Early- and intermediate-term results of surgical correction in 122 patients with total anomalous pulmonary venous connection and biventricular physiology.

BACKGROUND: We retrospectively reported our 26-year experience with operative repair of total anomalous pulmonary venous connection (TAPVC) with biventricular physiology. METHODS: Between December 1982 and December 2008, 122 TAPVC patients with biventricular heart underwent surgical repair in our department. Moderate or deep hypothermia was induced at the time of cardiopulmonary bypass (CPB). Follow-up was conducted for 5 postoperative years. Surgical outcomes of early and intermediate deaths after TAPVC repair were retrospectively analyzed. RESULTS: Six deaths occurred operatively; and three deaths, during follow-up. The 5-year survival rates after TAPVC repair was 92.6 %, without gradient across the anastomosis. The survival rate of the patients who were younger was 78.8 %, significantly lower than those older than 1 year. It was also lower in those who were less than 6 kg in weight. Three patients died during follow-up. Three patients died of ventricular arrhythmia, right heart failure, and pneumonia, respectively, during follow-up. If the left atrium pressure was higher than 15 mm Hg, the snare of the vertical vein was loosened after CPB ceased in the patients with supracardiac connection. It decreased from 21 ± 5 to 13 ± 3 mm Hg. The vertical vein was ligated in 57 cases and left open in 20 cases. A patient with an intact vertical vein had a large shunt and was cured by intervention afterward. Supraventricular arrhythmia occurred in 19 patients with the supercardiac type repaired through a biatrial incision. One patient died of ventricular arrhythmia, and none of the remaining patients had arrhythmias. CONCLUSION: Surgical treatment of TAPVC carried a low operative risk and had satisfactory immediate and intermediate results. Age younger than 1 year and weight less than 6 kg were risk factors. It was a good choice to leave the vertical vein open in the patients with a left atrial pressure higher than 15 mm Hg.

[The anatomy features and surgical significance of the pulmonary circuits of pulmonary atresia with ventricular septal defect and major aortopulmonary collateral arteries].

OBJECTIVES: To analyze the anatomy features of the pulmonary circuits in the patients with pulmonary atresia (PA) with ventricular septal defect (VSD) and major aortopulmonary collateral arteries (MAPCA), and discuss the clinical significance. METHODS: From April 2002 to June 2010, the anatomy features of pulmonary circuits in 33 patients with PA/VSD/MAPCA were examined and analyzed. There were 21 male and 12 female patients. The age ranged from 11 months to 29 years. The anatomic types of PA/VSD included group B for 22 cases, group C for 11 cases. Thirty-one patients of them underwent 33 operative procedures. The operations included aorta-pulmonary shunt in 8 cases, one stage unifocalization with VSD open in 2 cases, complete repair in 23 cases. RESULTS: Twenty-nine (87.9%) patients had native pulmonary arteries, 6 of them were normal size and 23 were hypoplastic size. Four patients (12.1%) had no native pulmonary arteries. The postoperative oxygen saturation of the patients undergone shunt and one stage unifocalization was increased to 83% to 90%. There was one early death after complete repair because of multiorgan function failure. There were 4 cases of severe low cardiac output and 3 cases of respiratory function failure. Sixteen patients after complete repair were followed up more than one year. The postoperative right ventricular pressure was 41 to 99 mmHg (1 mmHg = 0.133 kPa). The ejection fraction value was more than 50% in 14 patients and less than 50% in 2 patients. Two patients had medium pulmonary insufficiency. CONCLUSIONS: An individualized approach based on the anatomy of the pulmonary circuits permits achievement in the patients with PA/VSD/MAPCA. The surgical strategy for PA/VSD/MAPCA mainly depends on the anatomy features of native pulmonary arteries, confluent pulmonary arteries and MAPCA.

Combination of ELISA and dried blood spot technique for the quantification of large molecules using exenatide as a model.

