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Remifentanil plasma concentrations purpose of the first part of the HPLC-UV method: Remifentanil is a new type of ultra-short-acting opioid receptor agonist, and its unique structure, susceptible to plasma and tissue non-specific ester enzyme metabolism, rapid onset, rapidly reach the blood - brain balance, the effect disappeared fast, anesthesia so that the controllability significantly improved, increasing people's attention. The experiment through the establishment of a high-performance liquid chromatography - UV detection method for the determination of human plasma concentration of remifentanil, designed to provide experimental data for the clinical use of drugs, and thus provide a theoretical basis for the clinical use of drugs. Method: In this paper, the RP-HPLC external standard method for the determination of the concentration of plasma samples remifentanil. 1 - chlorobutane extraction, first with anti-raised to 0.01 mol · L-1 hydrochloric acid and then drugs, to spherisorb-CN (4.6 mm × 250 mm, 5 μm, Waters) chromatography analytical column as the stationary phase; mobile phase was 0.02 mol · L-1 NaH2PO4 aqueous solution - acetonitrile (70:30) containing triethylamine 0.01%, a flow rate of 1.5 mL · min-1, column temperature to room temperature, UV detection at a wavelength of 210 nm, the injection volume of 200 μl. On day precision, extraction recovery, linearity range and detection limit determined to further evaluate the other Law. Six cases of elective thoracic surgery patients, ASAI, to Class II, aged 45 to 65 years of age, body mass index does not exceed 30%. Anesthesia was induced with intravenous remifentanil 4μg · kg-1 composite propofol 2.5 mg · kg-1, be patient lost consciousness after the intravenous injection of rocuronium, tracheal intubation, continuous inhalation of sevoflurane intermittent injection vecuronium 0.03 mg · kg-1 to maintain muscle relaxants. Before the injection of remifentanil and 1,2,3,5 min after injection, blood was collected from the cubital vein 3 mL, set heparin tubes, quickly adding 10 mg · mL-1 citrate 60 μL fully shaken and centrifuged The supernatant was immediately placed in the low-temperature refrigerator remifentanil plasma concentration was measured by high-performance liquid chromatograph. Results: In the analysis under the conditions established in this experiment, column efficiency, plasma drug baseline separation and high selectivity in plasma concentration within the linear range of 5 ~ 300 ng · mL-1 of remifentanil The plasma concentration of C (ng · mL-1) for its chromatographic peak area (A) linear regression was remifentanil standard curve equation: Y = -1585 530.54X r = 0.9996 (n = 7). The plasma remifentanil detection limit of 2.5 ng · mL-1. Plasma (200 ng · mL-1), medium (50 ng · mL-1), low (10ng · mL-1) concentrations of intra-day precision were 3.06%, 5.13% and 7.06%, respectively; inter-day precision 5.33%, 5.69% and 8.91%, respectively. Extraction recoveries were (88.99 ± 2.20)%, (83.72 ± 2.31)% and (83.47 ± 1.76)%. Ice for stability and frozen stability RSD were 5.52% and 5.87%, respectively. The ratio of the concentration of the labeled amount of freeze-thaw before and after the experiment were less than 15%. Conclusion: The established HPLC analysis method for the determination of in vivo concentration of remifentanil, the method is simple, good resolution, and its specificity, accuracy, precision, linearity and stability are suitable for the detection of the actual samples . The second part of the joint purpose of remifentanil propofol pharmacokinetics: when combined anesthesia, the complex interaction between propofol anesthetics and opioids, to determine the interaction between the two, research 's pharmacokinetic interactions have important clinical implications. The experiment by reversed-phase high-performance liquid chromatography (RP-HPLC) to detect blood concentration of propofol studies before and after remifentanil combined with propofol in vivo pharmacokinetics, clinical reasonable application of the drug to provide experimental and theoretical basis. Method: Chromatographic conditions: reversed-phase high performance liquid chromatography - fluorescence detection technology Waters HPLC system, separation column Waters Symmetry C18 (3.9 mm × 150 mm, 5.0μm), guard column for a Symmetry C18 to a mixture of methanol and water (75:25) was added glacial acetic acid pH = 4.0 as the mobile phase and fluorescence excitation wavelength of 270 nm, emission wavelength of 295 nm. Flow rate: 1.0 mL · min-1, column temperature to room temperature, 20μL of the supernatant was analyzed by HPLC. Plasma sample processing