title: A validated HPLC-ESI-MS/MS method for quantification of 2-hydroxy-4-methoxy benzoic acid from rat plasma and its application to pharmacokinetic study using sparse sampling methodology.
abstract: The phenolic compound, 2-hydroxy-4-methoxy benzoic acid (HMBA), is one of the major phytoconstituents of Decalepis arayalpathra (Joseph & Chandra.) Venter, a rare and endemic medicinal plant found in the Western Ghats of India. HMBA has been attributed to possess several biological effects including anti-inflammatory, anti-pyretic, anti-oxidant and anti-diabetic. The present article describes a rapid and sensitive liquid chromatography-tandem mass spectrometric method (HPLC-MS/MS) for the determination of HMBA in rat plasma. In brief, the developed assay involves pre-treatment of the plasma samples by an optimized solid phase extraction method (recoveries for HMBA greater than 90%) followed by chromatographic separation on a Cosmosil C18 (150mm×4.6mm i.d.; 5μm particle size) analytical column with mobile phase of methanol and 10mM ammonium formate (95:5 v/v; 0.2% formic acid) delivered at a constant flow rate of 1.0mL/min. The detection and quantification was performed using an Applied Biosystems Hybrid Q-Trap API 2000 mass spectrometer equipped with electrospray ionization source (ESI) functioning in negative mode. The developed assay was validated as per the US FDA bioanalytical guidelines with the calibration curve linear over the concentration range of 5.05-2019.60ng/mL (r(2)≥0.9936) for HMBA from rat plasma. Further, the validated HPLC-MS/MS method was successfully applied to pharmacokinetic study of HMBA after oral administration of D. arayalpathra tuber extracts to female albino Wistar rats using sparse sampling methodology. Following oral administration, the maximum mean concentration in rat plasma (Cmax -1301.57±128.22ng/mL) was achieved at 1.5h (Tmax) and the area under the curve (AUC0-48h) was 8985.02±229.54ngh/mL. The elimination half-life (t1/2) and terminal elimination rate constant (Kel) were 2.48h and 0.28 L/h, respectively.
authors.affiliation: Ramnarain Ruia College, Matunga (East), Mumbai 400019, Maharashtra, India. Electronic address: sreenathnandakumar@gmail.com.
authors.forename: Sreenath Nandakumar
authors.initials: SN
authors.lastname: Nair
authors.affiliation: Ramnarain Ruia College, Matunga (East), Mumbai 400019, Maharashtra, India.
authors.forename: Mandar
authors.initials: M
authors.lastname: Mhatre
authors.affiliation: Institute for Advanced Training and Research in Interdisciplinary Sciences (TDM Laboratory), Sion (East), Mumbai 400 022, Maharashtra, India.
authors.forename: Sasikumar
authors.initials: S
authors.lastname: Menon
authors.affiliation: Herbal Research Laboratory, Ramnarain Ruia College, Matunga (East), Mumbai 400019, Maharashtra, India.
authors.forename: Sunita
authors.initials: S
authors.lastname: Shailajan
mesh_terms: D000284:Administration, Oral; D000818:Animals; D029062:Apocynaceae; D019540:Area Under Curve; D001565:Benzoates / Q000008:administration & dosage / Q000097:blood* / Q000493:pharmacokinetics*; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods* / Q000592:standards; D004355:Drug Stability; D005260:Female; D006207:Half-Life; D016014:Linear Models; D008657:Metabolic Clearance Rate; D008517:Phytotherapy; D010936:Plant Extracts / Q000008:administration & dosage / Q000097:blood* / Q000493:pharmacokinetics*; D035281:Plant Tubers; D010946:Plants, Medicinal; D017208:Rats, Wistar; D012015:Reference Standards; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods* / Q000592:standards; D053719:Tandem Mass Spectrometry / Q000379:methods* / Q000592:standards
chemical_list: C112536:2-hydroxy-4-methoxy benzoic acid; D001565:Benzoates; D010936:Plant Extracts
keywords: 2-Hydroxy-4-methoxy benzoic acid; Decalepis arayalpathra (Joseph & Chandra.) Venter; HPLC–MS/MS; Pharmacokinetics; Sparse sampling methodology