Determination Of Citalopram In Human Plasma By HPLC With Fluorescence Detector And Study On Its Pharmacokinetics And Relative Bioavailability

Objective: Depression is a most common illness that affects a large number of individuals in many countries. However, depression is under diagnosed and frequently under treated. Recent evidence suggests that depressive episodes, if left untreated, may heighten severity of subsequent episodes and may increase need for more health care resources. Citalopram was manufactured by Lundbeck Company of Denmark and was approved by FDA of America in 1998. Citalopram is an serotonin reuptake inhibitors(SSRIs) of novel antidepressants with a more specific and selective pharmacological profile than other antidepressants of its class. Same to others SSRIs, citalopram inhibit serotonin reuptake of synaptic space as its major pharmacological action. Citalopram is used at therapy of depression widespread in clinical. This study was designed to establish an HPLC-fluorescence method for determination of citalopram in human plasma. To assay concentration of citalopram in human plasma after 22 healthy volunteers administration of citalopram 40mg (test or reference) and to draw the concentration-time curves. The study calculates the pharmacokinetics of citalopram and discusses the behavior of pharmacolinetics in Chinese healthy subject. The study is base on the absorption of two citalopram tablets to calculate the relative bioavailability and to compare its bioequivalent. Methods: Twenty-two male healthy subjects were all students, aged 22.5±1.0 years and weighting 67.4±8.3kg. Medical Ethical Committee of Hebei General Hospital approved the study. Each subject signed the informed consent before participating in the study. Physical examinations and laboratory test was done before the study. Twenty-two volunteers were divided into 2 groups and participated in the study conducted according to an open randomized two-way crossover design. In the first part of study, subjects received either citaloprams(test or reference). A crossover study was followed by a washout period of 2 weeks. In the second part of study, two groups exchange the drugs. Venous blood samples (5ml) were drawn into heparinized tubes before and 0.5, 1, 2, 4, 6, 8, 16, 24, 36, 48, 72, 96, 144 h after the drug was administrated. The samples were centrifuged and frozen at -20℃before analysis. The waters HPLC system was used with 474 fluorescence detector. The analytical column was Zorbax SB C8 particle side 5μm, 150×4.6 mm ID. The mobile phase was a mixture of phosphoric acid-dihydrogenmine(20 mmol/L, pH 3.5) and acetonitrile as 65:35 at a flow-rate of 1ml/min. The parameters of the detector were as follows: Ex 240nm and Em 302nm. The column temperature was maintained at 30oC. Millennium 32TM chromatography workstation was used to print thechromatography of citalopram and calculate the concentration of citalopram in plasma. The initial data was processed by 3P97 software (Chinese Pharmacological Society). C_max and t_max were obtained directly from concentration-time curve. Other pharmacokinetic parameters were calculated by non-compartment model method. The study used ladder-shaped method to calculate AUC_0-144h , t_1/2=0.693/λ_z, AUC_144h-∞=C_t/λ_z, AUC_0-∞=AUC_0-144h+ AUC_144h-∞, CL/F=dosage of drug/ AUC_0-∞, V_d/F= CL/F/λ_z。Major parameters of pharmacokinetic were transformed into ln_(parameters), and analyzed with following statistical analysis: ANOVA, paired two one-side t-test, 90% confidence interval. To evaluating the bioequivalence of two citalopram tablets. Result: Citalopram and internal standard were well separated from the biological background under the described chromatographic conditions, and the retention time were 4.0 min and 6.9 min respectively. The peaks were of good shape and completely resolved. No interference from plasma matrix was observed. The limit of quantification (s/n) was 0.4μg/L. The calibration curve was linear over a concentration range of 1.0 –100.0μg/L and the limit of detection was 1μg/L. The method recovery was 97%–100%. So this HPLC method can be used at therapeutic drug monitoring (TDM) completely. Twenty-two volunteers after administrated 40mg citalopram (test or reference), the parameters of pharmacokinetic was showed on following: tmax was 4.14±2.01 h(mean±S.D) and 4.45±1.74 h,...