Relationship between normal oral glucose tolerance test in women at risk for gestational diabetes and large for gestational age infants.

OBJECTIVE: To determine whether the glycemic status of pregnant women with a normal 3-h 100-g oral glucose tolerance test (OGTT) is related to outcome with respect to large for gestational age (LGA) infants. RESEARCH DESIGN AND METHODS: A prospective study of 2631 women was conducted. One hundred seventy-six women had an OGTT based either on a 1-h 50-g OGTT (n = 105) or clinical risk factors (n = 71). Thirty-three women were diagnosed as having gestational diabetes mellitus. RESULTS: Negligible discriminatory capacity for the variables with respect to prediction of LGA infants was indicated by the areas under the receiver operating characteristic (ROC) curves for fasting blood glucose, 2-h OGTT blood glucose, and the OGTT response curve area for women with a normal OGTT (n = 143). However, a statistically significant increased incidence of LGA infants was established for both the OGTT-positive and normal OGTT groups (P less than 0.0001). Multiparity, a maternal weight greater than or equal to 70 kg, and birth of a male infant were other factors associated with a significantly increased frequency of LGA infants. CONCLUSIONS: The results may be interpreted as either indicating a role for confounding variables, i.e., maternal weight, multiparity, and birth of a male infant, or the imprecision of the OGTT in assessing physiologically important changes in maternal hyperglycemia.

Variability of urinary albumin excretion in patients with microalbuminuria.

OBJECTIVE: To estimate the within-person variability (SDi) for the overnight urinary albumin excretion rate (AER) in diabetic patients with persistent microalbuminuria. RESEARCH DESIGN AND METHODS: Thirteen normotensive diabetic patients in stable medical control, with normal renal function and without any associated cardiovascular or other clinical disorders, collected overnight urines at monthly intervals during one year. AER was determined by radioimmunoassay. RESULTS: Analysis of individual series of AER showed a significant trend (P < or = 0.01) in three cases. The remaining patients (eight men, two women; seven with insulin-dependent diabetes mellitus) had a median mean AER 102 micrograms/min (range 30-238 micrograms/min). Because the individual mean AERs were significantly associated with their respective SDis, the data was loge transformed. LogeSDi was estimated as 0.420, and its 90% probability range (0.353-0.490) was calculated using the bootstrap method. CONCLUSIONS: The high within-person variance for AER means that only people with an initial AER in the range of 53-76 micrograms/min have a high probability (P > or = 0.95) of being classified as microalbuminuric (20-200 micrograms/min) on a subsequent specimen. However, subjects with an initial AER > or = 53 or > or = 80 micrograms/min have a 95 and 99% probability of persistent microalbuminuria. The large variability of AER limits its potential as a serial marker to detect any gradual deterioration of established renal dysfunction.

Evidence for a hypothalamic site of action of clomiphene citrate in women.

To examine the site of action of clomiphene citrate (CC), LH and FSH pulsatile amplitude, frequency, and responsiveness to GnRH (10 micrograms, iv) were studied in 11 women during the early follicular phase of the menstrual cycle. Six women received CC (150 mg/day) on cycle days 2, 3, and 4, while 5 women received placebo tablets. Blood samples were drawn at 10-min intervals for 8 h before and after the treatment regimen on cycle days 2 and 5, respectively. All women treated with CC had multiple follicular development, as determined by ultrasound. Peripheral levels of estradiol did not change after CC treatment, while progesterone levels decreased slightly. Mean levels of LH increased from 7.5 +/- 0.9 (+/- SEM) to 10.7 +/- 1.4 mIU/ml (P less than 0.05), and FSH increased from 6.7 +/- 0.9 to 10.1 +/- 0.9 mIU/ml (P less than 0.01). After exposure to CC, the pulse frequency of LH during an 8-h period increased significantly (3.3 +/- 0.7 on day 2 vs. 6.8 +/- 0.8 on day 5; P less than 0.01), while the pulse frequency of FSH increased from 3.8 +/- 0.6 to 5 +/- 1.4, as determined by computer pulse analyses. The pulse amplitude of LH and FSH was not significantly altered. In the placebo-treated group, neither pulse amplitude nor pulse frequency changed significantly between cycle days 2 and 5. Pituitary sensitivity to exogenous GnRH did not change after CC treatment. Since the pulsatile frequency of LH is governed by hypothalamic influences, these findings provide compelling evidence for a hypothalamic site of action for CC, probably by inducing an increase in the frequency of GnRH secretion.