INTRODUCTION: To explore the feasibility of coupling dried blood spot (DBS) technique with ELISA for the quantification of large molecules, exenatide was used as a model. A method for the quantification of exenatide in human blood was developed and evaluated. METHODS: Exenatide standard and quality control samples prepared in fresh human blood were spotted on DBS cards and then extracted. The extraction conditions were optimized by comparing different extraction solutions, with/without protease inhibitors, and various incubation times. A competitive ELISA assay was used for quantification of exenatide from DBS samples. RESULTS: The assay range of exenatide standards in blood was 100-5000 pg/mL. The intra-assay precision (%CV) was from 1.2% to 16.3%, and the accuracy (%Recovery) was from 87.5% to 117.0%. The inter assay precision (%CV) was from 1.7% to 14.3%, and the accuracy was from 95.0% to 115.5%. All the above assay parameters met acceptance criteria. Furthermore, the storage stability of exenatide on DBS cards was tested at ambient temperature as well as at 4°C and -70°C, and it was found that change of storage temperature did not affect the stability of exenatide significantly. DISCUSSION: Our results demonstrated a successful coupling of DBS technique with ELISA for quantification of exenatide in human blood, and the DBS-ELISA combination has a great potential to be further applied for the quantification of other large molecule drugs or biomarkers.

Effective pulmonary artery perfusion mode during cardiopulmonary bypass.

AIM: Reducing lung injury during cardiopulmonary bypass (CPB) is important for patients' recovery. The present study was designed to research convenient and effective pulmonary artery perfusion mode during CPB in an animal model. METHODS: Twelve healthy mongrel dogs were randomly divided into 2 groups: a control group and a perfusion group designed to simulate clinical CPB-induced lung injury. During CPB, pulmonary artery perfusion with modified low-potassium dextran (LPD) solution was performed immediately after the initiation of ischemia and before reperfusion for 3 to 4 minutes each time, with pressure maintained at 15 to 20 mmHg; animals in the control group were not perfused. After pulmonary reperfusion, the changes in pulmonary function and tissue biochemical data were determined. RESULTS: Compared with the control group, lung compliance, oxygenation, and vascular resistance after reperfusion were significantly improved in the perfusion group. The malonaldehyde concentration, neutrophil sequestration ratio, and tissue water content also decreased significantly in the perfusion group. CONCLUSION: The pulmonary artery perfusion mode used in this experiment could relieve CPB-induced lung injury effectively. Improving cellular tolerance to hypoxia and decreasing inflammatory reaction may be the important mechanisms. Moreover, this mode is convenient and does not interfere with the intended operations, which is promising for clinical use.

[Clinical analysis of surgical procedures and outcomes for corrected transposition of great arteries with heart anomaly].

OBJECTIVE: To determine the outcome of anatomically corrective repair and traditional repair of corrected transposition of great arteries (c-TGA) with heart anomaly. METHODS: From April 2002 to December 2006, nineteen patients including fourteen male and five female with c-TGA, underwent operations, age ranged from 2 to 22 years old and weight ranged from 10 to 48 kg. Fifteen of them received anatomically corrective repair and the other four received traditional repair. Eighteen patients were referred to SLL (segmental anatomy) in situs solitus while fifteen of them with levocardia and three with dextrocardia. One patient was referred to IDD (segmental anatomy) in situs inversus with levocardia. Associated cardiac lesions included ventricular defect in eighteen patients, double outlet of right ventricle in one patient, pulmonary stenosis in seventeen patients and pulmonary hypertension in two patients. The operative procedures to anatomically correct atrioventricular discordance included an atrial switch plus a ventricle-arterial switch. The atrial switch was performed using the modified Senning procedure (n=13), Senning procedure (n=1) and Mustard procedure (n=1). The ventricle-arterial switch was performed using a Rastelli procedure (n=13) or an arterial switch (n=2). The patients underwent Mustard and Rastelli procedure had received bi-direct Gleen shunt due to postoperative high pressure of superior vena cava. Three patients underwent traditional cardiac repair because of small ventricular septal defect and one patient was reoperated to undergo traditional cardiac repair because of left ventricular failure after received anatomically corrective repair. RESULTS: In the patients received anatomically corrective repair, there was one early operative death received a modified Senning atrial switch and an arterial switch. The cause of death was acute myocardial failure due to imperfect coronary transfer. The postoperative complications included severe low cardiac output syndrome (n=1), temporary atrioventricular block (n=1) and thorax cavity fluidify (n=1). The survivors were followed up for 6 months to 4 years. All were sinus cardiac rhythm and in NYHA class I or II. There was no death in the patients received traditional repair. Four patients were followed up for 1 year. Three patients were in NYHA I or II class and one patient in class II. CONCLUSIONS: Anatomically corrective repair of c-TGA can be performed with good operative survival and intermediate-term outcome. The patients with good right ventricular function and well developed tricuspid valve who were difficult to undergo anatomically corrective repair might be fit to receive traditional repair.

