundance and diversity of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli in faeces and tonsils of pigs at slaughter.

This cross-sectional study investigates the abundance of cefotaxime-resistant Escherichia coli (CREC) in the faeces and tonsils of 96 pigs during slaughter. Moreover, different isolates from a selected number of pigs were tested to study the diversity of blaESBL genes within E. coli isolates from one pig. Cefotaxime-resistant bacteria (based on enumeration results on MacConkey agar supplemented with 1mg/L cefotaxime) were found in the faeces of 77 pigs (80%; 95% CI: 70-87%) and the tonsils of 91 pigs (95%; 95% CI: 88%-98%). Cefotaxime-resistant E. coli (based on enumeration results on Tryptone Bile X-glucuronide agar supplemented with 1mg/L cefotaxime) were detected in 72 faecal samples (75%; 95% CI: 64-83%) and 45 tonsil samples (47%; 95% CI: 35-59%), in numbers up to 5.5 and 5.6log10 CFU/g, respectively. On average, around 1/10,000 E. coli in both faeces and tonsils were cefotaxime-resistant, though large variations were observed between pigs. Within one sample, CREC isolates with up to five different combinations of ESBL genes were observed. In three out of 16 faecal samples and six out of 14 tonsil samples, only one ESBL gene profile was found. The high numbers of CREC that are occasionally found in the faeces and tonsils of pigs during slaughter may represent an important source of contamination of carcasses and subsequently pork.

Albizia lebbeck seed methanolic extract as a complementary therapy to manage local toxicity of Echis carinatus venom in a murine model.

CONTEXT AND OBJECTIVE: Viperid venom-induced chronic local-toxicity continues even after anti-snake venom treatment. Therefore, traditional antidote Albizia lebbeck L. (Fabaceae) seed extract was tested against Echis carinatus S. (Viperidae) venom (ECV)-induced local toxicity to evaluate its complementary remedy. MATERIALS AND METHODS: Soxhlet extraction of A. lebbeck seeds was performed with the increasing polarity of solvents (n-hexane to water); the extract was screened for phytochemicals (alkaloids, anthraquinones, flavonoids, glycosides, phenolics, saponins, steroids and tannins). In preliminary in vitro analysis, A. lebbeck methanolic extract (ALME) demonstrated significant inhibition of ECV proteases, the major enzyme-toxin responsible for local- toxicity. Therefore, in vitro neutralizing potential of ALME was further evaluated against hyaluronidases and phospholipase A2 (1:1-1:100 w/w). In addition, alleviation of ECV induced characteristic local- toxicity [haemorrhage (i.d.) and myotoxicity (i.m.)] was determined in mice. RESULTS: ALME contained high concentrations of phenolics and flavonoids and demonstrated significant in vitro inhibition of ECV protease (IC50 = 36.32 µg, p < 0.0001) and hyaluronidase (IC50 = 91.95 µg, p < 0.0001) at 1:100 w/w. ALME significantly neutralized ECV induced haemorrhage (ED50 = 26.37 µg, p < 0.0001) and myotoxicity by significantly reducing serum creatinine kinase (ED50 = 37.5 µg, p < 0.0001) and lactate dehydrogenase (ED50 = 31.44 µg, p = 0.0021) levels at 1:50 w/w. DISCUSSION AND CONCLUSION: ALME demonstrated significant neutralization of ECV enzymes that contribute in local tissue damage and haemostatic alterations. The study scientifically supports the anecdotal use of A. lebbeck in complementary medicine and identifies ALME as principle fraction responsible for antivenom properties.

Virtual analysis of structurally diverse synthetic analogs as inhibitors of snake venom secretory phospholipase A2.

Due to the toxic pathophysiological role of snake venom phospholipase A2 (PLA2 ), its compelling limitations to anti-venom therapy in humans and the need for alternative therapy foster considerable pharmacological interest towards search of PLA2 specific inhibitors. In this study, an integrated approach involving homology modeling, molecular dynamics and molecular docking studies on VRV-PL-V (Vipera russellii venom phospholipase A2 fraction-V) belonging to Group II-B secretory PLA2 from Daboia russelli pulchella is carried out in order to study the structure-based inhibitor design. The accuracy of the model was validated using multiple computational approaches. The molecular docking study of this protein was undertaken using different classes of experimentally proven, structurally diverse synthetic inhibitors of secretory PLA2 whose selection is based on IC50 value that ranges from 25 µM to 100 µM. Estimation of protein-ligand contacts by docking analysis sheds light on the importance of His 47 and Asp 48 within the VRV-PL-V binding pocket as key residue for hydrogen bond interaction with ligands. Our virtual analysis revealed that compounds with different scaffold binds to the same active site region. ADME analysis was also further performed to filter and identify the best potential specific inhibitor against VRV-PL-V. Additionally, the e-pharmacophore was generated for the best potential specific inhibitor against VRV-PL-V and reported here. The present study should therefore play a guiding role in the experimental design of VRV-PL-V inhibitors that may provide better therapeutic molecular models for PLA2 recognition and anti-ophidian activity.

