Quercitrin neutralizes sPLA2IIa activity, reduces the inflammatory IL-6 level in PC3 cell lines, and exhibits anti-tumor activity in the EAC-bearing mice model.

Human phospholipase A2 group IIa (sPLA2IIa) is an inflammatory enzyme that plays a significant role in tumorigenesis. Inhibiting the sPLA2IIa enzyme with an effective molecule can reduce the inflammatory response and halt cancer progression. The present study evaluates quercitrin, a biflavonoid, for sPLA2IIa inhibition and anticancer activity. Quercitrin inhibited sPLA2IIa activity to a greater extent-at 86.24% ± 1.41 with an IC50 value of 8.77 µM ± 0.9. The nature of sPLA2IIa inhibition was evaluated by increasing calcium concentration from 2.5 to 15 µM and substrate from 20 to 120 nM, which did not alter the level of inhibition. Intrinsic fluorescence and far UV-CD studies confirmed the direct interaction of quercitrin with the sPLA2IIa enzyme. This significantly reduced the sPLA2IIa-induced hemolytic activity and mouse paw edema from 97.32% ± 1.23-16.91% ± 2.03 and 172.87% ± 1.9-118.41% ± 2.53, respectively. As an anticancer activity, quercitrin reduced PC-3 cell viability from 98.66% ± 2.51-18.3% ± 1.52 and significantly decreased the IL-6 level in a dose-dependent manner from 98.35% ± 2.2-37.12% ± 2.4. It increased the mean survival time (MST) of EAC-bearing Swiss albino mice from 30 to 35 days. It obeyed Lipinski's rule of five, suggesting a druggable property. Thus, all the above experimental results were promising and encouraged further investigation into developing quercitrin as a therapeutic drug for both inflammatory diseases and cancers.

Group IIA secreted phospholipase A2 inhibition by elemolic acid as a function of anti-inflammatory activity.

Human group IIA secreted phospholipase A2 (GIIA) is a key enzyme in inflammatory reactions, worsening the condition of several chronic inflammatory diseases. The natural inhibitors of GIIA potentially block the production of inflammatory mediators. In the present study, elemolic acid, a triterpenoid from Boswellia serrata inhibited the GIIA enzyme in a concentration-dependent manner with IC50 value of 5.70 ± 0.02 µM. The mode of GIIA inhibition was studied by increasing the concentration of the substrate from 30 to 120 nM, and calcium from 2.5 to 15 mM, the level of inhibition was not changed. The inhibitor-enzyme interaction was examined by fluorimetry and Circular Dichroism (CD) studies; elemolic acid altered intrinsic fluorescence intensity and shifted far UV- CD spectra of GIIA enzyme, suggesting the direct interaction with GIIA. Elemolic acid neutralized the GIIA mediated indirect hemolytic activity from 94.5 to 9.8% and reduced GIIA induced mouse paw edema from 171.75 to 113.68%. Elemolic acid also reduced the hemorrhagic effect of GIIA along with Vipera russelii neurotoxic non-enzymatic peptide -VNTx-II (VR-HC-I). Thus, the elemolic acid has been proven as a potent inhibitor of GIIA enzyme and modulated the GIIA induced inflammatory response by in situ and in vivo methods.

Synthesis and characterization of Zinc oxide nanoparticles utilizing seed source of Ricinus communis and study of its antioxidant, antifungal and anticancer activity.

ZnO nanoparticles have been synthesized using solution combustion technique and its antioxidant, antifungal, anticancer activity was studied. Ricinus communis plant seed extract used as fuel in synthesis by the solution combustion technique. Powder X-ray diffraction (PXRD) demonstrates the arrangement of a crystalline hexagonal stage (ICDD card number 89-1397) with space aggregate P63mc (186) and cell parameters a = b = 3.253, c = 5.213 Å. The normal crystallite measure is 20 nm which is ascertained by Debye - Scherer's formula. The Purity of the sample and metal to oxygen bond development was affirmed by utilizing Fourier transformation infrared (FTIR) spectroscopy and the particle size and shape was confirmed by HRTEM. Antifungal action of ZnO NPs was studied against Aspergillus and Penicillium by well dispersion strategy. The antifungal activity shows that ZnO NPs constitute as an effective fungicidal agent against both Aspergillus (4 ±â€¯0.5 mm) and Penicillium (3 mm ±â€¯0.4 mm) at 30 µg/mL fixation. ZnO nanoparticles were subjected to antioxidant activity. The objective of the study was to analyze the anticancer property of ZnO NPs on MDA-MB 231 cancer cells. To check the efficacy of the synthesized drug ZnO NPs MTT assay was performed, that determines % viability and/or cytotoxicity. IC50 of ZnO NPs in case of MDA-MB-231 breast cancer was 7.103 µg/mL. Anticancer outcome demonstrates that ZnO NPs is active against in MDA-MB-231 cells.