[Study of cardiopulmonary adaptation during exercise in patients after extracardiac conduit total cavopulmonary connection].

OBJECTIVE: To study cardiopulmonary physiology during exercise in patients after extracardiac total cavopulmonary connection (ECTCPC). METHODS: Twenty-six patients were studied after ECTCPC by exercise testing with bicycle treadmill protocol. Heart rate (HR), blood pressure (BP), respiratory frequency (RF) and pulse oxygen saturation (SpO(2)) were measured continuously; twenty-six patients suffered from Fallot 4 underwent biventricular repair were also studied as control group. RESULTS: In ECTCPC group, HR, BP, SpO(2) and RF all increased with exercise below 3 grade; when exceed 4 grade, BP, SpO(2) decreased and RF kept increasing. Compared with control group, HR, RF were higher (t = 2.13, P < 0.05; t = 2.31, P < 0.05), SpO(2) was lower (t = 2.46, P < 0.05) under the quiescent condition; When exceed 3 grade, HR, BP, SpO(2) decreased more significantly, but RF increased continuously. In fenestration group after ECTCPC, HR reached the top at 5 grades, but in group without fenestration it reached the top at 3 grades; In the whole process of exercising, RF kept higher and SpO(2) kept lower in fenestration group. CONCLUSIONS: The ECTCPC patients showed obviously exercise limitation. Totally bypass of sinoatrial node in this operation may have some adverse effects on the integer regulation of HR.

[Clinical analysis on surgical treatment of single atrium].

OBJECTIVE: To summarize the experience of surgical treatment of single atrium. METHODS: From August 1984 to August 2004, there were 33 patients with single atrium in our study. Plastic surgery for mitral valves were performed for 30 cases with mitral insufficiency. Plastic surgery for tricuspid valves were performed for 18 cases with tricuspid valve insufficiency. There were 3 cases only with complete absence of atrial septum. There were 14 cases with left superior vena cava. All new atrial septums were made with patches including 24 autologous pericardial patches and 9 terylene patches. Complicate abnormalities were corrected in the same time. Tow suture techniques were used in operations to prevent conductive system block, and plastic surgery for mitral valves were performed until the mitral valves were sufficiency. RESULTS: There weren't death and conductive system block after operation in the group. One case was low-grade mitral insufficiency and the others weren't mitral insufficiency. Twenty-five cases were followed up from 3 months to 11 years, and they could work and study normally. CONCLUSIONS: Single atrium should be operated as early as possible. The key of surgery is to prevent conductive system block, to properly correct mitral insufficiency and to drastically correct complicated abnormality. The new atrial septum should be made by patch and an autologous pericardial patch is the first selection.

Stabilization and determination of a PPAR agonist in human urine using automated 96-well liquid-liquid extraction and liquid chromatography/tandem mass spectrometry.

Two stability challenges were encountered during development of an urine assay for a proliferator-activated receptor (PPAR) agonist, I (2-{[5,7-dipropyl-3-(trifluoromethyl)-1,2-benzisoxazol-6-yl]oxy}-2-methyl propionic acid), indicated for the treatment of Type II diabetes. First, the analyte was lost in urine samples due to adsorption on container surface which is a common problem during clinical sample handling. Secondly, the acylglucuronide metabolite (III), a major metabolite of I, displayed limited stability and effected the quantitation of parent drug due to the release of I through hydrolysis. Therefore, a clinical collection procedure was carefully established to stabilize I and its acylglucuronide metabolite, III, in human urine. The metabolite was not quantitated with this method. The urine samples are treated with bovine serum albumin (BSA) equal to 1.75% of the urine volume and formic acid equal to 1% of urine volume. Compound (I) and internal standard (II) were extracted from urine with 1 mL ethyl acetate using a fully automated liquid-liquid extraction in 96-well plate format. The analytes are separated by reverse phase high-performance liquid chromatography (HPLC) with tandem mass spectrometry in multiple-reaction-monitoring (MRM) mode used for detection. The urine method has a lower limit of quantitation (LLOQ) of 0.05 ng/mL with a linearity range of 0.05-20 ng/mL using 0.05 mL of urine. The method was validated and used to assay urine clinical samples.

