Unmethylated Insulin DNA Is Elevated After Total Pancreatectomy With Islet Autotransplantation: Assessment of a Novel Beta Cell Marker.

Beta cell death may occur both after islet isolation and during infusion back into recipients undergoing total pancreatectomy with islet autotransplantation (TPIAT) for chronic pancreatitis. We measured the novel beta cell death marker unmethylated insulin (INS) DNA in TPIAT recipients before and immediately after islet infusion (n = 21) and again 90 days after TPIAT, concurrent with metabolic functional assessments (n = 25). As expected, INS DNA decreased after pancreatectomy (p = 0.0002). All TPIAT recipients had an elevated unmethylated INS DNA ratio in the first hours following islet infusion. In four samples (three patients), INS DNA was also assessed immediately after islet isolation and again before islet infusion to assess the impact of the isolation process: Unmethylated and methylated INS DNA fractions both increased over this interval, suggesting death of beta cells and exocrine tissue before islet infusion. Higher glucose excursion with mixed-meal tolerance testing was associated with persistently elevated INS DNA at day 90. In conclusion, we observed universal early elevations in the beta cell death marker INS DNA after TPIAT, with pronounced elevations in the islet supernatant before infusion, likely reflecting beta cell death induced by islet isolation. Persistent posttransplant elevation of INS DNA predicted greater hyperglycemia at 90 days.

Sitagliptin Treatment After Total Pancreatectomy With Islet Autotransplantation: A Randomized, Placebo-Controlled Study.

Insulin independence after total pancreatectomy and islet autotransplant (TPIAT) for chronic pancreatitis is limited by a high rate of postprocedure beta cell apoptosis. Endogenous glucagon-like peptide-1 and glucose-dependent insulinotropic peptide, which are increased by dipeptidyl peptidase 4 inhibitor therapy (sitagliptin) may protect against beta cell apoptosis. To determine the effect of sitagliptin after TPIAT, 83 adult TPIAT recipients were randomized to receive sitagliptin (n = 54) or placebo (n = 29) for 12 months after TPIAT. At 12 and 18 months after TPIAT, participants were assessed for insulin independence; metabolic testing was performed with mixed meal tolerance testing and frequent sample intravenous glucose tolerance testing. Insulin independence did not differ between the sitagliptin and placebo groups at 12 months (42% vs. 45%, p = 0.82) or 18 months (36% vs. 44%, p = 0.48). At 12 months, insulin dose was 9.0 (standard error 1.7) units/day and 7.9 (2.2) units/day in the sitagliptin and placebo groups, respectively (p = 0.67) and at 18 months 10.3 (1.9) and 7.1 (2.6) units/day, respectively (p = 0.32). Hemoglobin A1c levels and insulin secretory measures were similar in the two groups, as were adverse events. In conclusion, sitagliptin could be safely administered but did not improve metabolic outcomes after TPIAT.

Metabolic assessment prior to total pancreatectomy and islet autotransplant: utility, limitations and potential.

Islet autotransplant (IAT) may ameliorate postsurgical diabetes following total pancreatectomy (TP), but outcomes are dependent upon islet mass, which is unknown prior to pancreatectomy. We evaluated whether preoperative metabolic testing could predict islet isolation outcomes and thus improve assessment of TPIAT candidates. We examined the relationship between measures from frequent sample IV glucose tolerance tests (FSIVGTT) and mixed meal tolerance tests (MMTT) and islet mass in 60 adult patients, with multivariate logistic regression modeling to identify predictors of islet mass ≥2500 IEQ/kg. The acute C-peptide response to glucose (ACRglu) and disposition index from FSIVGTT correlated modestly with the islet equivalents per kilogram body weight (IEQ/kg). Fasting and MMTT glucose levels and HbA1c correlated inversely with IEQ/kg (r values -0.33 to -0.40, p ≤ 0.05). In multivariate logistic regression modeling, normal fasting glucose (<100 mg/dL) and stimulated C-peptide on MMTT ≥4 ng/mL were associated with greater odds of receiving an islet mass ≥2500 IEQ/kg (OR 0.93 for fasting glucose, CI 0.87-1.0; OR 7.9 for C-peptide, CI 1.75-35.6). In conclusion, parameters obtained from FSIVGTT correlate modestly with islet isolation outcomes. Stimulated C-peptide ≥4 ng/mL on MMTT conveyed eight times the odds of receiving ≥2500 IEQ/kg, a threshold associated with reasonable metabolic control postoperatively.

Rapid determination of omeprazole in human plasma by protein precipitation and liquid chromatography-tandem mass spectrometry.

