Metabolic and renal clearance rates of purified human chorionic gonadotropin.

The metabolic clearance rate (MCR) and renal clearance rate (RCR) of human chorionic gonadotropin (hCG) were measured in healthy young men and women using techniques of continuous intravenous infusion and rapid intravenous injection of unlabeled, highly purified hCG. Seven subjects received 4 d of infusion at a rate of 0.2 microgram/min, followed by an additional 4 d of infusion at 0.8 microgram/min. Mean serum levels of hCG established at these rates of infusion were 61.1 +/- 3.3 and 237 +/- 16 ng/ml, respectively (mean +/- SEM). The MCR determined at the low infusion rate was not significantly different from that determined at the higher infusion rate (1.83 +/- 0.09 vs. 1.95 +/- 0.14 ml/min per m2). The mean MCR for all subjects was 1.88 +/- 0.08 ml/min per m2. The MCR was not significantly different between men amd women (2.04 +/- 0.13 vs. 1.76 +/- 0.07 ml/min per m2). The RCR also did not vary between low and high infusion rates (0.40 +/- 0.03 vs. 0.40 +/- 0.04 ml/min per m2). The mean RCR for all subjects was 0.40 +/- 0.02 ml/min per m2. There was no difference in RCR between men and women (0.42 +/- 0.05 vs. 0.39 +/- 0.03 ml/min per m2). Six subjects were given 1.0 mg of highly purified hCG by rapid intravenous injection. Initial serum levels of hCG were 300-400 ng/ml, and the subsequent disappearance curve was multiexponential over 8-10 d. The disappearance curve of hCG in each subject was fitted to a biexponential equation. The rapid component t1/2 was 5.97 +/- 0.63 h and the slow component t1/2 was 35.6 +/- 8.0 h. We conclude that the MCR of purified hCG in man is about 2 ml/min per m2 and the RCR is 0.4 ml/min per m2; these parameters are concentration independent and do not differ significantly between healthy young men and women.

Plasma clearance rates and renal clearance of 3H-labeled cyclic AMP and 3H-labeled cyclic GMP in the dog.

Previously, in an attempt to understand the mechanisms involved in the regulation of plasma cyclic nucleotides, we measured concentrations of adenosine 3',5'-monophosphate (cAMP) and guanosine 3',5'-monophosphate (cGMP) in plasma from selected blood vessels of anesthetized dogs. The observation that the renal venous plasma concentrations of both cyclic nucleotides were less than arterial concentrations suggested that the kidney might be an important site for the elimination of these compounds from plasma and prompted further investigation of the renal handling of these compounds. Tracer doses of either [(3)H]cAMP or [(3)H]cGMP were administered to anesthetized dogs by constant intravenous infusion, and metabolic clearance rates were determined. Concentrations of endogenous cyclic nucleotide and of cyclic nucleotide radioactivity were measured in aortic and renal venous plasma as well as in urine. Renal venous plasma [(3)H]cGMP was 39% and [(3)H]cAMP was 65% of the concentration in arterial plasma. Endogenous cyclic nucleotide levels showed a similar relationship. The plasma clearance rates (PCR) were 271+/-27 ml/min (mean+/-SE) for cGMP and 261+/-17 for cAMP. The total kidney clearance (calculated as the renal plasma flow x renal cyclic nucleotide extraction ratio) accounted for 52+/-4% and 30+/-2% of the PCR for cGMP and cAMP, respectively. Only about two-thirds of the total kidney clearance of each cyclic nucleotide could be accounted for by urinary excretion, the remainder presumably being the result of renal metabolism. The urinary clearances of (3)H-labeled cGMP (40.9+/-4.2 ml/min) and endogenous cGMP (45.0+/-2.3 ml/min) were not significantly different from each other. Both were approximately 50% greater than the glomerular filtration rate, which was 27.1+/-2.0 ml/min, indicating that a significant amount of urinary cGMP is derived from plasma by tubular secretion. In contrast, the urinary clearances of (3)H-labeled cAMP (23.7+/-1.9 ml/min) and endogenous cAMP (27.2+/-2.6 ml/min) were nearly equal both to each other and to the glomerular filtration rate, which was 24.6+/-1.7 ml/min. Thus, in the dog, glomerular filtration of plasma cAMP appears to be responsible for most of the cAMP found in urine. Renla production of cAMP, which in humans contributes from a third to a half of the urinary cAMP, was quantitatively of minor importance in the dog.Thus, under the conditions of these experiments in dogs, renal elimination appears to be responsible for half of the PCR of cGMP and about a third of the PCR of cAMP. About a third of the renal elimination of both cyclic nucleotides appears to be due to metabolic degradation within the kidney, and the balance is due to excretion in the urine.