Neutralization of Inflammation by Inhibiting In vitro and In vivo Secretory Phospholipase A2 by Ethanol Extract of Boerhaavia diffusa L.

BACKGROUND: Inflammation is a normal and necessary prerequisite to healing of the injured tissues. Inflammation contributes to all disease process including immunity, vascular pathology, trauma, sepsis, chemical, and metabolic injuries. The secretory phospholipase A2 (sPLA2) is a key enzyme in the production of pro-inflammatory mediators in chronic inflammatory disorders such as rheumatoid arthritis, coronary heart disease, diabetes, and asthma. The sPLA2 also contribute to neuroinflammatory disorders such as Parkinson's, Alzheimer's, and Crohn's disease. AIMS: The present study aims to investigate the inhibition of human sPLA2 by a popular medicinal herb Boerhaavia diffusa Linn. as a function of anti-inflammatory activity. MATERIALS AND METHODS: The aqueous and different organic solvents extracts of B. diffusa were prepared and evaluated for human synovial fluid, human pleural fluid, as well as Vipera russelli and Naja naja venom sPLA2 enzyme inhibition. RESULTS: Among the extracts, the ethanol extract of B. diffusa (EEBD) showed the highest sPLA2 inhibition and IC50 values ranging from 17.8 to 27.5 µg. Further, antioxidant and lipid peroxidation activities of B. diffusa extract were checked using 2,2-diphenyl-1-picrylhydrazyl radical, thiobarbituric acid, and rat liver homogenate. The antioxidant activity of EEBD was more or less directly proportional to in vitro sPLA2 inhibition. Eventually, the extract was subjected to neutralize sPLA2-induced mouse paw edema and indirect hemolytic activity. The EEBD showed similar potency in both the cases. CONCLUSIONS: The findings suggest that the bioactive molecule/s from the EEBD is/are potentially responsible for the observed in vitro and in vivo sPLA2 inhibition and antioxidant activity. SUMMARY: The present study aims to investigate the inhibition of human sPLA2 by a popular medicinal herb Boerhaavia diffusa Linn. as a function of anti inflammatory activity. Abbreviation Used: EEBD: Ethanolic extract of boerhaavia diffusa, sPLA2: Secretory phospholipase A2, HSF: Human synovial fluid, HPF: Human pleural fluid, VRV-PLA2-V: Vipera russelli phospholipase A2, NN-PLA2-I: Naja naja phospholipase A2.

Malabarase, a serine protease with anticoagulant activity from Trimeresurus malabaricus venom.

In the present study we describe the purification and characterization of Malabarase, a serine protease from Trimeresurus malabaricus venom. Purification was achieved by gel-permeation chromatography on Sephadex G-75 followed by ion-exchange chromatography on CM Sephadex C-25. Homogeneity of Malabarase was confirmed by RP-HPLC. Malabarase is a monomer that migrated as a single protein band on SDS-PAGE under both reducing and non-reducing conditions. The molecular mass of Malabarase was determined to be 23.4 kDa using MALDI-TOF mass spectrometry. Malabarase is the first serine protease purified from T. malabaricus venom and is selective for fibrinogen. Malabarase hydrolyzes Aα and Bß but not γ-chains of fibrinogen similar to the metalloproteases, Malabarin and Trimarin, isolated from the same venom. However, the action of Malabarase on plasma coagulation is opposite than those of Malabarin, Trimarin and the whole venom. Malabarase significantly prolonged plasma coagulation time from 152-341 s; whereas Malabarin, Trimarin, and whole venom, greatly reduce plasma clotting time from 152 to 12, 48, and 14 s, respectively. Malabarase did not show hemorrhagic or myotoxic activity. In contrast, Malabarin, Trimarin and whole venom are highly hemorrhagic and myotoxic. These observations support the specificity of Malabarase towards fibrinogen and its non-toxic nature. In conclusion, Malabarase is a fibrinogen-specific, anti-coagulant, and non-toxic serine protease. Its selective action and non-toxic nature might make it useful for treating thrombotic disorders.

Chemical modification of ascorbic acid and evaluation of its lipophilic derivatives as inhibitors of secretory phospholipase A(2) with anti-inflammatory activity.

The halo 6-fatty acid esters of L-ascorbic acid 3a, 3b and 6-fatty acid esters of L-ascorbic acid 5a-g were achieved from L-ascorbic acid 1. Compounds 3a, 3b and 5a-g were evaluated for anti-oxidant, anti-lipid peroxidation, and secretory phospholipase A(2) (sPLA(2)) inhibition in vitro, and sPLA(2) induced mouse paw edema. All the derivatives retained their anti-oxidant property compared to ascorbic acid at 6 × 10(-4)M and are good inhibitors of lipid peroxidation at 1 mg ml(-1) as evaluated by 2, 2-Diphenyl-1-picrylhydrazyl radical and thio-barbituric acid methods, respectively. Compounds 5e and 5f significantly inhibited purified group I sPLA(2) from Naja naja and group II sPLA(2) from Vipera russelli, human synovial fluid and human pleural fluid with IC(50) value ranging from 64 ± 1.95 to 82 ± 1.3 and 48 ± 2.27 to 61 ± 2.23 µM, respectively. The compounds 5e and 5f also showed varying degree of potency in neutralizing indirect hemolytic activity of sPLA(2) at 50 µM concentration, and sPLA(2) induced mouse paw edema at the dose 3 mg/kg. Further docking studies also confirmed that compounds 5e and 5f have maximum interaction with increasing negative energy value. Single molecule possessing both anti-oxidant and anti-inflammatory activities is of great therapeutic significance in inflammatory disorders.

