Timothy L. Bennett

Simultaneous measurement of adenosine and hypoxanthine in human umbilical cord plasma using reversed-phase high-performance liquid chromatography with photodiode-array detection and on-line validation of peak purity

A new, robust and sensitive reversed-phase high-performance liquid chromatographic method was developed for concomitant measurement of plasma concentrations of the ATP catabolites adenosine and hypoxanthine in human umbilical cord blood. Deproteinized cord plasma was chromatographed on Hypersil C18 columns, using UV photodiode-array detection, spectral analysis of peaks and on-line confirmation of peak purity. Elution with a gradient of acetonitrile-tetrahydrofuran in ammonium dihydrogen phosphate buffer pH 4.7, yielded sharp, well-resolved peaks of adenosine and hypoxanthine within 16 min. Peak areas were quantified from external calibration curves and converted to plasma concentrations via cord blood hematocrits. In seven deliveries, gestational ages 32-40 weeks, adenosine (range, 0.1-2.1 microM) was less than hypoxanthine (range, 1.6-18.5 microM) in the same cord plasma sample. Arteriovenous levels of each purine were similar, except in an abruptio placenta delivery.

Plasma endorphin-like immunoreactivity in pelvic and abdominal pain

&NA; This study was undertaken to determine the association, if any, of plasma endorphins with pelvic pain in a defined population. The study group was comprised of 37 patients who presented to the emergency room at Wishard Memorial Hospital for evaluation of gynecologic complaints. These subjects were divided into 3 groups depending on the degree of pelvic pain assessed both by the subject and the examining physician. Group I (control) consisted of those subjects who were pain‐free (9 subjects). Group II contained those whose pelvic pain was not felt to require hospital admission (18 subjects). Group III subjects represented those with pelvic pain whose diagnosis warranted hospitalization (10 subjects). Plasma endorphin levels were determined on each subject before and after pelvic examination using standard radioimmunoassay techniques. Data analysis was accomplished by analysis of variance methods. The timing of the pelvic examination made no difference on the plasma endorphin levels. Levels between groups were statistically similar, though differences between both group I and group II, and that of group III levels approached significance.

Measurement of hypoxanthine and xanthine in late-gestation human amniotic fluid by reversed-phase high-performance liquid chromatography with photodiode-array detection

A robust analytical method was developed for measurement of hypoxanthine and xanthine in late-gestation human amniotic fluid by reversed-phase high-performance liquid chromatography using diodearray detection. Purity of analyte peaks was confirmed via on-line analysis of peak spectra utilizing the purity parameter treatment of spectral data. Amniotic fluid obtained by amniocentesis was deproteinized by centrifugal ultrafiltration and chromatographed on an octadecylsilica column using isocratic elution with 1% (v/v) acetonitrile in 0.05 M ammonium dihydrogenphosphate pH 6.0; hypoxanthine and xanthine were resolved, but the hypoxanthine peak was not pure. Chromatography on a column of polar endcapped octadecylsilica, using similar mobile phase conditions, yielded spectrally pure peaks of hypoxanthine xanthine. Hypoxanthine and xanthine levels in amniotic fluid from fourteen patients, gestational age 34-39 weeks, ranged from 0.56 to 2.74 microM and 1.62 to 5.52 microM, respectively.

Atrial natriuretic factor responses to volume expansion in pregnant and nonpregnant sheep

Isotonic volume expansion results in atrial natriuretic factor release by cardiac myocytes. Because pregnancy produces well-established alterations in fluid homeostasis and cardiovascular function, changes in atrial natriuretic factor responses may also occur. This study compares plasma atrial natriuretic factor responses to short-term volume expansion in pregnant and nonpregnant sheep. Seven pregnant and six nonpregnant ewes were chronically instrumented and subjected to a series of four experiments consisting of a control group (no infusion) and groups that received 10 ml/kg, 25 ml/kg, and 40 ml/kg isotonic saline infusion over a 30-minute period. The order of the experiments was random and separated by greater than or equal to 48 hours. Plasma atrial natriuretic factor, osmolality, right atrial pressure, blood pressure, and urine flow were measured over a 150-minute observation period. After volume expansion, plasma atrial natriuretic factor levels rose significantly from 39 +/- 4 pg/ml (mean +/- SEM) to 49 +/- 7 pg/ml, 36 +/- 4 pg/ml to 62 +/- 19 pg/ml, and 39 +/- 6 pg/ml to 67 +/- 14 pg/ml in the nonpregnant group 10 ml/kg, 25 ml/kg, and 40 ml/kg experiments, respectively. In the pregnant groups, plasma atrial natriuretic factor levels rose from 50 +/- 2 pg/ml to 75 +/- 20 pg/ml, 43 +/- 5 pg/ml to 57 +/- 5 pg/ml, and 46 +/- 4 pg/ml to 67 +/- 7 pg/ml, respectively. Differences in atrial natriuretic factor responses were not seen between pregnant and nonpregnant groups at any volume expansion level. As expected, atrial pressure and urine flow significantly increased after all volume expansion experiments. Pregnant and nonpregnant groups were similar with respect to atrial pressure and urine flow responses. Over various volume expansion levels significant associations were seen between atrial pressure, atrial natriuretic factor, and urine flow. These relationships were unaltered by pregnancy. In summary, atrial natriuretic factor responses to volume expansion do not appear to differ between pregnant and nonpregnant sheep.