Absorption, metabolism, and excretion of [14C]MK-0767 (2-methoxy-5-(2,4-dioxo-5-thiazolidinyl)-N-[[4-(trifluoromethyl)phenyl] methyl]benzamide) in humans.

MK-0767 (KRP-297; 2-methoxy-5-(2,4-dioxo-5-thiazolidinyl)-N-[[4-(trifluoromethyl)phenyl] methyl]benzamide) is a thiazolidinedione (TZD)-containing dual agonist of the peroxisome proliferator-activated receptors alpha and gamma that has been studied as a potential treatment for patients with type 2 diabetes. The metabolism and excretion of [14C]MK-0767 were evaluated in six human volunteers after a 5-mg (200 microCi) oral dose. Excretion of 14C radioactivity was found to be nearly equal into the urine (approximately 50%) and feces (approximately 40%). Elimination of [14C]MK-0767 was primarily by metabolism, with minimal excretion of parent compound into the urine (<0.5% of dose) and feces (approximately 14% of the dose). [14C]MK-0767 was the major circulating compound-related entity (>96% of radioactivity) through 48 h postdose. It was also found that approximately 91% of the total radioactivity area under the curve was due to intact MK-0767. Several minor metabolites were detected in plasma (<1% of radioactivity, each), formed by cleavage of the TZD ring and subsequent S-methylation and oxidation. All the metabolites excreted into urine were formed by TZD cleavage, whereas the major metabolite in feces was the O-demethylated derivative of MK-0767.

A dual PPAR alpha/gamma agonist increases adiponectin and improves plasma lipid profiles in healthy subjects.

BACKGROUND: The objective of these studies was to evaluate the pharmacokinetics and pharmacodynamics of MK-0767, a prototypical dual peroxisome proliferator-activated receptor (PPAR) alpha/gamma agonist, following administration of single and multiple oral doses in healthy male subjects. METHODS: The first study was a double-blind, randomised, placebo-controlled, alternating two-panel, rising dose protocol in which single doses of 1-80 mg of MK-0767 were administered. The second study was a double-blind, randomised, placebo-controlled, staggered incremental dose, parallel-group protocol in which multiple doses of 0.3-25 mg of MK-0767 were administered once daily for 14 days. In both studies at each dose level, six subjects received MK-0767 and two subjects received placebo. RESULTS: Plasma area under the concentration-time curve and maximum plasma concentration increased with single and multiple doses of MK-0767 over the dose ranges studied. The apparent terminal half-life of MK-0767 averaged approximately 36 hours following single and multiple doses. Steady-state plasma concentrations were achieved following approximately 8 days of multiple doses. Compared with placebo, MK-0767 produced dose-dependent reductions in triglycerides (-26 +/- 8% [p = 0.002] and -33 +/- 13% [p = 0.008]) and free fatty acids (-50 +/- 11% [p < 0.001] and -67 +/- 23% [p = 0.008]) following single and multiple doses, respectively. Significant (p < or = 0.050) dose-dependent alterations in adiponectin (332 +/- 36%), low-density lipoprotein cholesterol (-29 +/- 5%), total cholesterol (-19 +/- 3%), non-high-density lipoprotein cholesterol (-28 +/- 4%), and fasting plasma glucose (-6 +/- 2%; only in the 25 mg group) were observed after multiple doses. CONCLUSIONS: The observed effects of MK-0767 on adiponectin, free fatty acids and lipids, even after single doses, demonstrate that this prototypical dual PPAR alpha/gamma agonist has clinically meaningful activity in vivo.

[The effects of inhaled nitric oxide on pulmonary vascular resistance in patients after total cavopulmonary connection].

OBJECTIVE: To study the effects of inhaled nitric oxide (NO) on pulmonary vascular resistance in patients after total cavopulmonary connection (TCPC). METHODS: Fifty-two patients after TCPC were evaluated, of them 24 patients were administered with inhaled nitric oxide in the early postoperative period. The cardiac index (CI) and pulmonary vascular resistance (PVR) were compared before and after inhaled NO. RESULTS: In experimental group, after inhaled NO, partial pressure of oxygen in artery/fraction of inspired oxygen increased from 161 +/- 17 to 193 +/- 23 (t = 2.75, P < 0.01); CI from (2.86 +/- 0.24) L.min(-1).m(-2) to (3.13 +/- 0.22) L.min(-1).m(-2) (t = 2.25, P < 0.05); PVR decreased from (4.2 +/- 0.5) U/m(2) to (3.8 +/- 1.4) U/m(2) (t = 2.29, P < 0.05); central venous pressure (CVP) from (17.0 +/- 1.8) mm Hg to (15.0 +/- 1.1) mm Hg, decreased 11.7%. Compared with the control group, respirator time decreased from (86 +/- 27) h to (54 +/- 18) h (t = 2.29, P < 0.05); ICU time from (6 +/- 2) d to (4 +/- 2) d (t = 2.32, P < 0.05); But hydrothorax drainage and length of stay had no significant difference. CONCLUSIONS: Though inhaled NO, there is no significant long-term effects in patients after TCPC, but it may play an important role in the management of low cardiac output syndrome and high cava pressure caused by reactive elevated pulmonary vascular resistance in the early postoperative period of TCPC.

