Serum progesterone is lower in ovarian stimulation with highly purified HMG compared to recombinant FSH owing to a different regulation of follicular steroidogenesis: a randomized controlled trial.

STUDY QUESTION: Does ovarian stimulation with highly purified (hp)-HMG protect from elevated progesterone in the follicular phase compared to recombinant FSH (r-FSH) cycles through a different regulation of follicular steroidogenesis? SUMMARY ANSWER: hp-HMG enhanced the Δ4 pathway from pregnenolone to androstenodione leading to lower serum progesterone at the end of the cycle, while r-FSH promoted the conversion of pregnenolone to progesterone causing higher follicular phase progesterone levels. WHAT IS KNOWN ALREADY: Elevated progesterone in the follicular phase has been related to lower clinical outcome in fresh IVF cycles. Progesterone levels are positively correlated to ovarian response, and some studies have shown that when r-FSH alone is used for ovarian stimulation serum progesterone levels on the day of triggering are higher than when hp-HMG is given. Whether this is caused by a lower ovarian response in hp-HMG cycles or to a difference in follicular steroidogenesis in the two ovarian stimulation regimens has not been well characterized. STUDY DESIGN, SIZE, DURATION: A randomized controlled trial including 112 oocyte donors undergoing ovarian stimulation with GnRH antagonists and 225 IU/day of r-FSH (n = 56) or hp-HMG (n = 56) was carried out in a university-affiliated private infertility clinic. Subjects were recruited between October 2016 and June 2018. PARTICIPANTS/MATERIALS, SETTING, METHODS: The women were aged 18-35 years with a regular menstrual cycle (25-35 days) and normal ovarian reserve (serum anti-Müllerian hormone (AMH) = 10-30 pMol/l) undergoing ovarian stimulation for oocyte donation. FSH, LH, estradiol (E2), estrone, progesterone, pregnenolone, 17-OH-progesterone, androstenodione, dehidroepiandrostenodione, and testosterone were determined on stimulation Days 1, 4, 6, and 8 and on day of triggering in serum and in follicular fluid. Samples were frozen at -20°C until assay. Total exposures across the follicular phase were compared by polynomic extrapolation. MAIN RESULTS AND THE ROLE OF CHANCE: Subjects in both groups were comparable in terms of age, BMI, and AMH levels. Ovarian response was also similar: 17.5 ± 7.9 (mean ± SD) versus 16.5 ± 7.5 oocytes with r-FSH and hp-HMG, respectively (P = 0.49). Serum progesterone (ng/ml) on day of trigger was 0.46 ± 0.27 in the hp-HMG group versus 0.68 ± 0.50 in the r-FSH group (P = 0.010). Differences for progesterone were also significant on stimulation days 6 and 8. The pregnenolone: progesterone ratio was significantly increased in the r-FSH group from stimulation day 8 to the day of trigger (P = 0.019). Serum androstenodione (ng/ml) on day of trigger was 3.0 ± 1.4 in the hp-HMG group versus 2.4 ± 1.1 in the r-FSH group (P = 0.015). Differences in adrostenodione were also significant on stimulation Day 8. The pregnenolone:androstenodione ratio was significantly higher in the hp-HMG group (P = 0.012) on Days 6 and 8 and trigger. There were no other significant differences between groups. Follicular fluid E2, FSH, LH, dehidroepioandrostenodione, androstenodione, and testosterone were significantly higher in the hp-HMG than r-FSH group. No differences were observed for progesterone, estrone, 17-OH-progesterone, and pregnenolone in follicular fluid. LIMITATIONS, REASONS FOR CAUTION: All women included in the study were young, not infertile, and had a normal BMI and a good ovarian reserve. The findings might be different in other patient subpopulations. Hormone analyses with immunoassays are subject to intra-assay variations that may influence the results. WIDER IMPLICATIONS OF THE FINDINGS: Stimulation with hp-HMG may prevent progesterone elevation at the end of the follicular phase because of a different follicular steroidogenesis pathway, regardless of ovarian response. This should be considered, particularly in patients at risk of having high progesterone levels at the end of the follicular phase when a fresh embryo transfer is planned. STUDY FUNDING/COMPETING INTEREST(S): Roche Diagnostics provided unrestricted funding for all serum and follicular fluid hormone determinations. J.L.R., M.M., and A.P. have nothing to declare. E.B. has received consulting fees from Ferring, Merck, Gedeon Richter, and Roche and has participated in a research cooperation with Gedeon-Richter. In addition, the author has participated in speakers' bureau and received fees from Ferring, Gedeon Richter, Merck, and Roche. P.A. has received consulting fees from MSD and has participated in speakers' bureau and received fees from Ferring. P.A. also declares travel/meeting support from MSD. E.L. has received consulting fees from Ferring and MSD. In addition, the author has participated in a research cooperation with Gedeon-Richter. Also, the author has participated in speakers' bureau and received fees from Ferring and IBSA, as well as travel/meeting support from IBSA and Gedeon Richter. E.B., P.A., and E.L. also own stocks in IVIRMA Valencia. TRIAL REGISTRATION NUMBER: NCT: NCT02738580. TRIAL REGISTER DATE: 19 February 2016. DATE OF FIRST PATIENT'S ENROLMENT: 03 October 2016.

