| A convenient method--target controlled infusion (TCI)--was developed and applied three decades ago. The basic principle of TCI is that the anesthesiologist sets and adjusts the target blood anesthetic concentration according to the depth of anesthesia required. As hemodynamic values vary widely because of the degree of surgical stress, the association of boluses with a variable opioid infusion rate allows a more stable hemodynamic profile with a satisfactory anesthetic state. Target-controlled infusion (TCI) has been developed to rapidly achieve and maintain the target plasma or effect-site concentrations of i.v. anesthetics deemed appropriate by the anesthetist.I.V. sufentanil, currently used for the induction and maintenance of anesthesia, has several favorable pharmacokinetic profiles for use with the TCI system. Sufentanil is a potent opioid with the long analgesic effect. The consequence of its long action is that sufentanil is not easy to infuse during a long period of time without risk of accumulation, delayed recovery and postoperative respiratory depression. As sufentanil has a long context-sensitive half-time (25 min after a 3-h infusion), TCI will prevent long-acting opioid-induced perioperative accumulation and will allow rapid recovery from anesthesia. Furthermore, TCI would target an extubation concentration associated with good analgesia without delaying tracheal extubation or causing postoperative respiratory depression. With this system, the tedious task of calculating the amount of administered drug required to achieve the target concentration is controlled by a microprocessor, which processes several pharmacokinetic models to predict sufentanil concentration.Although several models have been proposed and validated for their ability to predict sufentanil concentrations in human plasma, few of them were performed when volunteers were programmed to diverse effect-site sufentanil concentrations. Actually, target concentrations of anesthetic drugs are not always be fixed in clinical cases. Moreover, accurate measurement of the concentrations of this narcotic drug is the key to successful evaluation of the sufentanil TCI system. Thus, there is considerable demand for a specific, sensitive and rapid method for quantitative determination of sufentanil in human plasma. In this context, the aims of this work are (1) to build a UPLC-MS/MS method to determine the plasma levels of sufentanil in volunteer patients undergoing abdominal surgery and (2) to verify whether those levels can justify the accuracy of Gepts sets when the effect-site concentrations of sufentanil changed at multi-levels, from 0.06 to 0.8 ng/ml, during selective surgery.Materials and methods1. Volunteer selectionThe pharmacokinetic study was approved by the local ethics committee. Informed consent was obtained from 17 non-obese adult patients. Study exclusion criteria were:age< 20 yr or>60 yr, ASA physical status class IV or V, a history of drug or alcohol abuse, a history of allergy to opioids or any other drug used in the study, psychiatric disorders, pregnancy, breastfeeding, the presence of hepatic, kidney or pulmonary dysfunction, and participation in another research project.2. Anesthetic procedureAll patients underwent abdominal surgery for extended gynecologic or gastrointestinal tumors or multiple lymph-node resections. Patients received sufentanil TCI from the beginning of anesthesia using the pharmacokinetic sets reported by Gepts et al.. They also received other non-opioid anesthetics necessary for surgical procedures. For stable hemodynamic conditions throughout the entire surgery, target effect-site concentrations varied in multi-level (0.06-0.8 ng/ml) before some main steps of the surgery. The main steps of the surgery were:awake, loss of consciousness, no response to laryngoscope, tracheal intubation, scalpel incision, peritoneal traction, skin closure and spontaneous breathing.3. Sample collectionArterial blood samples were drawn 5min after the predicted blood concentrations reached the target effect-site concentrations. These samples were collected in separate sodium-heparin-containing tubes and centrifuged at 3℃with a rotation speed of 3000rpm for 15 min. The supernatant was transferred into a 1.0 ml Eppendorf tube and preserved at—80℃. After thawing, plasma samples were similarly centrifuged as mentioned above before further sample preparation.4. Sample extraction(1) Sample pre-treatment:a plasma aliquot of 880μl of plasma was combined with 20μl of fentanyl (2.5 ng/ml) as an internal standard and 100μl of 1% ammonium hydroxide before deproteinization.(2) Pre-condition of solid-phase extraction (SPE):each SPE cartridge was washed with 1ml of pure methanol and 1ml of purified water.(3) Sample loading:Loaded 1 ml prepared plasma sample to a single cartridge.(4) Analyte elution:Washed each SPE cartridge with 1ml of 1% methanol (v/v) and 1ml pure methanol.(5) Sample dryness:The sample was evaporated to dryness under vacuum condition for 24 h.