Applications of heavy isotope tracers to clinical studies: progesterone urinary production rate determination in the menstrual cycle and pregnancy.

A method for the in vivo determination of progesterone production rate using progesterone labeled with deuterium as the tracer and gas chromatography-mass spectrometry for assessing isotope dilution in a urinary metabolite is described. By measuring the ratio of labeled to unlabeled free progesterone in a 24-h urine sample, the problem of identifying a unique metabolite appears to have been avoided. Median progesterone production rates on day 7 of the follicular phase and day 7 of the luteal phase were 3.5 mumol/24 h and 72.6 mumol/24 h, respectively. Where repeated determinations were performed through the luteal phase in two nonconceptual cycles, the familiar midluteal plateau of progesterone production was noted. The MCR of progesterone calculated from luteal phase data was 2051 1/24 h (SD, 275 1/24 h). When determined in pregnancy, progesterone production varied from 250 mumol/24 h at 6 weeks in a singleton pregnancy, to 1565 mumol/24 h at 35 weeks in a triplet pregnancy. Repeated determinations before term in a twin pregnancy detected no evidence of decreasing progesterone production before labor. The value of urinary pregnanediol and plasma progesterone to indirectly monitor progesterone production were confirmed.

Urinary steroid metabolites and the overgrowth of lean and fat tissues in obese girls.

Studies were made of steroid metabolites excreted in the urine of 17 obese girls 11.4 to 16.8 yr and 17 normal girls 11 to 17 yr. Creatinine excretion (muscle mass), total body water (or deuterium space), lean body mass and body fat were determined in the obese girls. Extracellular volume (corrected bromide space) was also measured and by difference with body water, intracellular water or soft tissue cell mass was calculated. In normal girls 24-h creatinine excretion was determined, but body water was predicted from height and weight. It was found, as in previous studies, that the obese girls had excess muscle mass and soft tissue cell mass for height. The excess growth of muscle, lean tissue, and body length in obese girls correlated with increments in oxosteroid (17 ketosteroid) excretion. The overall weight increase correlated with increased excretion of corticosteroid metabolites--a finding of interest since a physiological Cushing's syndrome was postulated for fat girls many years ago. When the normal and obese girls were divided by age at 14 yr and the subgroups compared (normal obese) the younger girls showed differences with respect to height, weight, total body water, fat and percentage fat. Differences in steroid metabolites were not found. In older girls the same findings were made again, but here it was clear that the increments in body size, particularly muscle mass, correlated with augmented oxosteroid excretion. Evidence is cited that these findings are not just related to a larger steroid pool in obese girls.

Evaluation of a commercial radioassay for the simultaneous estimation of vitamin B12 and folate, with subsequent derivation of the normal reference range.

A commercial assay kit method for the simultaneous estimation of vitamin B12 and folate concentrations has been evaluated. Values derived for folate by a microbiological assay and vitamin B12 by a verified radioassay showed good correlation with the investigated method. The clinical sensitivity of the assay for detecting deficient concentrations of vitamin B12 and folate was comparable to that of the non-commercial methods and other more definitive clinical procedures. Establishment of reference ranges, based on accepted statistical criteria, are discussed and such ranges are contrasted with those proposed by the manufacturer. The kit method is time and labour saving compared with the non-commercial methods.

The quantitation of plasma phytanic acid by mass fragmentography.

Plasma phytanic acid has been quantitated by mass fragmentography after its successive reduction and derivatization to the corresponding t-butyldimethylsilyl ether. The latter was estimated by selective ion monitoring of its characteristic (M-57) ion (M/z 355). The technique has the advantage of being more rapid, specific and sensitive than existing methods, permitting the determination of phytanic acid at levels greater than 5 mug/ml plasma.

Specific quantitation of plasma medroxyprogesterone acetate by gas chromatography/mass spectrometry.