Sources of cyclic nucleotides in plasma.

In order to determine the sites of net production and removal of the cyclic nucleotides in plasma, various blood vessels were catheterized in 17 anesthetized dogs and arterial and venous concentrations of adenosine 3',5'-monophosphate (cAMP) and guanosine 3',5'-monophosphate (cGMP) were measured by radioimmunoassay. Aortic cAMP was 30+/-2 nM (mean+/-SE) and cGMP was 13+/-1 nM. There were no significant differences for either cyclic nucleotide between the concentration in the aorta and that in the inferior vena cava, coronary sinus, hepatic vein, and femoral vein. The concentration of cAMP in renal venous plasma was 25% lower than in aortic plasma, and renal venous cGMP was 51% lower than in the aorta. The pulmonary arterial concentrations of cAMP and cGMP were slightly lower than in the aorta. The concentration of cGMP in the superior mesenteric vein plasma was 83% greater than in aortic plasma; the concentration of cAMP in this vessel was only 16% greater than that in the aorta. Superior vena cava concentrations of both cyclic nucleotides were slightly greater than arterial concentrations. THE RESULTS SUGGEST THAT: (a) the kidneys are a major site of removal of both cyclic nucleotides from plasma. (b) The lungs may be a site of net addition of both cyclic nucleotides to plasma. (c) The small intestine is a site of net production of both cyclic nucleotides, particularly cGMP. (d) The liver probably removes cyclic nucleotides from plasma. (e) Since no other organs or regions studied added detectable net amounts of cyclic nucleotides to plasma, and since the turnover of these compounds in plasma is known to be rapid, the production of plasma cyclic nucleotides under basal conditions may well be the result of small net contributions may well be the result of small net contributions from many tissues or bidirectional fluxes between tissues and plasma, or both.

Characterization of a carboxyterminal peptide fragment of the human choriogonadotropin beta-subunit excreted in the urine of a woman with choriocarcinoma.

We have observed low-molecular weight carboxyterminal fragments of the human choriogonadotropin (hCG) beta-subunit in the urines of several women with choriocarcinoma, and we have characterized one fragment in detail. Its apparent molecular weight by gel chromatography on Sephadex G-100 was 14,200. The fragment was not adsorbed to concanavalin A-Sepharose, indicating that it lacked the asparagine-linked carbohydrate groups of intact hCG beta. It was active in radioimmunoassays (RIA) using antisera either to the hCG beta carboxyterminal peptide (CTP) or to the desialylated hCG beta CTP (hCG beta as-CTP), indicating the presence of not only the hCG beta carboxyterminus but also desialylated O-serine-linked carbohydrate side chains on the fragment. It lacked luteinizing hormone/choriogonadotropin radioreceptor activity and hCG beta conformational immunoreactivity (SB6 RIA). On Sephadex G-100 gel chromatography, the elution profiles of this fragment and the hCG beta as-CTP(115-145) prepared by trypsin digestion of as-hCG were essentially indistinguishable (apparent molecular weights 14,200 and 14,000, respectively). The immunological characteristics of the fragment in both hCG beta CTP and hCG beta as-CTP RIA were indistinguishable from those of the hCG beta as-CTP(115-145) glycopeptide. Carboxyterminal fragments of hCG beta were evident in urine specimens obtained from 10 of 11 patients with choriocarcinoma but not in those obtained from normal subjects who were given an intravenous infusion of highly purified hCG. Of six pregnant women, only the one at term excreted carboxyterminal fragments of hCG beta and then only in trace amounts. We conclude that hCG beta carboxyterminal fragments, including one that is indistinguishable from the tryptic glycopeptide hCG beta as-CTP(115-145), can occur naturally in the urine of patients with choriocarcinoma.

