Rifampicin d3

High-quality reference standards were sourced to ensure the precision and accuracy of our analytical methodology. PZA as a certified reference material, INH (≥99% purity), and RIF (>97% purity) were procured from Sigma-Aldrich (St. Louis, MO, USA). In addition, the following reference standards and internal standards were obtained from Toronto Research Chemicals (North York, ON, Canada): ETH (98% purity), 25-D-RIF (95.06% purity), RIF-Q (95.19% purity), and 3-F-RIF (96.42% purity); ethambutol-D4 (ETH-IS; 98% purity), rifampicin-D3 (RIF-IS; 98.15% purity), pyrazinamide-15N,D3 (PZA-IS; 99.5% purity). Acetonitrile, water, and methanol were procured from Sigma-Aldrich (St. Louis, MO, USA), while isopropanol was sourced from Fisher Chemical (Fair Lawn, NJ, USA). All solvents met LCMS grade specifications. Formic acid (FA) (>99.0% purity) was supplied by Fisher Chemical. Ammonium formate (≥99% purity) was obtained from Honeywell (Morristown, NJ, USA). Blank urine of individual donors for method validation was received from BioIVT (Westbury, NY, USA).

Pyrazinamide (lot: 100178–201104), isoniazid (lot: 100578–201502), rifampicin (lot: 130496–201403), and ethambutol dihydrochloride (lot: 100165–201705) were purchased from National Institutes for Food and Drug Control. Internal standards (ISs), pyrazinamide-D3 (lot: CRC-OW3-056), isoniazid-D4 (lot: 2-JLO-114-1), rifampicin-D3 (lot: 5-YEN-103-103-1), rifampicin-D8 (lot: NJ-07222019), and ethambutol-D4 (lot: 1-WEW-51-4), were purchased from Toronto Research Chemicals, Canada. The purities of these analytes were all above 98.0%. Molecule structures of the four analytes and corresponding ISs are presented in Figure 1.
Methanol (HPLC grade) and acetonitrile (HPLC grade) were purchased from Merck Drugs & Biotechnology, German. Trifluoroacetic acid (HPLC grade) was purchased from Sigma Aldrich Fluka, USA. Ammonium formate (HPLC grade) was purchased from Anaqua Chemical Supply, USA. Water was prepared by a Milli-Q deionization system (Millipore, Billerica, MA, USA). Blank human serum was obtained from the physical examination population of The First Affiliated Hospital, Zhejiang University School of Medicine.