Berberine hydrochloride bc

Berberine hydrochloride (BC) (>98% purity, Lot No. 20130306) was manufactured by Shanghai Yuanye Biotechnology Co., Ltd. (Shanghai, China). Clotrimazole (CLO) (99% purity, Lot No. 23593-75-1) was provided by BaDaTong Medical Company (Taizhou, Zhejiang, China). Time and concentration effects of berberine and clotrimazole on T. mentagrophytes were determined as proposed previously [42 (link)]. Briefly, the fungi were cultured at 30,000 cells/mL, in presence of berberine hydrochloride (0.5, 1, or 2 mg/mL) or clotrimazole (5.0 μg/mL) at 37 °C for 0 h, 10 h, 22 h, 34 h, 46 h, 58 h and 60 h, respectively. Duplicate 1 mL aliquots of homogenized samples were removed from each conical flask and transferred to pre-weighed Eppendorf tubes. After centrifugation was performed 13,300× g for 20 min, the sediments were dried in an oven at 60 °C. Differential weights were determined using an analytical balance. Each experiment was performed in duplicate. The growth of T. mentagrophytes was observed.

The M. canis strain (CMCC(F)M3h) used in this study was provided by the Institute of Internal Medicine, Chinese Academy of Medical Sciences (Nanjing, China) National Center. The M. canis was cultured in Tryptic Soy Broth (TSB) at 28°C for 96 h. The densities of the suspension were adjusted to primary inoculum of 1.0 × 10⁷ CFU/mL in the normal saline solution with total volume of 100 mL in conical flasks. berberine hydrochloride (BC) (HPLC>98%) was purchased from Yuan Ye Biological technology co., Ltd, Shanghai, China. The BC treated group (three samples) was subject to medicinal stress dose which means 1/2 MIC (minimum inhibitory concentration) dose (1 mg/mL berberine in DMSO), while the control group was fed with DMSO (Purity>99%) which was purchased from Sangon Biotech co., Ltd Shanghai, China only. Each group has three repeats. After 6 h incubation (28°C,150 rpm), the seeds were filtered with gauze and frozen after treatment for RNA isolation and further analysis.