Use of Micellar Liquid Chromatography to Determine Mebendazole in Dairy Products and Breeding Waste from Bovine Animals.

Mebendazole is an anthelmintic drug used in cattle production. However, residues may occur in produced food and in excretions, jeopardizing population health. A method based on micellar liquid chromatography (MLC) was developed to determine mebendazole in dairy products (milk, cheese, butter, and curd) and nitrogenous waste (urine and dung) from bovine animals. Sample treatment was expedited to simple dilution or solid-to-liquid extraction, followed by filtration and direct injection of the obtained solution. The analyte was resolved from matrix compounds in less than 8 min, using a C18 column and a mobile phase made up of 0.15 M sodium dodecyl sulfate (SDS)-6% 1-pentanol phosphate buffered at pH 7, and running at 1 mL/min under isocratic mode. Detection was performed by absorbance at 292 nm. The procedure was validated according to the guidelines of the EU Commission Decision 2002/657/EC in terms of: specificity, method calibration range (from the limit of quantification to 25-50 ppm), sensitivity (limit of detection 0.1-0.2 ppm; limit of quantification, 0.3-0.6 ppm), trueness (92.5-102.3%), precision (<7.5%, expressed at RSD), robustness, and stability. The method is reliable, sensitive, easy-to-handle, eco-friendly, safe, inexpensive, and provides a high sample-throughput. Therefore, it is useful for routine analysis as a screening or quantification method in a laboratory for drug-residue control.

Procedure for the Screening of Eggs and Egg Products to Detect Oxolonic Acid, Ciprofloxacin, Enrofloxacin, and Sarafloxacin Using Micellar Liquid Chromatography.

A method based on micellar liquid chromatography was developed to determine oxolinic acid, ciprofloxacin, enrofloxacin, and sarafloxacin in eggs and egg products. The antimicrobial drugs were obtained in a micellar solution which was directly injected. The analytes were resolved using a C18 column and a mobile phase of 0.05 M sodium dodecyl sulfate-7.5% 1-propanol-0.5% triethylamine, buffered at pH 3 with phosphate salt, running under the isocratic mode. The signal was monitored by fluorescence. Validation was successfully performed according to the EU Commission Decision 2002/657/EC in terms of specificity, calibration range (LOQ to 1 mg/kg), linearity (R2 > 0.9991), limit of detection and decision limit (0.01-0.05 mg/kg), limit of quantification (0.025-0.150 mg/kg), detection capability (<0.4 times decision limit), trueness (-14.2% to +9.8%), precision (<14.0%), robustness, and stability. The procedure was environmentally friendly, safe, easy-to-conduct, inexpensive, and had a high sample throughput, thus it is useful for routine analysis as a screening method in a laboratory for food residue control.

Micellar liquid chromatography determination of rivaroxaban in plasma and urine. Validation and theoretical aspects.

