Effect of 'Procumbenase' a serine protease from Tridax procumbens aqueous extract on wound healing: A scar free healing of full thickness wounds.

Wound healing involves several cellular and molecular pathways. Tridax procumbens activates genetic pathways with antibacterial, antioxidant, anticancer, and anti-inflammatory properties, aiding wound healing. This study purified Procumbenase, a serine protease from T. procumbens extract, using gel filtration (Sephadex G-75) and ion exchange (CM-Sephadex C-50) chromatography. Characterization involved analyses of protease activity, RP-HPLC, SDS-PAGE, gelatin zymogram, PAS staining, mass spectrometry, and circular dichroism. Optimal pH and temperature were determined. Protease type was identified using inhibitors. Wound-healing potential was evaluated through tensile strength, wound models, hydroxyproline estimation, and NIH 3T3 cell scratch analysis. In incision wound rat models, Procumbenase increased tensile strength on day 14 more than saline and Povidone­iodine. It increased wound contraction by 89 % after 10 days in excision wound models, attaining full contraction by day 15 and closure by day 21. Scarless wound healing was enhanced by 18 days of epithelialization against 22 and 21 days for saline and povidone­iodine. Procumbenase increased hydroxyproline concentration 2.53-fold (59.93 ± 2.89 mg/g) compared to saline (23.67 ± 1.86 mg/g). In NIH 3 T3 cell scratch assay, Procumbenase increased migration by 60.93 % (50 µg) and 60.57 % (150 µg) after 48 h. Thus, Procumbenase is the primary bioactive molecule in T. procumbens, demonstrates scar-free wound healing properties.

Thrombin- and plasmin-like and platelet-aggregation-inducing activities of Plumeria alba L. latex: Action of cysteine protease.

ETHNOPHARMACOLOGICAL RELEVANCE: In folk medicine, parts of Plumeria alba L. are used for the treatment of many diseases, with its latex being used for curing skin diseases and promoting wound healing. AIM OF THE STUDY: This study aimed to study the role of P. alba L. latex in hemostasis and platelet aggregation. MATERIALS AND METHODS: The latex of P. alba L. was processed to remove waxes and enrich protein content, and the final extract was named Plumeria alba L. natant latex (PaNL). PaNL was analyzed for protease activity against casein. The type of protease in PaNL was identified by using protease inhibitors such as E-64, phenylmethylsulfonyl fluoride, ethylenediaminetetraacetic acid, and pepstatin A. Human fibrinogen, fibrin, and collagen types I and IV were subjected to hydrolysis with different concentrations of PaNL. The thrombin-like activity of PaNL was determined by analyzing its fibrinogen-clotting and procoagulant activities. The role of PaNL in platelet aggregation was also investigated. Its hemorrhagic and edema-inducing activities were evaluated in a mouse model. Phytochemical compounds were identified by gas chromatography-mass spectroscopy. RESULTS: The findings of casein/gelatin zymography confirmed that PaNL possesses protease activity. The results of the protease inhibition study indicated the presence of a cysteine-type protease(s) in PaNL. PaNL hydrolyzed the subunits of fibrinogen, fibrin, and collagen types I and IV. Its fibrin-degradation activity indicated that PaNL possesses plasmin-like activity. PaNL induced clotting of citrated human plasma within 3 min of incubation in the absence of CaCl2, indicating the presence of thrombin-like activity, which was further confirmed by the results of the fibrinogen-clotting assay. PaNL induced platelet aggregation in the absence of agonists. There was no hemolytic activity. Mice injected with PaNL did not show edema/ hemorrhagic activity. CONCLUSION: PaNL possesses procoagulant, fibrino(geno)lytic, thrombin- and plasmin-like activities and induces platelet aggregation, which could explain its usage for wound treatment in folk medicine.