(6) Ahead of sample injection:The residue was dissolved with 1ml of the mobile-phase solution. After centrifugation,10μl of centrifuged supernatant was injected into the UPLC system.5. Measurement of sufentanil blood concentrations5.1 Chromatography and mass spectrometryAnalyses were performed on a Waters ACQUITYTM UPLC system (Waters Corp., Milford, MA, USA). Double-tandem-mass-spectrometric detection was carried out on a Waters Micromass(?) Quattro Premier XE ES mass spectrometer (Waters Corp., Milford, MA, USA) with an electrospray-ionization (ESI) interface. All data collected in centroid mode were acquired and processed using MassLynxTM NT 4.1 software with the QuanLynxTM program (Waters Corp.).5.2 Preparation of standard solutionsSolutions of sufentanil were prepared at concentrations of 0.071,0.142,0.235, 0.570,1.140,2.280 and 4.560 ng/ml, and these new solutions were used as a serial concentration calibration standard for sufentanil. Internal standard (IS) samples were prepared with distilled water at a concentration of 2.5 ng/ml, and all solutions were stored at—4℃after preparation.5.3 Method validationThe method was evaluated for specificity, linearity, precision and accuracy, extraction recovery, matrix effects and stability according to the FDA guidelines for the validation of bioanalytical methods.6. Predictive accuracy analysis of sufentanil TCI system6.1 Intrasubject data analysis(1) The prediction error (PE) is an indicator of the bias of the concentration achieved.(2) The median prediction error (MDPE) and median absolute prediction error (MDAPE) are reflections of the bias of the TCI model.(3) The wobble is a measurement of the intrasubject variability of PE.(4) The divergence is an indicator of the TCI inaccuracy during infusion. 6.2 Intersubject data analysis(1) Measured values divided by the predicted values plotted over time:this graph provided a means to visually assess the extent of bias and precision in the study group.(2) Plots of the regression lines for PEij-MDPEi on time:this graph illustrated how the errors in each patient tended to differ over time from that patient's own MDPE,.(3) Plots of (PEij-MDPE,) over time (where j= 1,...Ni in the ith subject):this graph illustrated the magnitude of the terms used to estimate the wobblei of each patient. It is the median of these absolute values that provides the wobblei of each patient.6.3 Intra-model comparisonUnlike many study of the sufentanil TCI models, which were set to reach only one steady concentration of sufentanil or two, this study was performed when volunteers were programmed to diverse effect-site sufentanil concentrations. Except for the work of Slepchenko et al., they were successful in building and analyzing a nonlinear mixed model for obese people with target concentrations ranging from 0.3 to 0.65 ng/ml. Thus we listed the results of predictive accuracy analysis of nonlinear model from Slepchenko's study and Gepts model from the present study.Results 1. Sample extractionIn our study, we found that SPE gave good results for the separation of low amounts of sufentanil from human plasma.2. Method validation(1) SpecificityNo interference from endogenous substances or other metabolites was observed in the retention time of sufentanil.(2) LinearityThe calibration curve of sufentanil (peak area ratios of analytes to IS versus the concentration of analytes in human plasma) was linear over the concentration range of 0.07-4.56 ng/ml. A typical equation for the calibration curve is y=-0.02965+ 0.5515 x,r2> 0.999.(3) Precision and accuracyThe study examined the intra-day accuracy and the inter-day precision for sufentanil from QC samples. The RSDs at three QC concentration levels (0.071,0.57 and 4.56ng/ml) were less than 15%.(4) Extraction recovery and matrix effectThe consistency of the recovery of sufentanil and IS supports the application of this procedure to routine analysis. Also, the matrix effects of sufentanil and the IS were between 85% and 115%, indicating that no co-eluting substance influenced the ionization of the analytes and IS.(5) StabilityThe results from all stability tests demonstrated the good stability of sufentanil over all determination steps.3. Predictive accuracy analysis3.1 Intrasubject data analysis To our knowledge, a mean 20-30% variation can be considered clinically acceptable. Therefore, our study suggested that the Gepts pharmacokinetic sets for the sufentanil TCI model were accurate for predicting plasma concentrations during surgery, with a variation of target effect-site concentrations from 0.06 to 0.8 ng/ml.3.2 Intersubject data analysis(1) Measured values divided by the predicted values plotted over time:many curves were lining above the base line, suggesting that measured concentrations of sufentanil were greater than predictive concentrations in many cases.(2) Plots of the regression lines for PEij-MDPEi on time:this graph illustrated how the errors in each patient tended to differ over time from that patient's own MDPE;. A negative slope in this study means the difference between PEij and MDPEi tends to shrink over time.(3) Plots of (PEij-MDPEi) over time (where j= 1,...Ni in the ith subject):this graph illustrated the magnitude of the terms used to estimate the wobblei of each patient. It is the median of these absolute values that provides the wobblei of each patient. In this study, most curves were stretching around the base line, suggesting that the Gepts parameters are available for prediction and target-control of the plasma concentration of sufentanil.3.3 Intra-model comparisonBoth models were appropriate for the use of sufentanil TCI system, with a variation of target concentrations, besides the wobble percentage of the Slepchenko's study was smaller than that in our study.Conclusion1. In this work, we present a highly sensitive UPLC-MS/MS method for the analysis of sufentanil. To the best of our knowledge, this work is the first report of a UPLC-MS/MS-based approach for measurement of sufentanil in human fluids. 2. The Gepts pharmacokinetic sets for the sufentanil TCI model were accurate for predicting plasma concentrations during surgery, with a variation of target effect-site concentrations from 0.06 to 0.8 ng/ml. |
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