Thrombin like activity of Asclepias curassavica L. latex: action of cysteine proteases.

AIM OF THE STUDY: To validate the scientific basis of plant latex to stop bleeding on fresh cuts. Cysteine protease(s) from Asclepias curassavica (Asclepiadaceae) plant latex was assessed for pro-coagulant and thrombin like activities. MATERIALS AND METHODS: A waxy material from the latex of Asclepias curassavica latex was removed by freezing and thawing. The resulted latex enzyme fraction was assayed for proteolytic activity using denatured casein as substrate. Its coagulant activity and thrombin like activity were determined using citrated plasma and pure fibrinogen, respectively. Inhibition studies were performed using specific protease inhibitors to know the type of protease. RESULTS: The latex enzyme fraction exhibited strong proteolytic activity when compared to trypsin and exerted pro-coagulant action by reducing plasma clotting time from 195 to 58 s whereas trypsin reduced clotting time marginally from 195 to 155 s. The pro-coagulant activity of this enzyme fraction was exerted by selectively hydrolyzing A alpha and B beta subunits of fibrinogen to form fibrin clot when pure fibrinogen was used as substrate as assessed by fibrinogen-agarose plate method and fibrinogen polymerization assay. Trypsin failed to induce any fibrin clot under similar conditions. The electrophoretic pattern of latex enzyme fraction-induced fibrin clot was very much similar to that of thrombin-induced fibrin clot and mimic thrombin like action. The proteolytic activity including thrombin like activity of Asclepias curassavica latex enzyme fraction was completely inhibited by iodoaceticacid (IAA). CONCLUSION: Cysteine proteases from Asclepias curassavica latex exhibited strong pro-coagulant action and were found to be specific in its action (Thrombin like). This could be the basis for the use of plant latex in pharmacological applications that justify their use as folk medicine.

Cysteine proteases from the Asclepiadaceae plants latex exhibited thrombin and plasmin like activities.

In the present study we evaluated the presence of cysteine protease from the latex of four plants Asclepias curassavica L., Calotropis gigantea R.Br., Pergularia extensa R.Br. and Cynanchum puciflorum R.Br. belongs to the family Asclepiadaceae. Cysteine proteases from these plants latex exhibited both thrombin and plasmin like activities. Latex enzyme fraction in a concentration dependent manner induced the formation of clot in citrated blood plasma. Direct incubation of fibrinogen with latex enzyme fraction resulted in the formation of fibrin clot similar to thrombin enzyme. However prolonged incubation resulted in degradation of the formed fibrin clot suggesting plasmin like activity. Latex enzyme fraction preferentially hydrolyzed Aalpha and Bbeta chains of fibrinogen to form fibrin clot. Latex enzyme fraction also hydrolyzed the subunits of fully cross linked fibrin efficiently, the order of hydrolysis was alpha-polymer > alpha-chains > beta-chain and gamma-gamma dimer. Cysteine proteases from all the four Asclepiadaceae plants latex exhibited similar action on fibrinogen and fibrin. This study scientifically validate the use of plant latex in stop bleeding and wound healing by traditional healers all over the world.

Synthesis and evaluation of tricyclic dipyrido diazepinone derivatives as inhibitors of secretory phospholipase A2 with anti-inflammatory activity.

A series of tricyclic dipyrido diazepinone derivatives 6(a-f) bearing different substituents at the tenth position of diazepinone ring were designed and are characterized by 1H NMR, FTIR and X-Ray crystallography studies. The synthesised derivatives are tested in-vitro phospholipase A2 (PLA2) enzyme inhibitory activity and in-vivo anti-inflammatory activity against purified group I and group II PLA2 enzymes from the snake venom and human pleural fluid. Compounds bearing aromatic ring with different substituents at different positions shown varied specificity. The 6f derivative with strong electron withdrawing nitro (-NO2) and trifluoromethyl (-CF3) groups at ortho and para positions respectively shown greater inhibitory activity. Inhibitory effect of the compound appeared to be direct interaction with active site and likely competes with substrates as supported by substrate dependent and calcium independent assays. The IC50 value of potent PLA2 inhibitor 6f was 22.1 microM and showed similar potency in the neutralization of in vivo PLA2 induced mouse paw edema and hemolytic activity.