A Micellar Chromatographic method to determine rivaroxaban in plasma and urine has been developed. The samples were dissolved in the mobile phase (SDS 0.05 M - 1-propanol 12.5%, phosphate buffered at pH 7) and 20 µL directly injected, avoiding the extraction and purification steps. Using a C18 column and running under isocratic mode at 1 mL/min, analyte was eluted without interference from the matrix in <6.0 min. The detection absorbance wavelength was set to 250 nm. The procedure was validated by Food and Drug Administration guidelines in terms of: system suitability, calibration range (0.05-5 mg/L), linearity, sensitivity, robustness, carry-over effect, specificity, accuracy (-11.1 to 4.2%), precision (<19.9%), stability and analysis of incurred samples. The method was found reliable, practical, easy-to-conduct, rapid, relatively eco-friendly, safe, inexpensive, widely available and with a high sample throughput. The method was applied to the analysis of incurred samples, including incurred sample reanalysis, to verify that the instrumentation works correctly. In addition, the constants of the different partition equilibria occurring in the column were elucidated in order to have a better comprehension of the theoretical aspects of the retention mechanism. A moderately strong association between rivaroxaban and the stationary phase and the micelles was found, weakened by short chain alcohol.

Stability studies of rifampicin in plasma and urine of tuberculosis patients according to the European Medicines Agency Guidelines.

Aim: The macrolide antibiotic rifampicin is prescribed against several infections, like tuberculosis disease. This drug decays to rifampicin quinone. Results/methodology: The biological fluids were diluted in a micellar solution and directly injected. Using a C18 column and a mobile phase of 0.15 M SDS-6% 1-pentanol phosphate-buffered at pH 7, running at 1 ml/min, the analytes were resolved in less than 15 min. The detection was by absorbance at 337 nm. Method was validated by the guidelines of the European Medicines Agency. Decomposition of rifampicin to rifampicin quinone was also studied. Discussion/conclusion: Procedure is rapid, easy-to-handle, economic, eco-friendly and with a high sample throughput. It was successfully used to monitor rifampicin in the plasma and urine of tubercular patients.

Variability of revascularization techniques among Catalan hospitals and impact on leg salvage in patients with peripheral arterial disease.

BACKGROUND: Indications for the revascularization treatment of peripheral arterial disease (PAD) generate much discussion, and practice varies significantly among hospitals. This study looked at patients with PAD admitted to all hospitals of the Catalan Health Service and analyzed patterns of revascularization techniques with subsequent amputation procedures. METHODS: We used the clinical-administrative registry of admissions of all patients in the hospitals of Catalonia, north-east Spain, between 2009 and 2014. We analyzed the clinical course of patients admitted with PAD throughout their successive hospital admissions. Variability between hospitals was described for the revascularization techniques and amputations performed. Endovascular outcomes were compared with those from open surgery. RESULTS: Annually, there were 9,828 admissions with PAD and 631 major amputations. Eight hospitals accounted for 52% of all admissions, and endovascular techniques occurred predominantly in high-tech, high-resolution or reference hospitals. The ratio of admissions involving endovascular techniques/open surgery varied from 0.02 to 3.73 according to the hospital, and had a correlation of -0.175 (P=0.447) with the percentage of performed major amputations and of 0.122 (P=0.598) ratio of minor / major amputations. At the end of the 6 studied years, endovascular revascularization resulted in lower patency and more minor amputations than open surgery, but had the same percentage of major amputations (10.3% vs. 10.7%, P=0.526) and lower in-hospital mortality (7.1% vs. 9.5%, P<0.0001). CONCLUSIONS: Interventions of PAD are centralized in complex hospitals and have an important variability depending on the treating hospital. Hospital variability in revascularization techniques seems to have no impact on leg salvage. Endovascular and surgical revascularization would result in similar percentages of major amputations.

A rapid and reliable assay to determine flumequine, marbofloxacin, difloxacin, and sarafloxacin in commonly consumed meat by micellar liquid chromatography.

BACKGROUND: Micellar liquid chromatography - fluorescence detection was used to determine the antibiotics flumequine, marbofloxacin, difloxacin, and sarafloxacin in porcine, bovine, poultry, ovine, caprine, rabbit, and equine meat, to verify compliance with EU Regulation 37/2010 with regard to the occurrence of veterinary drugs in food. RESULTS: The analytes were isolated from the matrix by ultrasonication-assisted leaching in a micellar solution, and the supernatant was filtered and directly injected. The fluoroquinolones were resolved in < 19 min using a C18 column, with an isocratic mobile phase of 0.05 mol L-1 sodium dodecyl sulfate - 8% 1-butanol - 0.5% triethylamine buffered at pH 3. The limits of quantification (0.01-0.05 mg kg-1 ) were below the maximum residue limits (0.15-0.4 mg kg-1 ). The method was validated by EU Commission Decision 2002/657/EC guidelines. CONCLUSION: The method shows practical advantages such as simplicity, low cost, eco-friendliness, safety, and applicability for routine analysis, and is useful for surveillance programs. © 2018 Society of Chemical Industry.