A rapid, sensitive and reliable method was developed to quantitate omeprazole in human plasma using liquid chromatography-tandem mass spectrometry. The assay is based on protein precipitation with acetonitrile and reversed-phase liquid chromatography performed on an octadecylsilica column (55 mm x 2mm, 3 microm particles), the mobile phase consisted of methanol-10 mM ammonium acetate (60:40, v/v). Omeprazole and flunitrazepam, the internal standard, elute at 0.80+/-0.10 min with a total run time 1.35 min. Quantification was through positive ion mode and selected reaction monitoring mode at m/z 346.1-->197.9 for omeprazole and m/z 314.0-->268.0 for flunitrazepam, respectively. The lower limit of quantitation was 1.2 ng/ml using 0.25 ml of plasma and linearity was observed from 1.2 to 1200 ng/ml. Within-day and between-day precision expressed by relative standard deviation was less than 5% and inaccuracy did not exceed 12%. The assay was applied to the analysis of samples from a pharmacokinetic study.

Development and validation of a liquid chromatography-tandem mass spectrometry method for the determination of phenylephrine in human plasma and its application to a pharmacokinetic study.

A sensitive and reliable method was developed to quantitate phenylephrine in human plasma using liquid chromatography-electrospray tandem mass spectrometry. The assay was based on solid-phase extraction with C18 cartridges and hydrophilic interaction chromatography performed on a pentafluorophenylpropylsilica column (50 mm x 4 mm, 3 microm particles), the mobile phase consisted of methanol-10 mM ammonium acetate (90:10, v/v). Quantification was through positive-ion mode and selected reaction monitoring at m/z 168.1-->135.0 for phenylephrine and m/z 182.1-->135.0 for internal standard etilefrin, respectively. The lower limit of quantitation was 51 pg/ml using 0.25 ml of plasma and linearity was observed from 51 to 5500 pg/ml. Within-day and between-day precision expressed by relative standard deviation was less than 12% and inaccuracy did not exceed 8% at all levels. The assay was applied to the analysis of samples from a pharmacokinetic study.

Rapid determination of valsartan in human plasma by protein precipitation and high-performance liquid chromatography.

A high-performance liquid chromatographic (HPLC) method for the determination of valsartan in human plasma is reported. The assay is based on protein precipitation with methanol and reversed-phase chromatography with fluorimetric detection. The preparation of a batch of 24 samples takes 20 min. The liquid chromatography was performed on an octadecylsilica column (50 mm x 4 mm, 5 microm particles), the mobile phase consisted of acetonitrile -15 mM dihydrogenpotassium phosphate, pH 2.0 (45:55, v/v). The run time was 2.8 min. The fluorimetric detector was operated at 234/374 nm (excitation/emission wavelength). The limit of quantitation was 98 ng/ml using 0.2 ml of plasma. Within-day and between-day precision expressed by relative standard deviation was less than 5% and inaccuracy did not exceed 8%. The assay was applied to the analysis of samples from a pharmacokinetic study.

Rapid determination of tamsulosin in human plasma by high-performance liquid chromatography using extraction with butyl acetate.

A high-performance liquid chromatographic method with fluorescence detection for the determination of tamsulosin in human plasma is reported. The sample preparation involved liquid-liquid extraction of tamsulosin from alkalised plasma with butyl acetate and back-extraction of the drug to the phosphate buffer (pH 2). Butyl acetate is preferable to more commonly used ethyl acetate because of its much lower solubility in water. Liquid chromatography was performed on an octadecylsilica column (55 mm x 4 mm, 3 microm particles), the mobile phase consisted of acetonitrile-30 mM dihydrogenpotassium phosphate (25:75 v/v). The run time was 3.5 min. The fluorimetric detector was operated at 228/326 nm (excitation/emission wavelength). An analogue of tamsulosin, (R)-5-[2-[(3-(2-ethoxyphenoxy)propyl)amino]-2-methylethyl]-2-methoxybenzensulfonamide was used as the internal standard. The limit of quantitation was 0.4 ng/ml using 1 ml of plasma. Within-day and between-day precision expressed by relative standard deviation was less than 10% and inaccuracy did not exceed 5%. The assay was applied to the analysis of samples from several pharmacokinetic studies.

Liquid chromatographic determination of carvedilol in human plasma.

