title: Precise simultaneous quantification of methadone and cocaine in rat serum and brain tissue samples following their successive i.p. administration.
abstract: A sensitive high-performance liquid chromatography (HPLC) assay with dual UV detection has been developed and validated for the simultaneous quantification of methadone and cocaine in rat serum and brain tissue samples. Liquid-liquid extraction using hexanes was applied for samples extraction with Levo-Tetrahydropalmatine (L-THP) as the internal standard. Chromatographic separation of the analytes was achieved on a reversed-phase Waters Symmetry®C18column (150mm×4.6mm, 5μm). A gradient elution was employed with a mobile phase consisting of 5mM potassium phosphate containing 0.1% triethylamine (pH=6.5) (A) and acetonitrile (B) with a flow rate of 1mL/min. UV detection was employed at 215nm and 235nm for the determination of methadone and cocaine, respectively. The calibration curves were linear over the range of 0.05-10μg/mL for both methadone and cocaine. The assay was validated according to FDA guidelines for bioanalytical method validation and results were satisfactory and met FDA criteria. Inter-day accuracy values of serum and brain samples ranged from 96.97 to 105.59% while intra-day accuracy values ranged from 91.49 to 111.92%. Stability assays showed that both methadone and cocaine were stable during sample storage, preparation, and analytical procedures. The method was successfully used to analyze biological samples obtained from a drug- drug interaction pharmacokinetics (PK) study conducted in rats to investigate the effect of methadone on cocaine PK. Our method not only can be used for bioanalysis of samples obtained from rats but also can potentially be applied to human biological serum samples to monitor compliance to methadone maintenance therapy (MMT) and to detect possible cocaine-methadone co-abuse.
authors.affiliation: Department of Pharmaceutical Sciences, University of Maryland, School of Pharmacy, Baltimore, MD, United States; Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Misr International University, Cairo, Egypt.
authors.forename: David S
authors.initials: DS
authors.lastname: Nakhla
authors.affiliation: Department of Pharmaceutical Analytical Chemistry, Faculty of Pharmacy, Ain Shams University, Cairo, Egypt.
authors.forename: Lobna A
authors.initials: LA
authors.lastname: Hussein
authors.affiliation: Department of Pharmaceutical Analytical Chemistry, Faculty of Pharmacy, Ain Shams University, Cairo, Egypt.
authors.forename: N
authors.initials: N
authors.lastname: Magdy
authors.affiliation: Department of Pharmaceutical Sciences, University of Maryland, School of Pharmacy, Baltimore, MD, United States; Department of Analytical Chemistry, Faculty of Pharmacy, University of Sadat City, Sadat City, Egypt.
authors.forename: Inas A
authors.initials: IA
authors.lastname: Abdallah
authors.affiliation: Department of Pharmaceutical Sciences, University of Maryland, School of Pharmacy, Baltimore, MD, United States; Department of Pharmaceutics and Industrial Pharmacy, Faculty of Pharmacy, Helwan University, Cairo, Egypt. Electronic address: hhassan@rx.umaryland.edu.
authors.forename: Hazem E
authors.initials: HE
authors.lastname: Hassan
mesh_terms: D000701:Analgesics, Opioid / Q000008:administration & dosage / Q000032:analysis / Q000097:blood / Q000493:pharmacokinetics*; D000818:Animals; D001921:Brain / Q000378:metabolism; D000697:Central Nervous System Stimulants / Q000008:administration & dosage / Q000032:analysis / Q000097:blood / Q000493:pharmacokinetics*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003042:Cocaine / Q000008:administration & dosage / Q000032:analysis / Q000097:blood / Q000493:pharmacokinetics*; D006801:Humans; D059625:Liquid-Liquid Extraction / Q000379:methods; D008297:Male; D008691:Methadone / Q000008:administration & dosage / Q000032:analysis / Q000097:blood / Q000493:pharmacokinetics*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D015813:Substance Abuse Detection / Q000379:methods
chemical_list: D000701:Analgesics, Opioid; D000697:Central Nervous System Stimulants; D003042:Cocaine; D008691:Methadone
keywords: Brain; Cocaine; HPLC; Methadone; Rats; Serum