Optimization and Validation of a Chromatographic Method for the Quantification of Isoniazid in Urine of Tuberculosis Patients According to the European Medicines Agency Guideline.

Isoniazid is a drug that is widely used against tuberculosis. However, it shows high interpatient variability in metabolism kinetics and clinical effect, which complicates the prescription of the medication and jeopardizes the success of the therapy. Therefore, in a specific patient, the pharmacokinetics of the drug must be elucidated to decide the proper dosage and intake frequency to make the drug suitable for therapeutic drug monitoring. This can be performed by the quantification of the drug in urine as this process is non-invasive and allows the effects of long-time exposure to be inferred. The paper describes the development of a micellar liquid chromatographic method to quantify isoniazid in urine samples. Extraction steps were avoided, making the procedure easy to handle and reducing the waste of toxic organic solvents. Isoniazid was eluted in less than 5 min without interference from other compounds of the urine using a mobile phase containing 0.15 SDS⁻12.5% 1-propanol (v/v)⁻Na2HPO4 0.01 M buffered at pH 7, running at 1 mL/min under isocratic mode through a C18 column with the detection wavelength at 265 nm. The method was validated by following the requirements of the Guidelines on Bioanalytical Method Validation issued by the European Medicines Agency (EMA) in terms of selectivity, calibration curve (r² = 0.9998 in the calibration range (0.03⁻10.0 µg/mL), limit of detection and quantification (10 and 30 ng/mL respectively), precision (<16.0%), accuracy (-0.9 to +8.5%), carry-over, matrix effect, and robustness. The developed method was applied to quantify isoniazid in urine samples of patients of an Indian hospital with good results. The method was found to be useful for routine analysis to check the amount of isoniazid in these patients and could be used in its therapeutic monitoring.

Development of a method to determine axitinib, lapatinib and afatinib in plasma by micellar liquid chromatography and validation by the European Medicines Agency guidelines.

A method based on micellar liquid chromatography to quantify the tyrosine kinase inhibitors axitinib, lapatinib and afatinib in plasma is reported. The sample pretreatment was a simple 1/5-dilution in a pure micellar solution, filtration and direct injection, without requiring extraction or purification steps. The three drugs were resolved from the matrix in 17min, using an aqueous solution of 0.07M sodium dodecyl sulfate - 6.0% 1-pentanol, buffered at pH7 with 0.01M phosphate salt as mobile phase, running under isocratic mode at 1mL/min through a C18 column. The detection was performed by absorbance at 260nm. An accurate mathematical relationship was established between the retention factor of each drug and the surfactant/organic solvent concentration in the mobile phase, achieved with a limited number of experiments, in order to optimize these factors. A binding behavior of the analytes face to the micelles was found out. The method was successfully validated by the guidelines of the European Medicines Agency in terms of: selectivity, linearity (r2>0.9995), calibration range (0.5 to 10mg/L), limit of detection (0.2mg/L), carry-over effect, accuracy (-8.1 to +6.9%), precision (<13.8%), dilution integrity, matrix effect, stability and robustness. The procedure was found reliable, practical, economic, accessible, short-time, easy-to-handle, inexpensive, environmental-friendly, safe, useful for the analysis of many samples per day. Finally, the method was applied to the analysis of incurred, using quality control samples in the same analytical run, with adequate results. Therefore, it can be implementable for routine analysis in clinical laboratories.

Validation of a procedure to quantify oxolinic acid, danofloxacin, ciprofloxacin and enrofloxacin in selected meats by micellar liquid chromatography according to EU Commission Decision 2002/657/EC.