Biochemical and pharmacological characterization of three toxic phospholipase A2s from Daboia russelii snake venom.

Three isoenzymes of phospholipase A2 (PLA2), VRV-PL-IIIc, VRV-PL-VII, and VRV-PL-IX were isolated from Daboia russelii snake venom. The venom, upon gel filtration on Sephadex G-75 column, resolved into six peaks (DRG75 I-VI). The VRV-PL-IIIc was purified by subjecting DRG75II to homogeneity by rechromatography in the presence of 8M urea on Sephadex G-75 column. The other two isoenzymes VRV-PL-VII and VRV-PL-IX were purified by subjecting DRG75III to ion exchange chromatography on CM-Sephadex C-25 column. Mol wt. for the three PLA2s, VRV-PL-IIIc, VRV-PL-VII, and VRV-PL-IX are 13.003kDa, 13.100kDa and 12.531kDa respectively. The VRV-PL-IIIc is not lethal to mice up to 14mg/kg body weight but it affects blood sinusoids and causes necrosis of the hepatocytes in liver. It causes hemorrhage in kidney and shrinkage of renal corpuscles and renal tubules. The LD50s for VRV-PL-VII and VRV-PL-IX are 7 and 7.5mg/kg body weight respectively. They induced neurotoxic symptoms similar to VRV-PL-V. All the three PLA2s are anticoagulant and induced varying degree of edema in the foot pads of mice. VRV-PL-V and VRV-PL-VII are shown to act as pre and post synaptic toxins, while VRV-PL-IX acts as presynaptic toxin. This is evident from experiments conducted on cultured hippocampal neurons by patch clamp electrophysiology.

Evidence for existence of venom 5' nucleotidase in multiple forms through inhibition of concanavalin A.

Pharmacologically active 5' nucleotidase is a ubiquitously distributed enzyme in snake venoms. In this study the effect of concanavalin A (Con-A) on different snake venoms 5' nucleotidase activity is tested in order to know the protein nature which will ultimately help in purification of the enzyme with high yield. Con-A inhibited Naja naja, Naja kauthia, Naja melanoleuca, Naja naja sputatrix, Agistrodon halys blomhoffii, Bothrops asper and Oxyranus scutellas venom 5' nucleotidase activity at different concentrations. This indicates the presence of glycopart in the protein, thus glycoprotein in nature. Vipera russellii, Vipera plaestenae, Agistrodon contratrix, Bitis orientis, Echis carinatus and Trimeresures malabaricus was not inhibited by Con-A, indicating absence of glycopart in the protein. This study for the first time shows existence of 5' nucleotidase in multimeric forms.

INN-toxin, a highly lethal peptide from the venom of Indian cobra (Naja naja) venom-Isolation, characterization and pharmacological actions.

A novel toxic polypeptide, INN-toxin, is purified from the venom of Naja naja using combination of gel-permeation and ion-exchange chromatography. It has a molecular mass of 6951.6Da as determined by MALDI-TOF/MS and the N-terminal sequence of LKXNKLVPLF. It showed both neurotoxic as well as cytotoxic activities. INN-toxin is lethal to mice with a LD(50) of 1.2mg/kg body weight. IgY raised in chicks against basic peptide pool neutralized the toxicity of INN-toxin. INN-toxin did not inhibit cholinesterase activity. It is toxic to Ehrlich ascites tumor (EAT) cells, but it is not toxic to leukocyte culture. The toxin appears to be specific in its mode of action. Interaction of N-bromosuccinamide (NBS) with the peptide resulted in the modification of tryptophan residues and loss of lethal toxicity of INN-toxin.

Isolation and characterization of "Reprotoxin", a novel protein complex from Daboia russelii snake venom.