Investigation of various derivatives (O-methyloxime, trifluoroenol acetate and t-butyldimethylsilyl enol ether) coupled with the efficient preparation of a trideuterated analogue of medroxyprogesterone acetate, has allowed the development of a rapid and accurate assay for its quantitation in plasma by isotope dilution gas chromatography/mass spectrometry. High oral doses (2.8 g weekly) of medroxyprogesterone acetate are shown to lead consistently to plasma levels of less than 16 ng.ml-1.

PIP: An assay, developed to quantitate plasma medroxyprogesterone acetate by isotope dilution gas chromatography/mass spectrometry, is reported. The assay couples derivatives of perfluoroacyl and t-butyldimethylsilyl (BDMS) to an efficient preparation of a trideuterated analog of medroxyprogesterone acetate. The method relies on a very weak, acidic catalyst, the use of which is outlined in the experimental section of the paper. With this assay plasma levels of medroxyprogesterone acetate were quantitated after administration of various doses via various routes. Doses of 2.8 gm/week, it was found, consistently produce, regardless of administration route, plasma levels of 16 ng/ml 1.

Preparation of androsta - 2,4-dien-17beta -ol.

A simple preparation of androsta-2,4-dien-17beta-ol from testosteroneis described...

Measurement of plasma progesterone and its O-methyloxime by gas chromatography-nitrogen detection.

A routine method for the quantitation of plasma progesterone, using only 1 ml of sample, has been developed based on gas chromatography-nitrogen detection of the O-methyloxime derivative. Simple solvent extraction of progesterone from plasma using cyclohexane as solvent and medrogestone as internal standard allows measurement at the 3 nmol/l level, while the coefficient of variation at 16, 64, and 127 nmol/l progesterone is 14, 7.5, and 6.5% respectively. Comparison with a radioimmunoassay gave a linear regression equation of Y = 0.84x + 1.4, Syx = 7.8, r = 0.9255 (n = 55).

Capillary gas chromatographic measurement of neutral urinary steroids: evaluation and comparison with alternative methodology.

The major neutral urinary steroids have been quantitated by capillary gas chromatography and the reproducibility and accuracy of the method have been fully evaluated. Typical intra- and inter-assay coefficients of variation for individual steroids ranged from 4.9 to 10.1% and from 15.6 to 21.5%, respectively. Comparison of steroid values with alternate and commonly employed procedures for measuring urinary 17-oxosteroids, total 17-oxogenic steroids, and cortisol and plasma dehydroepiandrosterone-sulphate yielded correlation coefficients between 0.71 and 0.79. A definite relationship between serum cortisol and urinary trihydroxy-pregnanedione was also shown. Urinary androstenedione metabolites, however, could not be significantly (P greater than 0.05) correlated with the normalised androgen ratio in plasma.

Subnanogram detection of t-butyldimethylsilyl fatty acid esters by mass fragmentography.

The mass spectra of t-butyldimethylsilyl fatty acid esters all display a pronounced (M-C4H9)+ ion. The proportion of the total ionization carried by this fragment, particularly for saturated and mono-, di-, and tri-unsaturated acid derivatives, facilitates their qualitative analysis at the subanogram level by mass fragmentography.

Derivatisation of steroid hormones with t-butyldimethylsilyl imidazole.

An efficient procedure for the derivatisation of hydroxyl and alpha,beta-unsaturated ketonic steroids using t-butyldimethylsilyl imidazole is described. The conditions which employ potassium acetate as a basic catalyst result in quantitative formation of BDMS ethers and greater than 96% yield of BDMS-enol ethers. Various mechanisms involving the catalysed and uncatalysed reactions of t-butyldimethylsilyl imidazole with alcohols and enols are also discussed. The mass spectra of the BDMS-enol ethers, in contrast to the BDMS ethers, are dominated by an intense molecular ion, which facilitates the quantitation of these derivatives in biological samples. An example of the method is shown by the determination of 4-androstene-3,17-dione in urine using a stable-isotope internal standard.

Evaluation of allyldimethylsilyl ethers as steroid derivatives for gas chromatography-mass spectrometry.

The preparation of allyldimethysilyl steroid ethers for the analysis of steroids by GC-MS has been studied in depth. The application of these derivatives is shown to be unsatisfactory due to competitive formation of allylsiloxane derivatives.