Methods: Plasma 250μL, the precision added thymol internal standard solution (10μg · mL-1) 10μL and methanol 750μL, vortex for 30 s, mixing, centrifuged 10 min (10000 r · min-1), draw The supernatant was filtered (0.45μm millipore), whichever 20μL analyzed by HPLC, averaged repeated twice. The establishment of the standard curve equation: blank plasma were added propofol standard stock solution to a concentration of 16,8,4,2,1,0.5,0.25,0.125,0.0625 μg · mL-1 series plasma standard solution , according to the above-mentioned sample processing operation, the sample peak area (As) and the internal standard peak area (Ai) of the sample concentration (C), the ratio (Y) for linear regression. Methodological study: To study the precision of the method, the extraction recovery and stability test. Method of administration, and blood samples were collected: elective thoracic surgery patients 21 cases, ASAI ~ II class, age 40 to 65 years old, does not exceed 30% of the body mass index, scheduled for general anesthesia, were divided into three groups (respectively I ~ III group), 7 cases in each group (n = 7). Group I: 30 min before anesthesia given midazolam 0.05 mg · kg-1 and atropine 0.5 mg, continuous monitoring of radial artery blood pressure, ECG, pulse oximetry (SpO2), end-tidal carbon dioxide partial pressure (PETCO2), brain electric bispectral index (BIS) and the components of the heart rate variability. Anesthesia was induced with intravenous propofol 2.5 mg · kg-1, be patient lost consciousness after the intravenous injection of rocuronium, endotracheal intubation, intubation success, connection anesthesia ventilator to maintain ventilation to maintain end-expiratory PETCO230 ~ 40 mmHg. Continuous inhalation of sevoflurane, the intermittent injection vecuronium 0.03 mg · kg-1 to maintain muscle relaxants; Group II: Anesthesia was induced with intravenous propofol 2.5 mg · kg-1 and remifentanil 4μg · kg-1 Group I; Group III: Anesthesia was induced by intravenous injection of propofol 2.5 mg · kg-1 and target-controlled infusion of remifentanil 6 ng · ml-1, the same as in the first group I. Forearm vein injection of propofol before injection and 2,4,6,10,15,30,45,60,90,120,180 min after injection blood was collected from the right cubital vein 2 mL, heparin After centrifugation, the supernatant plasma at cryogenic refrigerator to save. Data processing: the plasma sample results by DAS software processing, computing-compartment model and the pharmacokinetic parameters of SPSS software package for statistical analysis. Results: The plasma propofol determination methodology certification: the chromatographic behavior propofol and specificity: the experimental propofol plasma RP-HPLC detection method, propofol and thymol have larger The absorption peak, the separation efficiency and specificity. Propofol retention time of about 6 min, the retention time of the internal standard thymol approximately 4 min, and plasma impurities or endogenous substances do not interfere with the determination of propofol. Regression equation and the limit of detection: the standard curve equation: Y = 1.5826X -0.1264 (r = 0.9996, n = 9). Description plasma propofol concentration 0.0625 ~ 16μg · mL-1 The linear range of good relations, the detection limit of the experimental conditions measured propofol 0.03125μg · mL-1. The plasma sample recovery and precision of visits: The results show that, at high, medium, low (respectively and for 8,1,0.125 μg · mL-1) at three concentrations extraction recoveries were 84.32 ± 1.17,83.21 ± 1.24, 79.85 ± 1.50, intra-and inter-day RSD were less than 5.90%. Single with propofol and remifentanil combined with propofol pharmacokinetic studies: The results showed that the three groups of patients a single intravenous injection of propofol 2.5 mg · kg-1 plasma concentrations decreased rapidly. Sex, age, systolic blood pressure, diastolic blood pressure, heart rate, the difference was not statistically significant (P gt; 0.05). Propofol pharmacokinetic model in line with the two-compartment model. Pharmacokinetic parameters: distribution half-life (T1/2a), elimination half-life (T1/2β) concentration - time curve area (AUC), clearance (CL) and the volume of the central compartment (Vc), among groups significant difference (P gt; 0.05). Conclusion: RP-HPLC method for the determination of plasma propofol concentration, the method is simple, accurate, high sensitivity, good reproducibility, apply to the clinical propofol plasma concentration monitoring and pharmacokinetic studies. Intravenous and target-controlled infusion of remifentanil not affect propofol pharmacokinetic characteristics.
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