Dipotassium hydrogen phosphate

Cetyl trimethyl ammonium bromide (CTAB), calcium nitrate (Ca(NO₃)₂·4H₂O), ethyl acetate, tetraethyl orthosilicate, anhydrous toluene, 3-aminopropyl triethoxysilane (APTES), polyacrylic acid (PAA), calcium chloride, disodium hydrogen phosphate, dipotassium hydrogen phosphate, glacial acetic acid, potassium hydroxide and fluoride (NaF) were all purchased from Sinopharm Chemical Reagent Co., Ltd. Artificial saliva was provided by Beijing Leigen Biotechnology Co., Ltd. All of the chemicals used in the experiments were of analytical grade. CPP-ACP dental protectors were purchased from GC Corporation Tokyo, Japan.

Citric acid (≥99.5%), thiourea (≥99%), potassium dihydrogen phosphate (≥99%), dipotassium hydrogen phosphate (≥99%), sodium hydroxide (≥98%), orthophosphate (≥85%), ethanol (≥99%), and other pure analytical reagents were procured from Sinopharm Chemical Reagent Co., LTD (Shanghai, China). HPLC-grade rutin standard (≥98%) was obtained from Shanghai Titan Technology Co., LTD (Shanghai, China).. Quercetin, luteolin, genistein, baicalin, chlorogenic acid, and kaempferol standard (≥98%) was obtained from Shanghai Aladdin Technology Co.LTD (Shanghai, China).. Ultrapure water (with a resistivity of 18.2 MΩ·cm) was used in all experiments.

Anthrone-sulfuric acid, glucose, sulfuric acid, sodium tetraborate, carbazole, ferric tartrate, sodium dihydrogen phosphate, dipotassium hydrogen phosphate, potassium hydroxide, diamine tetraacetic acid, ferric chloride, hydrogen peroxide, ascorbic acid, hydrochloric acid, 95% ethanol, and coomasylan bright blue (CBB) G-250 were purchased from Sinopharm Group Chemical Reagents Co. (Shanghai, China). The superoxide dismutase (SOD) test kit was purchased from Jiancheng Biological Engineering Institute (Nanjing, China). The bovine serum protein was purchased from Wuhan Tianyuan Biotechnology Co. (Wuhan, China). D-ribose was purchased from Sigma-Aldrich (Beijing, China).

Aspergillus niger HS-5 with high enzyme yield was screened by selective medium and compared its β-glucanase activity. The preparation and isolation of A. niger HS-5 β-glucanase was performed in house by using microbial fermentation. In brief, after fermentation, the cells in the fermented broth were removed by centrifugation to obtained crude enzyme solution, then ammonium sulfate salting out method was used to concentrate and Sephadex G-100 chromatography column was used for further purification.
Poria cocos were purchased from the Jiuzihe brand (Huanggang, China). Sulfuric acid, phenol, citric acid, sodium citrate, glucose, sodium dihydrogen phosphate, potassium ferricyanide, ferric chloride, trichloroacetic acid, 1,1-diphenyl-2-picryl-hydrazyl (DPPH), anhydrous ethanol, hydrochloric acid, trometamol, pyrogallic acid, salicylic acid, 1,10-phenanthroline, potassium dihydrogen phosphate, dipotassium hydrogenphosphate, sodium chloride, ferrous sulfate, and hydrogen peroxide (30%) were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China). All reagents were of analytical grade.

Huoxuerongluo Pill II was provided by the First Affiliated Hospital of Hunan University of Chinese Medicine. GR formula granules were purchased from a local drugstore (Changsha, China). The samples of GR decoction piece were from different provinces, including Hubei (No. S1–S3), Hunan (No. S4–S6), Sichuan (No. S7–S8), Jilin (No. S9–S11), and Yunnan (No. S12–S14) (the detailed information is listed in Table S4), which met the technical requirements for quality control and standard formulation of traditional Chinese medicine formula granules. These decoction pieces were further verified as GR by Professor Desheng Zhou from the First Affiliated Hospital of Hunan University of Traditional Chinese Medicine. Gastrodin, p-hydroxybenzyl alcohol, parishin A, parishin B, parishin C, and parishin E standards were all purchased from Chengdu Kloma Biotechnology Co., Ltd. (Chengdu, China) and their chemical structures are shown in Figure S4. Methanol and acetonitrile were both purchased from Beijing Inoke Technology Co., Ltd. (Beijing, China) Phosphoric acid, potassium dihydrogen phosphate, dipotassium hydrogen phosphate, and sodium hydroxide were all purchased from Sinopharm. The experimental water was made by SZ-93 automatic double pure water distiller.

All standard chemicals used for analyses were purchased from Dr. Ehrenstorfer GmbH (Augsburg, Germany), except for the internal standards (erythromycin-13C-d3, lincomycin-d3, sulfamethoxazole-d4, ciprofloxacin-d8, doxycycline-d3, and chloramphenicol-d5), which were purchased from Sigma-Aldrich (St. Louis, MO, USA). High performance liquid chromatography (HPLC)-grade solvents, including methanol, formic acid, and acetonitrile, were provided by Merck & Co., Inc. (Kenilworth, NJ, USA). Guaranteed reagents, including ammonium acetate, hydrochloric acid, sodium hydroxide, dipotassium hydrogen phosphate, and disodium ethylenediaminetetraacetic acid (EDTA), were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China). Ultrapure water was prepared using a water purification system. The internal standards were dissolved in acetonitrile as stock solutions and stored in a freezer at −40 °C. Working standard solutions were prepared immediately before measurements were taken by diluting the stock solutions with a mixture of menthol and water (9:1, v/v). For solid-phase extraction (SPE), Oasis HLB cartridges (6 mL/500 mg) were purchased from Water Oasis Co., Ltd. (Milford, MA, USA), glass microfiber filters (Whatman GF/F) were obtained from Sigma-Aldrich (St. Louis, MO, USA), and syringe-driven filters (PTFE) from Millipore Corp. (Bedford, MA, USA).