Optimization of extraction protocol of Parmelia perlata and its validation for protective effects against oxalate-induced renal injury in NRK-52E cells

Abstract

The objectives of the present study are to optimize the extraction protocol for preparing a hydro-ethanolic extract of Parmelia perlata, to investigate the extract for its protective effects against calcium oxalate (CaOx) crystallization and oxalate-induced injury in NRK-52E cells. The full factorial design of experiment was used to optimize the extraction process. Three extraction variables (i.e. extraction time, temperature and drug-solvent ratio) were studied at two levels and one central point was included. A total of nine experimental runs were set by randomization. A polynomial mathematical equation was obtained by regression analysis using JMP software for predicting the experimental model. The response (yield of extract) was found to be significantly affected by two factors i.e. time and temperature. The optimized P. perlata extract (PPE) so prepared was screened by HPTLC, evaluated for its inhibitory effects on CaOx crystallization by turbidimetric assay and on oxalate-induced injury in NRK-52E cells. PPE significantly (p < 0.05) inhibited the CaOx crystallization in a concentration-dependent manner. Further, the oxalates injured the NRK-52E cells, promoted the cellular damage, increased LDH (lactate dehydrogenase) release and decreased the cell viability to approximately 46.01%. PPE significantly (p < 0.05) prevented this cellular injury, reduced the LDH release and improved the cell viability to 92.45%. PPE showed antiurolithiatic effects, probably by inhibiting the crystallization of CaOx and preventing the tubular retention of crystals by protecting the renal epithelial cells against oxalate-induced injury. Such experimental substantiation of PPE for antiurolithiatic potential has not been previously carried out and this is the first scientific report.