beta-Core chemical and clinical properties.

beta-Core is the most abundant hCG-related molecule in pregnancy urine. The structure of beta-core as well as aspects of its metabolic clearance suggest that beta-core is a metabolic fragment of the hCG-beta subunit. The occurrence of beta-core in the urine of patients with a broad spectrum of malignancies imparts an important role to beta-core as a tumor marker. The recent development of antisera with enhanced specificity and sensitivity for beta-core will facilitate further studies on the clinical significance of this molecule.

Estimation of a physiologic replacement dose of levothyroxine in elderly patients with hypothyroidism.

We gave graded doses of levothyroxine sodium to 11 elderly hypothyroid subjects (mean age, 66.1 years). The daily levothyroxine sodium dose was initially 75 micrograms or less, and was increased by 25 micrograms every six weeks. Serum total thyroxine, total triiodothyronine, and basal thyrotropin levels were measured at the start of the study and at the end of each six-week dose period. A protirelin (thyrotropin-releasing hormone) test was performed when the thyrotropin level returned to normal. Mean daily levothyroxine sodium doses that normalized serum thyrotropin levels and protirelin test were 110 +/- 8 micrograms and 113 +/- 9 micrograms, respectively. Serial basal thyrotropin determinations during stepwise increments in levothyroxine dose indicated physiologic hormone replacement. As determined in our elderly patients, levothyroxine replacement dose was a third less than that formerly recommended.

Carbohydrate composition of beta-core.

beta-Core is a major component of the hCG-related molecules found in pregnancy urine. We previously have purified the beta-core molecule and have deduced portions of its carbohydrate structure based on lectin binding data. In the present study we used recently developed technology to determine the carbohydrate composition of beta-core and hCG beta (CR119). For direct compositional analysis, parallel samples were hydrolyzed in trifluoroacetic acid and analyzed for sialic acid and neutral sugars without prior derivatization. Separation of the monosaccharides was achieved by HPLC on a Dionex CarboPac column eluted at high pH, and the resolved monosaccharides were quantified by pulsed amperometric detection. The amounts of sugar that were found relative to peptide indicated the presence of two N-linked oligosaccharides per molecule on both beta-core and hCG beta. hCG beta contained additional sugars consistent with the presence of four O-linked oligosaccharides. Compared to hCG beta, beta-core contained negligible sialic acid, galactose, or N-acetylgalactosamine. The compositional data suggest that beta-core does not contain N-acetylglucosamine at the nonreducing end of the molecule, whereas the trimannosyl-chitobiose core is apparently intact at both glycosylation sites, consistent with the ability of the molecule to bind to Concanavalin-A. Comparable fucose contents and abilities of beta-core and hCG beta to bind to Lens culinaris indicate a similar extent of fucosylation on the internal N-acetylglucosamine in both molecules. We propose that the N-linked oligosaccharides on beta-core closely resemble the underlying N-linked structures of hCG beta with the antennary sialic acid, galactose, and N-acetylglucosamine removed.

Purification of beta-core fragment from pregnancy urine and demonstration that its carbohydrate moieties differ from those of native human chorionic gonadotropin-beta.

