| Objective: Opioids have been widely used in gynecologic surgery, anesthesiaand analgesia in labor. More detailed pharmacokinetic data, completemonitoring measures and application of opioid antagonist drugs reversed theability of its adverse reactions makes the harmful side effects of drug use hasbeen reduced. The pharmacokinetic characteristics of the study drug transferthrough the placenta can help determine fetal exposure from the parent drug.However, due to difficult for the placenta in vivo studies and taking intoaccount the problem of maternal and fetal safety, limiting the direct study ofthe human placenta. The purpose of this study is investigates thepharmacokinetics of the placental transfer of homegrown sufentanil using thedual-perfused, sing-cotyledon human placental model.Method: Select ten maternal of healthy full-term elective cesareansection. The placentas were persused via an umbilical arterty with heparinizedice-cold Krebs-Ringer bicarbonate （NaCl,120mmol/L；NaHCO3,30mmol/L；KCL,4.7mmol/L；CaCl22H2O,3.3mmol/L；KH2PO4,1.2mmol/L；MgSO47H2O,2.4mmol/L；Glucose,1g/L；Heparin,25IU/ml；pH=7.4） within tenminutes, make sure the blood of endovascular was washed out and saved thefunction of the placenta. An umbilical arterial branch and correspondingvenous vessel that provided the principal blood supply to a non-peripheralcotyledon with an intact maternal surface were identified and catheterized.Trocars in children were used as the indwelling catheters. If the arterysupplied more than one cotyledon, both arterial and venous were ligated. Putthe cotyledon in the special placenta box after make sure the cotyledon noleakage. Then put the placenta box into constant temperature water bath.Maternal flow rates were maintained at10-12ml/min, producing perfusatepressure of35-45mmHg. Fetal flow rate （1-3ml/min） were adjusted toprovide perfusate pressure of60-75mmHg. Perfused by peristaltic pump. Both maternal side and fetal side have a reservoir, The reservoir into the waterbath tank. During the whole experiment, constant temperature water bathmaintained at37℃. The perfusates were aerated with a mixture of95%oxygen and approximately5%carbon dioxide. The pH of both perfusates washeld constant at7.4±0.05by manipulating the amount of carbon dioxide in thegas mixture. Establish the dual-perfused, sing-cotyledon human placentalmodel. The closed （recirculating） method or open（single-pass ornon-recirculating） method of placental perfusion was used to study domesticsufentanil placental transfer mechanism, placental tissue accumulation ofsufentanil and characteristics of placental transfer of sufentanil close to thephysiological state.The concentration of sufentanil and antipyrine was detected byHPLC/MS/MS. Sample （clossed method） extraction procedure:1ml samplewas transferred to a15ml glass tube, followed by addition of internalstandard（IS） solution15μl （fentanyl1ng/μl）, acetonitrile2ml. Samples werevortex-mixed for5min. Then dichloromethane2ml and chloroform2ml wasadded, all samples were vortex-mixed for5min again. Centrifuged for10min（2000r/min）. The lower organic solvent was placed in a10ml glass centrifugetube, evaporated under a nitrogen stream with temperature of30℃. Residuewas dissolved with1ml ultrapure water and vortex-mixed for1min. Thesamples were transferred to auto-sampler vials and10ul was injected into theHPLC system. Sample （open method） extraction procedure:1ml sample wastransferred to a15ml glass tube, followed by addition of IS solution15μl（fentanyl1ng/μl）, dichloromethane2ml, stand for1min. The lower organicsolvent is placed in a10ml glass centrifuge tube after stand for1min.Evaporated under a nitrogen stream with temperature of30℃. Residue wasdissolved with1ml ultrapure water and vortex-mixed for1min. The sampleswere transferred to auto-sampler vials and10ul was injected into the HPLCsystem. Chromatographic conditions: Column: Phenomenex SB-C18（2.1mm×30mm,1.8μm）； Mobile phase: A-methanol, B-0.1%formic acid, thegradient elution; Flow rate:0.2ml/min; Column temperature:25℃; Injection volume:10ul; MS condition: Ion source: electrospray ionization source （EIS）;Nebulizer:25.0psi; Ion source temperature:100℃；Ionization mode: positive;Scanning: Multiple reaction monitoring（MRM）. Glucose consumption andlactate formation was detected by biochemical analysis.Result: Sufentanil crossed the placenta rapidly in both （M→F and F→M）directions. Sufentanil transfer rate （Sufentanil transfer rate/Antipyrine tran-sfer rate） M→F （0.48±0.02） and F→M （0.56±0.07） were notsignificantly different （p＞0.05）. Sufentanil be bound by the placental tissue.Sufentanil transfer through the placenta to the fetal transfer rate increasedwith maternal sufentanil concentration.Conclusion: Establishment the dual-perfused, single-cotyledon humanplacental model contribute to the study drug pharmacokineticcharacteristics of the placental transfer. Sufentanil crossed the placenta rapidlyin both （M→F and F→M） directions. Absolute transfer ratio of sufentanilincreased with the perfusion time. Sufentanil be bound by the placental tissue. |
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