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CAS:136656-07-0
Appearance:White to off-white solid
Storage:Powder : 2-8℃, 2 years; In solvent(mother liquid): -20℃, 1 month; -80℃, 6 months
Purity:HPLC≥98%
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Examples of using this product（for reference only）
In Vitro：
In Vitro Incubation of Timosaponin BII with Gut Microbiota:
After 7 SD rats were sacrificed, the colon contents were collected and added to the sterilized anaerobic medium (Solarbio Life Sciences Co., Ltd., Beijing, China) at a ratio of 1.0 g:20 mL, and stirred gently. After filtering, the culture medium containing gut microbiota (mixed medium) was placed in a N2 atmosphere, and pre-incubated at 37 ℃ for 60 min before use. Accurately weigh 1.0 mg of timosaponin BII and dissolve with methanol to obtain a solution of 1 mg/mL. The incubation system consists of 10 μL of timosaponin BII in methanol (1 mg/mL), and 990 μL of mixed medium. The incubation was conducted in a 37 ℃, 200 rpm shaking incubator. The incubation system must be maintained completed in an anaerobic environment during the experiment. In the first incubation experiment, the timosaponin BII and mixed medium were incubated for 0, 1, 2, 6, 12, and 24 h, respectively. In the second incubation experiment, the drugs and mixed medium were incubated for 0, 0.25, 0.5, 1, 1.5, and 2 h, respectively. In addition, the negative control group was introduced consisting of twice boiled mixed medium incubated with the same amount of timosaponin BII. The termination reaction was carried out by adding 3-fold volume of 100 ng/mL glipizide methanol solution (IS), shaking it evenly and then precipitating the protein. After each sample was centrifuged at 13,400× g rpm in a 4 ℃ refrigerated centrifuge for 10 min, 10 μL of supernatant was injected for LC-MS/MS analysis, and 7 μL of supernatant was injected for LC/MS-Q-TOF analysis.
In VitroIncubation of Timosaponin BII with Liver Microsomes and Liver Homogenate:
The liver microsome incubation system was consisted of the following: 5 μL SD rat liver microsomes (20 mg/mL), 2 μL timosaponin BII (1 mM), 20 μL of NADPH, and 0.05 mM Tris/HCl (pH = 7.4), with a total volume of 200 μL. The incubation was conducted in a shaking incubator at 37 ℃ and 800 rpm with supply of oxygen. After the incubation, 3-fold volume of glipizide methanol solution (IS, 100 ng/mL) was added to the incubation system and mixed to stop the reaction at 0, 0.25, 0.5, 1, 1, 5, and 2 h. After centrifugation at 13,400× g rpm in a refrigerated centrifuge at 4 ℃ for 10 min, 10 μL of the supernatant was injected for LC-MS/MS analysis.
Liver homogenate was prepared by homogenizing freshly collected SD rat livers and adding ice-cold normal saline with weight/volume = 1:3. The liver homogenate incubation system was consisted as follows: 2 μL timosaponin BII (1 mM), 198 μL of freshly prepared liver homogenate, with a total volume of 200 μL.
Incubation was conducted in a shaking incubator at 37 ℃ and 800 rpm with supply of oxygen. After the incubation, 3-fold volume of glipizide methanol solution (IS, 100 ng/mL) was added to the incubation system and mixed to stop the reaction at 0, 0.25, 0.5, 1, 1, 5, and 2 h. After centrifugation at 13,400× g rpm in a refrigerated centrifuge at 4 ℃ for 10 min, 10 μL of the supernatant was injected for LC-MS/MS analysis.
Reference：
Dong GM, Yu H, Pan LB, Ma SR, Xu H, Zhang ZW, Han P, Fu J, Yang XY, Keranmu A, Niu HT, Jiang JD, Wang Y. Biotransformation of Timosaponin BII into Seven Characteristic Metabolites by the Gut Microbiota. Molecules. 2021 Jun 24;26(13):3861. doi: 10.3390/molecules26133861. PMID: 34202717; PMCID: PMC8270264.