Pregnancy urine contains large quantities of hCG, free beta-subunit, free alpha-subunit, and a population of fragments of beta-subunit known as beta-core. This beta-core population, which can account for as much as 70% of the total beta-immunoreactivity in pregnancy urine, is of interest as both a normal metabolite of pregnancy and a potential marker for malignancy. We have purified the beta-core fragment from pregnancy urine (P-core) and have characterized it with respect to size and carbohydrate composition. P-Core was purified by chromatography on Sephadex G-100, Concanavalin A (Con A)-Sepharose, DEAE-Sephacel, and Sephadex G-75 (superfine). The purified P-core has an apparent mol wt of 17,500 and 17,000, as determined by gel filtration on Sephadex G-75 (superfine) and sodium dodecyl sulfate-polyacrylamide gel electrophoresis under nonreducing conditions, respectively. The sialic acid content of P-core was assayed chemically and was less than 0.07 mumol sialic acid/mumole P-core. For comparison to P-core, we have prepared a trypsin fragment of beta-subunit that retains the beta-core immunological determinant recognized by SB6 antiserum and lacks the carboxy-terminal immunological determinant. We have designated this beta-core molecule as T-core (tryptic fragment of beta-subunit) to distinguish it from the beta-core molecule that we have purified from pregnancy urine (i.e. P-core). Most of the P-core and T-core molecules bind to Con A (84% and 86%, respectively). The Con A-bound material was used for subsequent characterizations. Neither P-core nor T-core binds to DEAE using conditions under which intact hCG beta binds to DEAE. A variety of agarose-bound lectins were used to further investigate the carbohydrate nature of the Con A-bound P-core and T-core molecules. The lectin binding data indicate that the antennae on P-core do not contain appreciable sialic acid or galactose, in contrast to the antennae on T-core, which contain both. We conclude that P-core, the naturally occurring beta-core fragment in pregnancy, has been processed to a form in which nearly all of the sialic acid and galactose residues are removed, but the Con A-binding site (consisting of the core sugars) and most of the core fucose are retained.

Stimulation of testosterone production in the cynomolgus monkey in vivo by deglycosylated and desialylated human choriogonadotropin.

Modifications of carbohydrate structures of hCG, such as deglycosylation or desialylation, have been shown to reduce the biological activity of the hormone derivatives in vivo. We posed the question of whether deglycosylated hCG (dg-hCG) and desialylated hCG (ds-hCG) would behave as agonists at the LH/CG receptor in the primate in vivo, as this would bear on their potential clinical utility as LH/CG agonists or antagonists. Thus, we administered large doses (approximately 3 nmol) of highly purified dg-hCG, ds-hCG, hCG, or normal saline as a rapid iv injection to adult male cynomolgus monkeys (n = 3/group). Mean areas under the curves of plasma T over the first 6 h achieved with dg-hCG and ds-hCG were about 5-fold, significantly (P less than 0.05) greater than that in the saline controls and not significantly (P greater than 0.05) different from that in hCG-injected animals. Despite comparable plasma T responses in the first 6 h, mean plasma concentrations of ds-hCG, dg-hCG, and hCG differed dramatically among the groups. Plasma ds-hCG and dg-hCG levels were undetectable by 15 and 180 min, respectively, while the mean plasma hCG level was more than 2.10 nmol/L at 360 min. These data indicate that 1) dg-hCG is a full agonist at the LH/CG receptor in the primate in vivo, despite having minimal intrinsic activity in the rat Leydig cell adenyl cyclase assay and being able to near-completely antagonize hCG action therein; and 2) ds-hCG is a full agonist in the monkey in vivo, capable of stimulating a full testicular response over 6 h, despite being cleared from the circulation in 15 min. We conclude that the signal transduction system at the monkey LH/CG receptor is capable of achieving full steroidogenesis despite dramatically shortened exposure to stimulus or exposure to a stimulus with markedly reduced adenyl cyclase-stimulating activity in vitro.

Metabolic clearance rates of the subunits of human chorionic gonadotropin in man.