In snake venoms, non-covalent protein-protein interaction leads to protein complexes with synergistic and, at times, distinct pharmacological activities. Here we describe a new protein complex containing phospholipaseA(2) (PLA(2)), protease, and a trypsin inhibitor. It is isolated from the venom of Daboia russelii by gel permeation chromatography, on a Sephadex G-75 column. This 44.6 kDa complex exhibits only phospholipase A(2) activity. In the presence of 8M urea it is well resolved into protease (29.1 kDa), PLA(2) (13 kDa), and trypsin inhibitor (6.5 kDa) peaks. The complex showed an LD(50) of 5.06 mg/kg body weight in mice. It inhibited the frequency of spontaneous release of neurotransmitter in hippocampal neurons. It also caused peritoneal bleeding, and edema in the mouse foot pads. Interestingly, the complex caused degeneration of both the germ cells and the mouse Leydig cells of mouse testis. A significant reduction in both the diameter of the seminiferous tubules and height of the seminiferous epithelia were observed following intraperitoneal injection of the sub-lethal dose (3 mg/kg body weight). This effect of the toxin is supported by the increase in the activities of acid and alkaline phosphatases and the nitric oxide content in the testes, and a decrease in the ATPase activity. Because of its potent organ atrophic effects on the reproductive organs, the toxin is named "Reprotoxin". This is the first report demonstrating toxicity to the reproductive system by a toxin isolated from snake venom.

Papaya shoot tip associated endophytic bacteria isolated from in vitro cultures and host-endophyte interaction in vitro and in vivo.

Fourteen distinct bacterial clones were isolated from surface-sterilized shoot tips (approximately 1 cm) of papaya (Carica papaya L. 'Surya') planted on Murashige and Skoog (MS)-based papaya culture medium (23/50 nos.) during the 2-4 week period following in vitro culturing. These isolates were ascribed to six Gram-negative genera, namely Pantoea (P. ananatis), Enterobacter (E. cloacae), Brevundimonas (B. aurantiaca), Sphingomonas, Methylobacterium (M. rhodesianum), and Agrobacterium (A. tumefaciens) or two Gram-positive genera, Microbacterium (M. esteraromaticum) and Bacillus (B. benzoevorans) based on 16S rDNA sequence analysis. Pantoea ananatis was the most frequently isolated organism (70% of the cultures) followed by B. benzoevorans (13%), while others were isolated from single stocks. Bacteria-harboring in vitro cultures often showed a single organism. Pantoea, Enterobacter, and Agrobacterium spp. grew actively on MS-based normal papaya medium, while Microbacterium, Brevundimonas, Bacillus, Sphingomonas, and Methylobacterium spp. failed to grow in the absence of host tissue. Supplying MS medium with tissue extract enhanced the growth of all the organisms in a dose-dependent manner, indicating reliance of the endophyte on its host. Inoculation of papaya seeds with the endophytes (20 h at OD550=0.5) led to delayed germination or slow seedling growth initially. However, the inhibition was overcome by 3 months and the seedlings inoculated with Pantoea, Microbacterium, or Sphingomonas spp. displayed significantly better root and shoot growths.

Chemistry and structural evaluation of different phospholipase A2 inhibitors in arachidonic acid pathway mediated inflammation and snake venom toxicity.

PLA2 inhibitors specific to Group I and II PLA2 isoforms are therapeutically important as anti-inflammatory molecules and against venom toxicity. From various natural sources diversified molecules with PLA2 inhibition and concomitant neutralization of inflammatory reactions and venom toxicity were characterized. Using these molecules, lead compounds are generated in several laboratories. Analogues of lead molecules were generated by substituting different types of functional groups in order to obtain a molecule with optimal PLA2 inhibition. The lead molecules characterized as PLA2 inhibitors are indoles, azetidinones, piperazines, isoxazolidines, isoxazolines, diazepinones, acenaphthenes and several substrate analogues. The lead optimization involves relative hydrophobicity and substitution of functional groups, such as electron withdrawing or donating. Many such groups are placed on hydrophobic moiety and their positional bioisosters are characterized. Among these analogue piperazine derivatives on optimization with respect to hydrophobicity and electronegativity showed inhibition at nanomolar levels. Structural analysis of many lead molecules indicated that a PLA2 inhibitor should have both hydrophobic moiety and polar functional groups. Each lead molecule requires optimization in this regard for effective inhibition.

The anti-snake venom properties of Tamarindus indica (leguminosae) seed extract.

