Hemostatic, milk clotting and blood stain removal potential of cysteine proteases from Calotropis gigantea (L.) R. Br. Latex

Abstract

Introduction:Plant latex is a natural source of biologically active compounds and several hydrolytic enzymes responsible for their diverse health benefits. Recent past has witnessed substantial progress in understanding their supplementary industrial and pharmaceutical utility. Calotropis gigantea is one of the important latex producing plants belonging to asclepediaceae family with wide ethnopharmacological applications and is rich in proteolytic enzymes. Present study investigates hemostatic, milk clotting and blood stain removal potential of C. gigantea latex proteases.Materials and Methods:The protease activity of crude enzyme (CE), obtained by centrifugation followed by ammonium sulphate precipitation and dialysis, was assayed using casein as the substrate. Effect of pH, temperature and specific inhibitors on protease activity was determined. Native PAGE and in gel protease activity of CE was performed. Hemostatic (Fibrinogen polymerization, fibrinogen agarose plate and blood clot lysis assays), milk clotting and blood stain removal efficacies of CE were determined.Results:CE exhibited high caseinolytic activity. Enzyme activity was optimum at 37-50ºC and pH 8.0. Fibrinogen polymerization assay showed concentration dependent increase in turbidity indicating thrombin like activity which was further confirmed by fibrinogen agarose plate assays. Clot lysis assay indicated 92.41% thrombolysis by CE in 90 min. CE also revealed significantly high ratio of milk clotting to protease activity (Milk Clotting Index, MCI = 827.59 ± 1.52). Complete destaining of blood stained fabric was observed when incubated with 1% detergent incorporated with 0.1mg/ml CE. The study highlights and validates the compound application potential of latex cysteine proteases from C. gigantea.

Detection and analysis of triterpenic compounds in apple extracts

“…[24] The extraction of triterpenic compounds from botanical raw material is most commonly carried out with the help of non-polar extractants, such as acetone, alcohols (ethanol, methanol or (less frequently) other lower alcohols), hexane, ethyl acetate, diethyl ether, or liquid chlorinated hydrocarbons (chloroform or dichloromethane) or their mixtures. [24][25][26] The analysis of scientific literature revealed a number of publications on the phytochemical composition of apples and its variation in whole apples and apple peel and flesh samples. Most commonly, phenolic compounds were analyzed, whereas other equally important biologically active compounds of other groups (such as pentacyclic triterpenic compounds) received much less attention.…”

Production, properties and applications of proteases from pongamia oil seed cakes

“…Later, 100 μL of partially purified enzyme was introduced to the substrate aseptically to test the enzyme capacity to coagulate the milk protein. 32…”

The Proteolytic Activity of Philibertia gilliesii Latex. Purification of Philibertain g II

“…Plant extracts with a high content of proteolytic enzymes can be applied in a variety of processes such as the manufacture of food, and biomedical and environmental applications [3]. Clarified latex extracts of Calotropis gigantea and Calotropis procera have been recently used for biomedical applications [33][34][35], and those of Carica papaya to biodegrade protein effluents [36] and to hydrolyze raw hide [37]. Philibertain g obtained from latex of P. gilliesii fruits containing several endopeptidases [21] was extensively characterized, taking into account its potential use in industrial processes.…”