The suitability of an analytical method to determine oxolinic acid, danofloxacin, ciprofloxacin and enrofloxacin in edible tissues, based on micellar liquid chromatography coupled with fluorescence detection, to be applied in chicken, turkey, duck, lamb, goat, rabbit and horse muscle, is described. The method was fully matrix-matched in-lab revalidated, for each antimicrobial drug and meat, following the guidelines of the EU Commission Decision 2002/657/EC. The permitted limits were the maximum residue limits stated by the EU Commission Regulation 37/2010. The results obtained for the studied validation parameters were in agreement with the guidelines: selectivity (the antibiotics were resolved), linearity (r2  > 0.995), limit of detection (0.004-0.02 mg/kg), limits of quantification (0.01-0.05 mg/kg), calibration range (up to 0.5 mg/kg), recovery (89.5-105.0%), precision (<8.3%), decision limit, detection capability, ruggedness, stability and application to incurred samples. The method was found to be able to provide reliable concentrations with low uncertainty within a large interval, including the maximum residue limits, and then was useful to find out prohibited contaminated samples. The method did not require to be adapted for these matrices, and then it maintained its interesting advantages: short-time, eco-friendly, safe, inexpensive, easy-to-conduct, minimal manipulation and useful for routine analysis.

Ingresos hospitalarios de desintoxicación en mujeres con alcoholismo y otras toxicomanías / Hospitalized inpatient detoxification of women with alcoholism and other drug addictions

Objetivos: Analizar las características descriptivas de los ingresos de Desintoxicación por Alcoholismo u otras Toxicomanías en mujeres que requieren un ingreso hospitalario de desintoxicación en nuestro medio. Diseño: Retrospectivo, incluyendo todas las mujeres ingresadas en la Unidad de Desintoxicación de la Corporació Sanitària i Universitària Parc Taulí de Sabadell (Barcelona) entre enero de 2010 y junio 2013. Utilizamos criterios diagnósticos DSM-IV-TR y una estadística descriptiva. De forma secundaria, se compararon mujeres que habían requerido un ingreso por alcohol como sustancia principal de desintoxicación (APD) y aquellas con alcohol como sustancia única de desintoxicación (AUD) en relación al resto de la muestra. Resultados: Desde Enero de 2010 a Junio de 2013 se realizaron 360 ingresos en la Unidad de Desintoxicación, de los cuales 82 fueron mujeres (22,7%). Se trataba de mujeres de edad media (43,3 años) que ingresaron principalmente para la desintoxicación de más de una sustancia. El tóxico principal fue el alcohol con 67,1% (n=55), seguido de las politoxicomanías que incluyeron casos con abuso de varias sustancias como motivo principal de desintoxicación. A nivel de otra comorbilidad psiquiátrica, la mayoría de las mujeres no presentaban ningún diagnostico DSM-IV-TR (53,7%, n=44). Las mujeres AUD (n=33) fueron significativamente mayores de edad, utilizaron más benzodiacepinas y tuvieron menos diagnósticos en el Eje II. Conclusiones: El principal motivo de desintoxicación en mujeres de nuestro medio es el alcohol. Existen diferencias significativas entre las mujeres con desintoxicación de alcohol y aquellas con consumos comórbidos de otras sustancias

Objectives: To analyze the characteristics of inpatient detoxification hospitalizations for alcoholism and other drug addictions in women from our area. Design: We use a retrospective design. The subjects were inpatient women hospitalized in the Detoxification Unit at the Corporació Sanitària i Universitària Parc Taulí de Sabadell (Barcelona) between January 2010 and June 2013. Diagnoses were made by DSM-IV-TR criteria. The statistics were descriptive. In a secondary analysis, we compared women with alcohol as a principal reason of admission (APD) with women with alcohol as an isolated reason of admission (AUD) in relation with the rest of the sample. Results: From January 2010 to June 2013 there were 360 admissions in the Detoxification Unit, 82 of whom were women (22.7%). These were middle-aged women (43.3 years), who were mainly admitted for detoxification from more than one substance. The main drug involved in the abuse was alcohol (67.1%, n=55), followed by multiple drug abuse as a main reason for detoxification. Considering psychiatric comorbidity, most of the women did not display any DSM-IV-TR diagnosis criteria (53.7%,= 44). Women in AUD (n=33) were significantly older, used more benzodiazepines and had fewer Axis II diagnoses. Conclusions: The main reason for inpatient detoxification in women in our area is alcohol. There are significant differences between women with alcohol detoxifications and women with comorbid abuse of other substances

Development and validation of a micellar liquid chromatographic method to determine three antitumorals in plasma.