Fully automated liquid-liquid extraction for the determination of a novel insulin sensitizer in human plasma by heated nebulizer and turbo ionspray liquid chromatography-tandem mass spectrometry.

I, 2-{[5,7-dipropyl-3-(trifluoromethyl)-1,2-benzisoxazol-6-yl]oxy}-2-methyl propionic acid is an alpha peroxisome proliferator-activated receptor (PPAR) agonist with some gamma activity being investigated for potential use in the treatment of Type II diabetes mellitus and dyslipidemia. Two automated liquid-liquid extraction methods were developed and validated for the determination of I in human plasma. Concentrations of I were determined over a wide range of clinical doses. For Method A, plasma was acidified and extracted with ethyl acetate using a fully automated procedure. Analysis was performed by LC-MS/MS with a turbo ionspray source in negative ion mode. For Method B, a larger volume of plasma was extracted and a heated nebulizer source was used on the mass spectrometer. Method A was linear from 0.05 to 50 ng/mL and Method B from 0.2 to 1000 ng/mL. Validation procedures showed that both methods were robust, specific and reproducible.

Quantitative determination of a novel dual PPAR alpha/gamma agonist using on-line turbulent flow extraction with liquid chromatography-tandem mass spectrometry.

Turbulent flow chromatograph (TFC) is a technique for the direct and efficient analysis of drugs and metabolites in biological matrices. We report here TFC on-line with an HPLC-MS/MS assay for the determination of 5-[2,4-dioxothiazolidin-5-yl)methyl]-2-methoxy-N-[[(4-trifluoromethyl)phenyl]methyl]benzamide (I, MK-0767, KRP297, Fig. 1) in plasma. Samples were transferred using an automated system followed by the addition of internal standard (II), prepared in 0.1 M ammonium acetate (pH 4.0). The plasma samples were directly injected onto a C18 turbulent flow column on-line with an HPLC-MS/MS system, and the analytical column used was a ThermoHypersil Keystone C18. Detection was achieved by MS/MS, using positive ionization on a TurboIonSpray probe, operated in multiple reaction monitoring (MRM) mode. The linear range was 4-2000 ng/mL for I when using 50 microL of plasma. The method exhibited good linearity and reproducibility. The method also showed good selectivity and ruggedness when applied to clinical samples, and was successfully cross-validated with a conventional off-line SPE, LC-MS/MS method.

Determination of MK-0767 enantiomers in human plasma by normal phase LC-MS/MS.

A sensitive and selective analytical method for the enantioselective determination of MK-0767, a dual peroxisome proliferator-activated receptor (PPAR) alpha/gamma agonist, in human plasma has been developed and validated. The chromatography is based on normal-phase chiral separation on a Kromasil, 5 microm, CHI-DMB 250 mm x 4.6 mm column. The detection involves the direct introduction of the normal phase eluent into MS/MS without the addition of a post-column reagent. Atmospheric pressure chemical ionization (APcI) mode was selected as the ion source in this method. With proper sample handling and processing procedures, ex vivo interconversion of the enantiomers was kept to minimum during sample collection, preparation and short term storage of frozen human plasma samples. The method was successfully utilized to determine the concentrations of MK-0767 enantiomers in human plasma to support pharmacokinetic investigation in man.

Quantitative determination of MK-0767, a dual alpha/gamma peroxisome proliferator-activated receptor (PPAR) agonist, in human plasma by liquid chromatography-tandem mass spectrometry.

