Standardization of the manufacturing procedure for Pinelliae Rhizoma Praeparatum cum Zingibere et Alumine

Pinelliae Rhizoma (PR), the dried tuber of Pinellia ternata (Thunb.) Breit., is a traditional Chinese medicinal herb. It is commonly used for treating cancer, cough and phlegm. To treat cancer, Chinese medicine practitioners often use raw PR; while to treat cough and phlegm, they usually use Pinelliae Rhizoma Praeparatum cum Zingibere et Alumine (PRZA, raw PR processed with ginger juice and alumen as adjuvant materials). Currently, the producing protocol of PRZA varies greatly among different places in China. This study aims to standardize the manufacturing procedure for PRZA. We also evaluated the impact of processing on the bioactivities and chemical profile of raw PR. We used the orthogonal design to optimize the manufacturing procedure of PRZA at bench scale, and validated the optimized procedure in pilot-scale production. The MTT assay was used to compare the cytotoxicities of raw PR and PRZA in hepatocellular carcinoma HepG2 cells. Animal models (ammonia liquor-induced cough model and phenol red secretion model) were used to compare the antitussive and expectorant effects of raw PR and PRZA, respectively. The chemical profiles of raw PR and PRZA samples were compared using a newly developed ultra-performance liquid chromatography/quadrupole-time-of-flight mass spectrometry (UPLC/Q-TOF-MS) method. The standardized manufacturing procedure for PRZA is as follows: soak raw PR in water until the center of the cut surface is devoid of a dry core, after that, boil the herb in water (for each 100kg raw PR, 12.5kg alumen and 25L freshly squeezed ginger juice are added) for 6h, and then take out and dry them. The cytotoxicity of PRZA was less potent than that of raw PR. Intragastric administration of raw PR or PRZA demonstrated antitussive and expectorant effects in mice. These effects of PRZA were more potent than that of raw PR at the dose of 3g/kg. By comparing the chemical profiles, we found that six peaks were lower, while nine other peaks were higher in PRZA than in raw PR. Six compounds corresponding to six individual changed peaks were tentatively identified by matching with empirical molecular formulae and mass fragments. The manufacturing procedure for PRZA was standardized. This protocol can be used for PRZA industrial production. The bioactivity assay results of raw PR and PRZA (produced using the standardized protocol) support the common practice for the clinical applications of these two decoction pieces. Moreover, raw PR and PRZA showed differen