AIM: A micellar liquid chromatographic method to determine several anticancer drugs (pazopanib, dabrafenib and regorafenib) in plasma was developed and validated by the guidelines of the EMA. EXPERIMENTAL: Plasma samples were directly injected, after a 1/5-dilution in a micellar solution. The drugs were resolved in <18 min using a C18 column. The mobile phase was an aqueous solution of 0.12 M SDS - 2% 1-pentanol, buffered at pH 7. The detection was performed by absorbance at 260 nm. RESULTS: The values of the main validation parameters were: LOD (0.1-1 mg/l), calibration range (0.2-2 to 80 mg/l), accuracy (-12.5 to +11.7%) and precision (<11.9%). CONCLUSION: The procedure was conducted by minimum cost, effort, manipulation, time and quantity of hazardous chemicals. The method was useful to determine the drugs at their respective target concentrations, and was found useful for clinical analysis.

Determination of oxolinic acid, danofloxacin, ciprofloxacin, and enrofloxacin in porcine and bovine meat by micellar liquid chromatography with fluorescence detection.

A method was developed for the determination of oxolinic acid, danofloxacin, ciprofloxacin and enrofloxacin by micellar liquid chromatography - fluorescence detection in commercial porcine and bovine meat. The samples were ultrasonicated in a micellar solution, free of organic solvent, to extract the analytes, and the supernatant was directly injected. The quinolones were resolved in <22min using a mobile phase of 0.05M SDS - 7.5% 1-propanol - 0.5% triethylamine buffered at pH 3, running through a C18 column at 1mL/min using isocratic mode. The method was validated by the in terms of: selectivity, calibration range (0.01-0.05 to 0.5mg/kg), linearity (r2>0.9998), trueness (89.3-105.1%), precision (<8.3%), decision limit (<12% over the maximum residue limit), detection capability (<21% over the maximum residue limit), ruggedness (<5.6%) and stability. The procedure was rapid, eco-friendly, safe and easy-to-handle.

Determination of diuron, terbuthylazine, and terbutryn in wastewater and soil by micellar liquid chromatography.

An analytical method for the quantification of the herbicides and algaecides diuron, terbuthylazine, and terbutryn in wastewater and soil by micellar liquid chromatography was developed. The sample preparation was expedited to reduce the number of intermediate steps and the use of chemicals. The analytes in soils were recovered by ultrasonication in the mobile phase. The obtained supernatant and the water samples were directly injected, thus avoiding intermediate steps. The chromatographic behavior of the analytes, depending on the surfactant and alcohol was studied, in order to optimize the chromatographic run, by a chemometrical approach. The herbicides were resolved in <16 min using a C18 column and a mobile phase of 0.07 M sodium dodecyl sulfate/6% 1-pentanol phosphate buffered at pH 3, running under isocratic mode at 1 mL/min. The detection absorbance wavelength was set to 240 nm. The method was successfully validated in terms of selectivity, detection limit (0.06 mg/L in water and 0.3 mg/kg in soil), quantitation range (0.2-2 mg/L in water and 1-10 mg/kg in soil), trueness (-6.1 to +5.0%), precision (<9.4%), and ruggedness (<8.3%). The procedure was reliable, practical, easy-to-handle, available, short-time and ecofriendly and useful for routine analysis. Its applicability to real samples was evaluated by analyzing several wastewater, decorative reservoir, and soil samples from agricultural and urban sources.

A review on development of analytical methods to determine monitorable drugs in serum and urine by micellar liquid chromatography using direct injection.

Therapeutic drug monitoring is a common practice in clinical studies. It requires the quantification of drugs in biological fluids. Micellar liquid chromatography (MLC), a well-established branch of Reverse Phase-High Performance Liquid Chromatography (RP-HPLC), has been proven by many researchers as a useful tool for the analysis of these matrices. This review presents several analytical methods, taken from the literature, devoted to the determination of several monitorable drugs in serum and urine by micellar liquid chromatography. The studied groups are: anticonvulsants, antiarrhythmics, tricyclic antidepressants, selective serotonin reuptake inhibitors, analgesics and bronchodilators. We detail the optimization strategy of the sample preparation and the main chromatographic conditions, such as the type of column, mobile phase composition (surfactant, organic solvent and pH), and detection. The finally selected experimental parameters, the validation, and some applications have also been described. In addition, their performances and advantages have been discussed. The main ones were the possibility of direct injection, and the efficient chromatographic elution, in spite of the complexity of the biological fluids. For each substance, the measured concentrations were accurate and precise at their respective therapeutic range. It was found that the MLC-procedures are fast, simple, inexpensive, ecofriendly, safe, selective, enough sensitive and reliable. Therefore, they represent an excellent alternative for the determination of drugs in serum and urine for monitoring purposes.