In Indian traditional medicine, various plants have been used widely as a remedy for treating snake bites. The aim of this study was to evaluate the effect of Tamarindus indica seed extract on the pharmacological as well as the enzymatic effects induced by V. russelli venom. Tamarind seed extract inhibited the PLA(2), protease, hyaluronidase, l-amino acid oxidase and 5'-nucleotidase enzyme activities of venom in a dose-dependent manner. These are the major hydrolytic enzymes responsible for the early effects of envenomation, such as local tissue damage, inflammation and hypotension. Furthermore, the extract neutralized the degradation of the Bbeta chain of human fibrinogen and indirect hemolysis caused by venom. It was also observed that the extract exerted a moderate effect on the clotting time, prolonging it only to a small extent. Edema, hemorrhage and myotoxic effects including lethality, induced by venom were neutralized significantly when different doses of the extract were preincubated with venom before the assays. On the other hand, animals that received extract 10 min after the injection of venom were protected from venom induced toxicity. Since it inhibits hydrolytic enzymes and pharmacological effects, it may be used as an alternative treatment to serum therapy and, in addition, as a rich source of potential inhibitors of PLA(2), metalloproteinases, serine proteases, hyaluronidases and 5 cent-nucleotidases, the enzymes involved in several physiopathological human and animal diseases.

Synthesis and efficacy of 1-[bis(4-fluorophenyl)-methyl]piperazine derivatives for acetylcholinesterase inhibition, as a stimulant of central cholinergic neurotransmission in Alzheimer's disease.

The cholinergic hypothesis of Alzheimer's disease (AD) has spurred the development of numerous structural classes of compounds with different pharmacological profile aimed at increasing central cholinergic neurotransmission. Thus proving a symptomatic treatment for this disease are cholinomimetics with the pharmacological profile of acetyl cholinesterase (AchE) inhibitors. The novel bioactive 1-[bis(4-fluorophenyl)-methyl]piperazine derivatives were synthesized under mild conditions using different aryl/alkyl halides and heterocyclic alkyl halides with 1-[bis(4-fluorophenyl)-methyl]piperazine in the presence of powdered potassium carbonate in N,N-dimethylformamide. All the synthesized compounds were characterized by spectroscopic techniques, elemental analysis and were screened for their efficacy as AchE inhibitor. Some derivatives in this class showed good inhibition against AchE as compared to neostigmine as standard.

A glycoprotein from a folk medicinal plant, Withania somnifera, inhibits hyaluronidase activity of snake venoms.

Venom hyaluronidases help in rapid spreading of the toxins by destroying the integrity of the extra-cellular matrix of the tissues in the victims. A hyaluronidase inhibitor (WSG) is purified from a folk medicinal plant, Withania somnifera. The glycoprotein inhibited the hyaluronidase activity of cobra (Naja naja) and viper (Daboia russelii) venoms, which was demonstrated by zymogram assay and staining of the skin tissues for differential activity. WSG completely inhibited the activity of the enzyme at a concentration of 1:1 w/w of venom to WSG. Thus we are able to demonstrate that the glycoprotein inhibits hyaluronidase activity of the venoms. External application of the plant extract as an antidote in rural parts of India to snakebite victims appears to have a scientific basis.

Purification of a post-synaptic neurotoxic phospholipase A2 from Naja naja venom and its inhibition by a glycoprotein from Withania somnifera.

A post-synaptic neurotoxic phospholipase A(2) (PLA(2)) has been purified from Indian cobra Naja naja venom. It was associated with a peptide in the venom. The association was disrupted using 8 M urea. It is denoted to be a basic protein by its behavior on both ion exchange chromatography and electrophoresis. It is toxic to mice, LD(50) 1.9 mg/kg body weight (ip). It is proved to be post-synaptic PLA(2) by chymographic experiment using frog nerve-muscle preparation. A glycoprotein, (WSG) was isolated from a folk medicinal plant Withania somnifera. The WSG inhibited the phospholipase A(2) activity of NN-XIa-PLA(2,) isolated from the cobra venom, completely at a mole-to-mole ratio of 1:2 (NN-XIa-PLA(2): WSG) but failed to neutralize the toxicity of the molecule. However, it reduced the toxicity as well as prolonged the death time of the experimental mice approximately 10 times when compared to venom alone. The WSG also inhibited several other PLA(2) isoforms from the venom to varying extent. The interaction of the WSG with the PLA(2) is confirmed by fluorescence quenching and gel-permeation chromatography. Chemical modification of the active histidine residue of PLA(2) using p-brophenacyl bromide resulted in the loss of both catalytic activity as well as neurotoxicity of the molecule. These findings suggest that the venom PLA(2) has multiple sites on it; perhaps some of them are overlapping. Application of the plant extract on snakebite wound confirms the medicinal value associated with the plant.

Novel non-enzymatic toxic peptide of Daboia russelii (Eastern region) venom renders commercial polyvalent antivenom ineffective.