A high-performance liquid chromatographic method for the quantitation of carvedilol in human plasma is presented. The method is based on protein precipitation with methanol, concentration of the supernatant by evaporation and reversed-phase chromatography with fluorimetric detection. The separation was performed on a Develosil 3 micro m ODS 100 x 4.6 mm I.D. column and the mobile phase consisted of acetonitrile-30 mM potassium dihydrogenphosphate buffer, pH 2 (30:70 v/v). With only 250 micro l of plasma used for sample preparation, the limit of quantitation 1.3 ng/ml was achieved. Dihydroergocristine mesylate was used as the internal standard. The between-day precision expressed by relative standard deviation was less than 6% and inaccuracy does not exceed 3%. The assay was used for pharmacokinetic studies.

Determination of mirtazapine in human plasma by liquid chromatography.

A rapid high-performance liquid chromatographic method for the quantitation of mirtazapine in human plasma is presented. The method is based on a liquid-liquid extraction and reversed-phase chromatography with fluorimetric detection. The separation was performed on a Luna microm C(18)(2) 50 x 4.6 mm I.D. column using an isocratic elution. Zolpidem hemitartrate was used as the internal standard. The between-day precision expressed by relative standard deviation was less than 5% and inaccuracy does not exceed 6%. A low limit of quantitation (1.5 ng/ml) and a short time of analysis (4 min) makes this assay suitable for pharmacokinetic studies.

Determination of alendronate in human urine as 9-fluorenylmethyl derivative by high-performance liquid chromatography.

A high-performance liquid chromatographic method for the quantitation of alendronate as the 9-fluorenylmethyl derivative (FMOC) in human urine is presented. The sample preparation involved coprecipitation with calcium phosphate, separation on diethylamine (DEA) solid-phase extraction (SPE) cartridge and derivatization with 9-fluorenylmethyl chloroformate in citrate buffer pH 11.9. Liquid chromatography was performed on an octadecylsilica column (150 x 4.6 mm, 3 microm particles); a gradient method with starting mobile phase acetonitrile-methanol-citrate/pyrophosphate buffer (20:15:65 v/v) was employed. The total run time was 21 min. The fluorimetric detector was operated at the following wavelengths: 260 nm (excitation) and 310 nm (emission). Pamdronate was used as the internal standard. The limit of quantitation was 3.5 ng/ml using 5 ml of urine. Within-day and between-day precision expressed by relative standard deviation was less than 8% and inaccuracy did not exceed 9%. The assay was applied to the analysis of samples from a pharmacokinetic study.

Auditory pattern recognition abilities of aphasic and normal subjects: a preliminary study.

A comparison of the ability of aphasic and normal subjects to recognize three element auditory patterns of duration was made under the following conditions: monaural right, monaural left; split left leading and split right leading: dichotic. There were eight possible combinations of elements for each condition. In all conditions the normal group clearly outperformed the aphasic subjects with practically no overlap in their performances. Implications of using nonlinguistic stimuli for testing aphasic individuals, assessing progress, and training in pattern recognition is discussed.

The fit between journals and theses.

The titles of a substantial sample of articles related to speech pathology and audiology over a period of 21 years were catalogued under one or more of 10 categories, e.g., normal audition, defective phonation, and the like. The titles of theses and dissertations in this field of six universities of Ohio over the same period were also catalogued. The articles appeared in 17 journals. The journals were treated as seven groups of "related" journals. The total output of journals was stable over the period studied; a "group" of journals tended to be consistent with itself over successive 3-year periods; the seven groups of journals tended to be unique, and not to replicate each other. The topics treated in the journals were accepted as a criterion for contemporaneousness. The student output of the universities varied in contemporaneousness from one school to another, ranging upward to r = 0.98 (10 categories). The overall correlation between the student and the journal outputs was r = 0.79.

Research trends in the field of speech and hearing.

Topics of research in the field of speech and hearing were identified and compared over a 21-yr period (1954--1974). These topics were identidied by a key-word analysis of approximately 8200 titles consisting of articles in national and international journals, and of theses and dissertations presented in the state of Ohio. Results of this analysis have pinpointed certain research trends in the field of speech and hearing. Attention to certain topics has either declined, increased, or reached a peak during the 21-yr period, while interest in some topics has been consistently maintained throughout the years. The information reported provides a perspective from which to view contributions made by researchers in the field of speech and during the last two decades.

[Attempt at a new look on the problems of traffic accidents type pedestrian--car]. / Pokus o nový pohled na problematiku dopravních nehod typu chodec--automobil.

