Elastase and inflammation inhibitory from Labisia pumila

Abstract

Labisia pumila (L. pumila) is widely reported to exhibit antioxidant, antimicrobialand anti-cancer activities. The goal of this study was to identify andquantify triterpenoid compounds from L. pumila leaves which are responsible foranti-inflammatory and anti-elastase activities by using bioassay-guided fractionation.The leaves of L. pumila were sequentially extracted with n-hexane, ethyl acetate,methanol, and water using serial exhaustive extraction procedure. Qualitativephytochemical analysis revealed the presence of phenolics, flavonoids, terpenoids,cardiac glycosides and saponins in the extracts. Each extract was assessed for totaltriterpenoids and polyphenols content by using vanillin-perchloric acid assay andFolin-Ciocalteu’s reagent, respectively. The ethyl acetate extract has the highest totaltriterpenoids content (78.94 mg oleanolic acid equivalent (OAE)/g dried extract) andappreciable amount of total polyphenols (38.93 mg gallic acid equivalent/g driedextract). The extracts were then tested for their anti-inflammatory and anti-elastaseactivities. The ethyl acetate extract showed the highest anti-inflammatory activity viathe inhibition of cyclooxygenase-2 (half-maximal inhibitory concentration (IC50) of34.32 μg/ml) and elastase (IC50 of 17.14 μg/ml). The correlation analysis of antiinflammatoryand anti-elastase ability of L. pumila with its total triterpenoids andtotal polyphenols content showed positive relationship. Further fractionation of theethyl acetate extract by solid phase extraction afforded sample fraction EAF-B withhigh total triterpenoids content (91.50 mg OAE/g dried fraction) and anti-elastaseactivity (IC50 of 13.27 μg/ml). Rapid liquid chromatography-electrospray ionizationtandemmass spectrometry method was developed and validated for the identificationand quantification of pentacyclic triterpenoids with elastase inhibition property fromthe sample fraction EAF-B. Three pentacyclic triterpenoids, namely betulinic acid,oleanolic acid and lupeol were identified. The content of betulinic acid, oleanolicacid and lupeol in the fraction were 2.67, 5.04 and 1.47 mg/g dried fraction,respectively. The limits of detection and quantification ranged from 0.04 to 0.12μg/ml and 0.11 to 0.65 μg/ml, respectively. The response surface methodologyanalysis showed that solvent-solid ratio (p<0.01), extraction time (p<0.05) andtemperature (p<0.01) were statistically significant factors affecting the totaltriterpenoids extraction. The optimum extraction conditions obtained were: solventsolidratio of 37.76, extraction time of 4.24 hours and extraction temperature of 55.27°C. The results suggested that L. pumila has promising anti-inflammatory and antielastaseactivity that could serve as potential source of natural elastase inhibitor intreating inflammatory skin disorders and photoaging.