5-[2,4-Dioxothiazolidin-5-yl)methyl]-2-methoxy-N-[[(4-trifluoromethyl)-phenyl]methyl]benzamide (I, MK-0767 or KRP-297, Fig. 1), is a dual alpha/gamma peroxisome proliferator-activated receptor (PPAR) agonist. A LC-MS/MS method for the determination of I in human plasma has been successfully developed, validated and applied to clinical programs. The analyte and internal standard (II) are extracted from 0.05 mL plasma via solid phase extraction (SPE). HPLC is used for the separation of I and II from possible co-extracted endogenous and other compounds. Detection is by MS/MS in multiple reaction monitoring (MRM) mode using a TurboIonSpray probe. The whole sample preparation is automated by using a Packard Multiprobe liquid handling system. The linear range is 4-2000 ng/mL in plasma. Recoveries were 71.1% and 69.4% for I and II, respectively. The method exhibited good linearity, reproducibility and sensitivity, selectivity and robustness when used for the analysis of clinical samples.

[Surgical treatment of secundum atrial septal defects in adults over 30 years old].

OBJECTIVE: To summarize the experience of surgical treatment of secundum atrial septal defects in adults over 30 years old. METHODS: There were 469 patients with secundum atrial septal defects in our study (male 144, female 325; ages 30-68, mean 38.6 years old). There were 105 cases with pulmonary hypertension and 458 cases with arrhythmia in the group. Surgical closure of defects were performed in all patients. Surgical closure of 358 cases were done by patches including 305 autologous pericardial patches. The low dose (6 x 10(-6)) nitric oxide inhalation was used in 25 postoperative patients with pulmonary hypertension. Right sided maze procedures were done in 5 cases with atrial fibrillation. RESULTS: Surgical mortality was 0.6% (3 cases), the others were healed. In the group, there were 180 cases with arrhythmia, 27 cases with left ventricular function amyoplasia, 28 cases with low cardiac output syndrome, 12 cases in secondary operation for bleeding and 1 case with air-embolism. The level of mean pulmonary artery pressure of 25 postoperative patients with pulmonary hypertension inhaled nitric oxide was down 28.5%. After right sided maze procedures were done in 5 cases with atrial fibrillation, atrial fibrillation disappeared. 352 cases were followed up from 3 months to 20 years (mean 5.6 years). Twenty-nine cases were in class I-II of cardiac function, and the others were better than class I of cardiac function. CONCLUSIONS: Atrial septal defects in adult should be operated as early as possible. When patch is needed, an autologous pericardial patch is the first selection. Inhaled nitric oxide is an effective method to postoperative pulmonary hypertension. The maze operation should be performed for atrial septal defect with atrial fibrillation while the surgical closure of defect was done. During and after operation, much attention should be paid to preventing and curing arrhythmia and protecting and supporting left heart function.

Quantitative determination of a novel insulin sensitizer and its para-hydroxylated metabolite in human plasma by LC-MS/MS.

I, 5-[3-[3-(4-phenoxy-2-propylphenoxy)-propoxy]-phenyl]-2,4-thiazolidinedione sodium salt, is a dual alpha/gamma peroxisome proliferator-activated receptor (PPAR) agonist for potential use in diabetic patients. The compound has a para-hydroxylated metabolite, II, which has also been shown to exhibit PPAR activity. An LC-MS/MS method for the simultaneous determination of I and its active metabolite (II) in human plasma has been successfully developed. The method consists of treating 0.5 ml plasma with ammonium acetate (pH 9.6; 50mM) and extracting I, II and internal standard (III, Fig. 2) with 5 ml ethyl acetate. The ethyl acetate is evaporated and the samples are reconstituted in 0.1 ml acetonitrile:0.1% formic acid (65:35, v/v). The entire extraction procedure, as well as sample collection, was performed in glass tubes and vials to overcome the analytes adherence to polypropylene. A linear HPLC gradient was used to separate the analyte, metabolite, internal standard, and other interfering, non-quantitated metabolites. Detection was by negative ionization MS/MS on a turbo ionspray probe. Precursor-->product ion combinations were monitored in multiple reaction monitoring (MRM) mode. The linear range is 0.05-20 ng/ml for I and 0.1-20 ng/ml for II. Recoveries were 59.4, 90.1 and 56.8% for I, II and III, respectively. Intraday variation using this method was <==7.0% for I and <==9.2% for II. The method exhibits good linearity and reproducibility for each analyte and good sensitivity, selectivity and robustness when used for the analysis of clinical samples.