Use of micellar liquid chromatography for rapid monitoring of fungicides post harvest applied to citrus wastewater.

A method based on micellar liquid chromatography has been developed to simultaneously monitor four pesticides largely post-harvest applied to citrus: thiabendazole, pyrimethanil, o-phenylphenol and imazalil. Water samples were filtered and directly injected without other treatment, thus avoiding extraction steps. The composition of the mobile phase was optimized using a chemometrical approach to achieve and excellent resolution to 0.07 mol/L SDS/5%, V/V 1-pentanol buffered at pH3. Mobile phase run through a C18 column at 1 mL/min at room temperature. The detection was performing by UV-Visible absorbance using a wavelength program: 0-10 min, 305 nm (for thiabendazole); 10-12; 265 nm (for pyrimethanil) and 12-18, 220 nm (o-phenylphenol and imazalil). The developed method was validated following the guidelines of the US Environmental Protection Agency in terms of: quantitation range, (0.5-4 to 15 µg/mL), linearity (r(2)>0.9995), sensitivity (LOD, 0.18-1.4 µg/mL), precision (<9.2%), trueness (93.9%-103.7%), and ruggedness (<9.9%). It was found that the fungicides remain up to eight days in surface water at outdoor conditions. The method was used to screen the presence of the analytes in several waste water samples, and was proved to be useful in routine analysis.

Development and validation of a method to determine thiabendazole and o-phenylphenol in wastewater using micellar liquid chromatography-fluorescence detection.

A micellar liquid chromatographic method to determine thiabendazole (TBZ) and o-phenylphenol in wastewater is described here. The sample was directly injected without any additional treatment other filtration. The pesticides were resolved in <11 min, using a mobile phase of 0.10 M SDS-6% 1-pentanol buffered at pH 3 running through a C18 column at 1 mL/min. The detection was performed by fluorescence at 305/360 and 245/345 nm excitation/emission wavelengths for TBZ and o-phenylphenol, respectively. The method was validated following the directives of the Validation and Peer Review of U.S. Environmental Protection Agency Chemical Methods of Analysis guidelines in terms of selectivity, quantitation range (0.01-0.02 to 2 mg/L), detection limit (0.005-0.008 mg/L), trueness (92.1-104.2%), precision (<13.9%), robustness (<6.6%), and stability under storage conditions. The procedure was applied to the screening of TBZ and o-phenylphenol in wastewater samples from citrus packing plants, agricultural gutters, urban sewage, as well as in influent and effluent wastewater treatment plants.

The Metabolomic Profile of Spent Culture Media from Day-3 Human Embryos Cultured under Low Oxygen Tension.

Despite efforts made to improve the in vitro embryo culture conditions used during assisted reproduction procedures, human embryos must adapt to different in vitro oxygen concentrations and the new metabolic milieu provided by the diverse culture media used for such protocols. It has been shown that the embryo culture environment can affect not only cellular metabolism, but also gene expression in different species of mammalian embryos. Therefore we wanted to compare the metabolic footprint left by human cleavage-stage embryos under two types of oxygen atmospheric culture conditions (6% and 20% O2). The spent culture media from 39 transferred and implanted embryos from a total of 22 patients undergoing egg donation treatment was analyzed; 23 embryos came from 13 patients in the 6% oxygen concentration group, and 16 embryos from 9 patients were used in the 20% oxygen concentration group. The multivariate statistics we used in our analysis showed that human cleavage-stage embryos grown under both types of oxygen concentration left a similar metabolic fingerprint. We failed to observe any change in the net depletion or release of relevant analytes, such as glucose and especially fatty acids, by human cleavage-stage embryos under either type of culture condition. Therefore it seems that low oxygen tension during embryo culture does not alter the global metabolism of human cleavage-stage embryos.