Development of a High Performance Liquid Chromatography with Coulometric Electrochemical 123 1 M.Sc., Sciencetist, 5 Ph.D., Associate Professor, 6 Ph.D., Lecturer, Faculty of Pharmaceutical Sciences, Khon Kaen University, Khon Kaen 2 M.Sc., Pharmacist, the Food and Drug Administration, the Ministry of Public Health, Bangkok 3 Ph.D., Associate Professor, Faculty of Pharmacy, Mahidol University, Bangkok 4 Ph.D., Associate Professor, Faculty of Pharmaceutical Science, Chulalongkorn University, Bangkok 1 «∑.¡. π—°«‘∑¬“»“ µ√å 5 Ph.D. √Õß»“ µ√“®“√¬å§≥–‡¿ —™»“ µ√å 6 Ph.D. Õ“®“√¬å §≥–‡¿ —™»“ µ√å ¡À“«‘∑¬“≈—¬¢Õπ·°àπ ¢Õπ·°àπ 2 ¿.¡. ‡¿ —™°√  ”π—°ß“π§≥–°√√¡°“√Õ“À“√·≈–¬“ °√–∑√«ß “∏“√≥ ÿ¢ °√ÿ߇∑æ¡À“π§√ 3 Ph.D. √Õß»“ µ√“®“√¬å §≥–‡¿ —™»“ µ√å ¡À“«‘∑¬“≈—¬¡À‘¥≈ °√ÿ߇∑æ¡À“π§√ 4 Ph.D. √Õß»“ µ√“®“√¬å §≥–‡¿ —™»“ µ√å ®ÿÓ≈ß°√≥å¡À“«‘∑¬“≈—¬ °√ÿ߇∑æ¡À“π§√ * Corresponding author: Detpon Preechagoon, Faculty of Pharmaceutical Sciences, Khon Kaen University, Khon Kaen 40002 Tel./Fax. +66 4336 2092 E-mail: detpre@kku.ac,th Original Article Development of a High Performance Liquid Chromatography with Coulometric Electrochemical Detector Method for the Analysis of Morphine in Human Plasma °“√æ—≤π“«‘∏’«‘‡§√“–Àå¡Õ√åøïπ„πæ≈“ ¡“¡πÿ…¬å‚¥¬°“√„™â‡∑§π‘°‚§√¡“‚∑°√“øï ¢Õ߇À≈« ¡√√∂π– Ÿß√à«¡°—∫‡§√◊ËÕßµ√«®«—¥·∫∫‡§¡’‰øøÑ“™π‘¥§Ÿ≈Õ¡‡¡µ√‘° Thanee Tessiri 1 , Viroj Sumyai 2 , Suvatana Chulavatnatol 3 , Poj Kulvanich4, Khanittha Tontisirin 2 , Thaned Pongjanyakul 5 , Sirikul Aumpon 2 , Verawan Uchaipichat 6 , Detpon Preechagoon 5 * ∏“π’ ‡∑»»‘√‘ 1 , «‘‚√®πå  ÿà¡„À≠à 2 ,  ÿ«—≤π“ ®ÿÓ«—≤π±≈ 3 , æ®πå °ÿ≈«“π‘™ 4 , ¢π‘…∞“ µ—𵑻‘√‘π∑√å 2 , ∏‡π» æß»å®√√¬“°ÿ≈ 5 , »‘√‘°ÿ≈ Õ”æ≈ 2 , «’√«√√≥ Õÿ™“¬Õ¿‘™“µ‘ 6 , ‡¥™æ≈ ª√’™“°ÿ≈ 5 * Received: 2 June 2009 Accepted: 25 July 2009 Abstract A HPLC method using reversed-phase chromatography coupled with a coulometric electrochemical detector (ECD) was developed for the determination of morphine in human plasma. Hydromorphone was selected as an internal standard. The compounds were extracted using solid-phase extraction with C 18 cartridges and separated on a reversed-phase C 18 column with the mobile phase consisting of 69% v/v phosphate buffer pH 2.2 (5 mM sodium phosphate monobasic and 0.7 mM sodium dodecyl sulfate) and 31% v/v acetonitrile. The working electrode of the ECD was set at +450 mV for analytical purposes. Under this condition, the limit of detection for morphine was 0.33 ng/mL, whereas the limit of quantitation was 1.88 ng/mL. The percent recovery, intra-day and inter-day variation of morphine determinations were in the percentage range of 85.07-93.41, 2.86-6.41 and 3.76-11.39, respectively. The frozen samples kept at -20 o C proved to be stable for at least one month prior to extraction. Also, the repeated use of cartridges for extraction was found to generate reliable results. Keywords: Morphine, HPLC, Electrochemical detector IJPS 2009; 5(2): 123-130