Simultaneous determination of losartan and EXP3174 in human plasma and urine utilizing liquid chromatography/tandem mass spectrometry.

Losartan is an orally active angiotensin II receptor antagonist indicated for the treatment of hypertension. EXP3174 is an active metabolite, which contributes to the overall activity of losartan. Analytical methods for the simultaneous determination of losartan and its active metabolite EXP3174 in human plasma and urine with limited plasma sample size have been developed and validated to support a pediatric clinical program. In both methods, analytes are extracted from the matrixes by liquid-liquid extraction and separated using reverse phase high-performance liquid chromatography (HPLC). A tandem mass spectrometer (MS/MS) with a Turbo ionspray (TIS) interface in multiple-reaction-monitoring (MRM) mode is used for detection of the analytes in both methods. The plasma method has a lower limit of quantitation (LOQ) of 1 ng/ml with a linearity range of 1-500 ng/ml for losartan and EXP3174 using 100 microl of plasma. For the urine method, the LOQ for both losartan and EXP3174 is 2 ng/ml using 0.5 ml of urine, and the linearity range for both analytes is 2-1000 ng/ml. Validation procedures have proven that both methods are robust, accurate, and reproducible. Both methods have been used to assay clinical samples and provided satisfactory results.

[Levels of plasma endothelin, nitric oxide and atrial natriuretic peptide after cardiopulmonary bypass and influence of inhaled nitric oxide in patients with ventricular septal defect and pulmonary hypertension].

OBJECTIVES: To assess the changes of the levels of plasma endothelin (ET-1), nitric oxide (NO) and atrial natriuretic peptide (ANP) after cardiopulmonary bypass (CPB) and the influence of inhaled nitric oxide in patients with ventricular septal defect (VSD) and pulmonary hypertension (PH). METHODS: Sixty patients with VSD were enrolled in this study. They were divided into 2 groups: group A [no-PH group, mean pulmonary artery pressure (mPAP) < 20 mm Hg (1 mm Hg = 0.133 kPa) n = 20] and group B (PH group, mPAP > 20 mm Hg, n = 40). Group B was subdivided into two groups by randomized block, group B(1) (inhaled NO group, n = 20) and group B(2) (contrast group, n = 20). The plasma ET-1, NO, ANP concentrations were assayed at 24 h pre-operation and 0 h, 1 h, 5 h, 12 h, 24 h, 48 h after CPB. RESULTS: The preoperative plasma ET-1, NO and ANP concentrations in group B were significantly higher than those in group A. In three groups, the plasma ET-1 concentration at 0 h after CPB was significantly higher than that at 24 h pre-operation, and the plasma NO concentration at 0 h after CPB was significantly lower than that at 24 h pre-operation. In group B, the plasma ANP concentration at 0 h after CPB was significantly higher than that at 24 h pre-operation. After CPB, the plasma ET-1 concentration in group B(1) decreased faster than that in group B(2), and the plasma NO concentration in group B(1) increased faster than that in group B(2). In group B, the preoperative plasma ET-1 concentration negatively correlated with the preoperative plasma NO concentration and positively correlated with the preoperative ANP concentration. CONCLUSIONS: The broken dynamic balance of ET-1/NO may take part in generation and development of pulmonary hypertension. ANP acts as a favorable physiological regulating factor in the pathogenesis of pulmonary hypertension. CPB can regulate the level of ET-1 up and NO and ANP down while inhaled NO can cause the level of ET-1 down and the level of NO up.

Simultaneous determination of a novel M3 muscarinic receptor antagonist and its active 5-OH metabolite in human plasma using liquid chromatography/tandem mass spectrometry.

A sensitive, specific, and robust liquid chromatography (LC)/mass spectrometry (MS)/MS method has been developed and validated for a novel M(3) muscarinic receptor antagonist (I) and its active 5-OH metabolite (II) in human plasma. The assay involves a two-step liquid-liquid extraction of the compounds from human plasma, high performance liquid chromatography (HPLC) separation, and MS/MS for the detection of the analytes. The method provides a linear response from a quantitation limit of 0.05-20 ng/ml for I and 0.1-20 ng/ml for II using 1 ml of plasma. The mean absolute recovery was 85.4% for I and 80.8% for II, respectively. The intra-assay accuracy of I and II averaged from 95.0 to 105.3% with coefficient of variation (CV) values