Milli q water purifying system

Working standards of Olmesartan acid (OLM) and OLM D₆ (OLM D₆) were procured from VIVAN Life Sciences (Thane-Mumbai, India) and Simson Pharma (Dahisar-Mumbai, India), respectively. Working standard of HCTZ was also procured from VIVAN Life Sciences and HCTZ ¹³C D₂ (HCTZ ¹³C D₂) was purchased from Splendid Lab (Pune, India). OLM D₆ and HCTZ ¹³C D₂ were used as internal standards (IS) for OLM and HCTZ, respectively. Methanol was purchased from Merck. Drug-free human plasma with ethylene diamine tetra acetic acid (K₂EDTA) was purchased from Laxmi Sai Clinical Labs (Hyderabad, India). Formic acid (SQ grade) and ammonium acetate (Excela R grade) were purchased from Qualigens Fine Chemicals (Mumbai, India). Diethyl ether and dichloromethane were obtained from S.D. Fine Chemicals Ltd. (Mumbai, India). Milli-Q water with resistivity of 8.2 milliohm at 25°C and total organic carbon (TOC) ≤500 ppb was used from the in-house Milli-Q water purifying system (Millipore, SAS, Molsheim, France).

LC-MS grade acetonitrile, methanol and trifluoroacetic acid were purchased from Carlo Erba (Milan, Italy). α-Cyano-4- hydroxycinnamic (CHCA) was purchased from Sigma-Aldrich Co. (St. Louis, Missouri, USA). Deionized water was produced using a Milli-Q water purifying system (Millipore Corp., Bedford, MA, USA).

Acetonitrile (HPLC grade) was purchased from J. T. Baker (USA). Laboratory-deionized water was produced by a Milli-Q water purifying system (Millipore, USA), iodoacetamide, somatic ACE (rabbit lung) and BK acetate were purchased from Sigma-Aldrich (USA). The synthetic peptide APQERGPPEIPP was purchased from FastBio Ltda (Brazil).

δ-ALA hydrochloride, acetylacetone reagent, and Formaldehyde (37%, V/V in water) were provided by Aladdin-Reagent (Shanghai, China). Porphobilinogen was purchased from Sigma-Aldrich (St. Louis, MO, United States). Methanol HPLC grade, glacial acetic acid, and hydrochloric acid were supplied by J&K (China). The aqueous solutions were prepared by purified water. The ultrapure water was obtained from a Millipore Milli-Q water purifying system (Millipore, Bedford, MA, United States), with a specific resistance of 18.2 MΩ cm.
Stock solutions for δ-ALA and PBG were prepared as 3 and 1 mM, respectively, in amber-colored volumetric flasks and were preserved in the refrigerator until use. Samples for both the analytes were prepared following a procedure reported previously (Ajmal et al., 2019 (link)). Calibration plots were obtained by spiking different solutions with concentrations as 0–5 µM for δ-ALA and 0–15 µM for PBG. Acetylacetone reagent was formulated by mixing acetylacetone, ethanol, and water in a ratio of 15:10:75, respectively. Formaldehyde solution was prepared by 3.7–fold dilution of commercially available formaldehyde solution and was stored in darkness (Oishi et al., 1996 (link)).

The chemicals were of analytical reagent grade or a higher purity (HPLC-grade). The solvents (methanol and acetonitrile) were supplied by Tedia® (Rio de Janeiro, Brazil). Ammonium acetate (CAS 631- 61-8, CH₃COONH₄), zinc sulfate heptahydrate (ZnSO₄·7H₂O), and acetic acid (CAS 64-19-7, CH₃COOH) were from Merck Millipore®. Sodium heparin (5,000 IU·mL⁻¹) was purchased from Hipolabor (Belo Horizonte, Brazil). Ultrapure water was provided daily by a Milli-Q® purifying water system (Merck-Millipore).