In view of the complexity of investigation of the ever rising number of traffic accidents the authors draw attention to the need of a narrower specialization of experts in the branch of forensic medicine and on the desirability of close collaboration and technical specialist when elucidating mechanisms of traffic injuries. In one of the most pretentious sections, in traffic accidents of the type pedestrian--car, they want to attempt an interdisciplinary study the purpose of which is to obtain certain basic data for expert evaluation of the mechanism of fatal injuries of pedestrians, and a basis for assessing speed limits at sites of increased danger of this type of accidents. As source of information they will use in particullar: the protocol of the accident incl. the map and photodocumentation, a questionnaire prepared for the investigating authorities, the results of necropsy incl. laboratory examinations, and if available, conclusions of expert opinions from other special branches.

Determination of ofloxacin in human plasma using high-performance liquid chromatography and fluorescence detection.

A new method for the determination of ofloxacin in human plasma was developed. Plasma proteins were precipitated with acetonitrile, the supernatant concentrated and injected into a reversed-phase C18 column. Enoxacin was used as an internal standard. The fluorimetric detection was performed at 282 nm for excitation and 450 nm for emission. Limit of quantitation was 20 ng/ml and the calibration curve was linear up to 6900 ng/ml.

Rapid and simple method for the determination of zolpidem in human plasma by high-performance liquid chromatography.

A simple and reproducible method for the determination of zolpidem in human plasma is presented. This method involves protein precipitation with methanol (2 ml of methanol are added to 0.5 ml of plasma) and reversed-phase chromatography with fluorescence detection (excitation wavelength 244 nm, emission wavelength 388 nm). The mobile phase consists of methanol-30 mM dihydrogen potassium phosphate-triethylamine (30:69:1). pH of the aqueous part of the mobile phase is 6.8. No internal standard is required. Limit of quantitation is 1.5 ng/ml and the calibration curve is linear up to 400 ng/ml. Within-day and between-day precision expressed by relative standard deviation is less than 5% and inaccuracy also does not exceed 9%. The assay is useful for pharmacokinetic studies.

Determination of omeprazole in human plasma by high-performance liquid chromatography.

A new method for the determination of omeprazole in human plasma was developed. Omeprazole was extracted from plasma with toluene-isoamylalcohol (95:5, v/v), the organic phase was evaporated, dissolved in the mobile phase and injected into a reversed-phase C18 column. Flunitrazepam was used as an internal standard. The mobile phase consisted of 47% methanol and 53% of 0.1 M dipotassium hydrogenphosphate, pH 7.8. The spectrophotometric detection was performed at 302 nm. Limit of quantitation was 9.7 ng/ml and the calibration curve was linear up to 1240 ng/ml.

Determination of finasteride in human plasma by liquid-liquid extraction and high-performance liquid chromatography.

A high-performance liquid chromatographic method for the quantitation of finasteride in human plasma is presented. The method is based on liquid-liquid extraction with hexane-isoamylalcohol (98:2, v/v) and reversed-phase chromatography with spectrophotometric detection at 210 nm. The mobile phase consists of acetonitrile-15 mM potassium dihydrogenphosphate (40:60, v/v). Clobazam is used as the internal standard. The limit of quantitation is 4 ng/ml and the calibration curve is linear up to 300 ng/ml. Within-day and between-day precision expressed by relative standard deviation is less than 5% and inaccuracy does not exceed 8%. The assay was used for pharmacokinetic studies.

Determination of roxithromycin in human plasma by high-performance liquid chromatography with spectrophotometric detection.

A simple and reproducible method for the determination of roxithromycin in human plasma is presented. This method is based on liquid-liquid extraction with hexane-isoamylalcohol (98:2, v:v) and reversed-phase chromatography with spectrophotometric detection at 220 nm. The mobile phase consists of methanol-15 mM dihydrogen potassium phosphate (70:30, v:v), pH of the aqueous part of the mobile phase is 6.0. The column is operated at 60 degrees C. Clarithromycin is used as the internal standard. The limit of quantitation is 0.5 microg/ml and the calibration curve is linear up to 30 microg/ml. Within-day and between-day precision expressed by relative standard deviation is less than 5% and inaccuracy does not exceed 9%. The assay was used for pharmacokinetic studies.

Determination of cetirizine in human plasma by high-performance liquid chromatography.

A high-performance liquid chromatographic method for the quantitation of cetirizine in human plasma is presented. The method is based on liquid-liquid extraction with dichloromethane and reversed-phase chromatography with spectrophotometric detection at 232 nm. Gradient elution was used to remove late eluting peaks. Diazepam was used as the internal standard. The limit of quantitation was 10 ng/ml using 0.5 ml of plasma. Within-day and between-day precision expressed by relative standard deviation was less than 10% and inaccuracy did not exceed 8%. The assay was applied to the analysis of samples from a pharmacokinetic study.