The snake venoms are typically complex mixtures of enzymes and non-enzymatic peptides. Regional variation in the non-enzymatic fraction of Russell's viper venom from three regions of India studied. The eastern, western and southern regional venom upon gel permeation chromatography on sephadex-G-75 column resolved into three peaks. All the three overlapping peaks differ in their lethality and enzymatic potency. Peak III of all the regional venom found to be non-enzymatic, Western and southern regional venom has trypsin inhibitory activity with varying potencies. Interestingly, the peak III of eastern region is devoid of trypsin inhibitory activity. But it is highly lethal with a LD50 0.7 mg/kg body weight and also it exhibited post-synaptic neurotoxicity. On the other hand southern and western regional venom's non-enzymatic peak is non-lethal and did not induce neurotoxic symptoms in experimental model. The antibodies developed against the eastern regional venom cross-reacted with the peaks I and II of other regional venom, but failed to cross-react with the peak III of western and southern regional Russell's viper venom. Commercial anti-venom prepared to neutralize the toxic effects of common poisonous snakes of India, showed positive cross-reaction against peaks I, II and III of all three regional venom tested, except peak III of eastern regional venom. Commercial anti-venom neutralized the lethal toxicity of both western and southern regional Russell's viper venom, and failed to neutralize the lethal effects of eastern regional Russell's viper venom.

Strong myotoxic activity of Trimeresurus malabaricus venom: role of metalloproteases.

Trimeresurus malabaricus is an endemic snake found in the Southern region of Western Ghats section of India along with the more widely distributed species like Naja naja and Daboia russelii. T. malabaricus venom is not lethal when injected (i.p.) up to 20 mg/kg body weight in mice, but causes extensive local tissue degeneration. N. naja and D. russelii are highly toxic (i.p.) with minimum local tissue damage in experimental mice. In this study a comparative analysis of local tissue damage of T. malabaricus venom is made with N. naja and D. russelii snake venoms of the Southern regions of Western Ghats. T. malabaricus venom exhibits caseinolytic activity 16 and 24 times more than N. naja and D. russelii venom. Inhibition studies with specific protease inhibitors reveal that the major proteases belong to metalloproteases. T. malabaricus venom hydrolyses gelatin and induces strong hemorrhagic activity in mice. Both N. naja and D. russelii fail to hydrolyze gelatin even at very high concentration and did not induce any hemorrhagic activity. With D. russelii venom small hemorrhagic spot was observed at the site of injection. The hemorrhagic activity of T. malabaricus venom is completely neutralized by metalloprotease inhibitors and not by serine protease inhibitor. The i.m. injection of T. malabaricus venom causes extensive degradation of muscle tissue within 24 h. The light microscopic observation of muscle tissue showed congestion of blood vessels and hemorrhage at the early stage followed by extensive necrosis of muscle fibers. The elevated levels of serum CK and LDH activity further supported the muscle degeneration. Such pathological symptoms were not seen with N. naja and D. russelii snake venom. The hemorrhagic and the muscle necrosis was completely neutralized by metalloprotease inhibitors and not by serine protease inhibitor strongly suggests that the major toxin component in the T. malabaricus venom is metalloprotease and its activity can be easily neutralized using chelating agents and its use in the first aid as chelation therapy is beneficial.

Inhibition of secretory phospholipase A(2) enzyme by bilirubin: a new role as endogenous anti-inflammatory molecule.

Bilirubin is a powerful antioxidant that suppresses the inflammatory process. However its interaction with proinflammatory PLA(2) enzyme is not known. Inhibition of several secretory phospholipase A(2) (sPLA(2)) enzyme activities by bilirubin was studied using (14)C-oleate labeled Escherichia coli as substrate. Bilirubin inhibits purified sPLA(2) enzyme from Vipera russellii and Naja naja venom and partially purified sPLA(2) enzymes from human ascitic fluid, pleural fluid and normal serum in a dose dependent manner. IC(50) values calculated for these enzymes ranges from 1.75 to 10.5 microM. Inflammatory human sPLA(2) enzymes are more sensitive to inhibition by bilirubin than snake venom sPLA(2)s. Inhibition of sPLA(2) activity by bilirubin is independent of calcium concentration. Increasing substrate concentration (upto 180 nmol) did not relieve the inhibition of sPLA(2) by bilirubin and it is irreversible. Bilirubin quenched the relative fluorescence intensity of sPLA(2) in a dose dependent manner in the same concentration range at which in vitro sPLA(2) inhibition was observed. In the presence of bilirubin, apparent shift in the far UV-CD spectra of sPLA(2) was observed, indicating a direct interaction with the enzyme. Inhibition of sPLA(2) induced mouse paw edema by bilirubin confirms its sPLA(2) inhibitory activity in vivo also. These findings indicate that inhibition of sPLA(2) by bilirubin is mediated by direct interaction with the enzyme and bilirubin may act as an endogenous regulator of sPLA(2) enzyme activity.