Highly purified preparations of the alpha- and beta-subunits of hCG (hCG alpha and hCG beta) were injected iv in normal subjects. After rapid injection, the disappearance of both subunits from serum was nonlinear when plotted on a semilog graph. A two-component exponential curve was fitted for each subject, and the curve parameters were used to estimate the MCR, apparent initial volume of distribution (Vd), and half-times of disappearance of the rapid and slow phase for each subunit. The Vd of hCG beta was indistinguishable from that of hCG alpha (1958 +/- 131 vs. 1729 +/- 99 ml/m2, respectively). The rapid phase half-time for hCG beta was significantly longer than that of hCG alpha (41.2 +/- 1.7 vs. 13.0 +/- 0.9 min; P less than 0.001), and the slow phase half-time of hCG beta was also significantly longer than that of hCG alpha (236 +/- 41 vs. 76 +/- 19 min; P less than 0.01). The estimated MCR of hCG alpha was 49.7 +/- 1.6 ml/min.m2; this value was significantly greater than that of hCG beta (19.0 +/- 0.7 ml/min.m2; P less than 0.001). No significant differences between sexes in parameters determined for the subunits were observed. Continuous infusion of subunits at a rate of 2.7 microgram/min achieved a steady state blood level of hCG beta that was significantly greater than that of hCG alpha (59.1 +/- 7.8 vs. 24.4 +/- 0.7 ng/ml; P less than 0.02) and gave a MCR of hCG alpha that was 3 times greater than the MCR of hCG beta (72.2 +/- 4.9 vs. 21.6 +/- 2.8 ml/min.m2; P less than 0.001). We conclude that the hCG subunits have similar Vds, but since hCG alpha has much shorter half-times of disappearance in both rapid and slow phases, the MCR of hCG alpha is much greater than that of hCG beta.

Pituitary-thyroid hormone economy in healthy aging men: basal indices of thyroid function and thyrotropin responses to constant infusions of thyrotropin releasing hormone.

In order to assess the effects of aging, as distinct from those of thyroid disease or extrathyroidal illness, on certain indices of thyroid function, we studied 74 healthy, ambulatory men recruited from the Baltimore Longitudinal Study on Aging. We determined basal serum values of T4, T3, rT3, thyroxine-binding globulin (TBG), and T3 resin uptake (T3RU) and calculated the free T4 index (FT4I = T4 X T3RU/100), free T3 index (FT3I = T3 X T3RU/100), and T4/TBG ratio for each subject. We used an ultrasensitive RIA to measure variations in basal concentrations of TSH within the normal range. We then infused TRH at a constant rate (0.4 microgram/min iv) for 240 min into 63 of the same men; serum samples, collected at 15-min intervals during the infusion, were analyzed for TSH by routine RIA. Subjects were divided into 3 groups according to age; A (n = 26, mean age = 39.4), B (n = 23, mean age = 60.0), and C (n = 25, mean age = 79.6). Analysis of variance with Duncan's multiple range test and regression analysis were used to evaluate data. There was no significant (P greater than 0.05) variation with age of basal serum values of T4, TBG, or T3RU. Comparison of groups A and C showed significant decreases of mean values of serum T3 (-11%, P less than 0.05), FT3I (-13%, P = 0.02), FT4I (-11%, P less than 0.01), and T4/TBG ratio (-12%, P less than 0.01) and an increase in serum TSH (+38%, P less than 0.05). For these variables, the mean values for group B were intermediate between, but not significantly different from, those of A and C. Regression analysis showed significant correlations of age with T3, FT3I, FT4I, T4/TBG, and TSH at P levels similar to those obtained by Duncan's test. No elderly individual exhibited a baseline elevation of TSH (greater than 7 microU/ml) or depression of T4 (less than 5 micrograms/dl), suggesting that primary hypothyroidism was not present in our old group. The basal TSH concentration did not correlate significantly with any index of thyroid function except with FT3I in group C (r = -0.43, P less than 0.05). In all age groups the TSH responses to TRH exhibited a biphasic pattern with early and late peaks.(ABSTRACT TRUNCATED AT 400 WORDS)

Metabolic clearance rate and urinary clearance of purified beta-core.

We injected a highly purified preparation of the beta-core molecule, a fragment of hCG beta excreted in pregnancy urine, into five men and three women to determine its kinetic parameters, MCR, and urinary clearance. The beta-core molecule was distributed in an initial volume [1950 +/- 156 (mean +/- SEM) mL/m2 body surface area] approximately equal to the estimated plasma volume. Its disappearance was multiexponential on a semilogarithmic plot, with a rapid phase t1/2 of 3.5 +/- 0.7 min and a slow phase t1/2 of 22.4 +/- 4.2 min. The transit time (the mean time spent by a molecule of beta-core in transit) was 20.6 +/- 2.1 min. The MCR was 192.0 +/- 8.0 mL/min.m2 body surface area. About 5% of the injected dose of beta-core was excreted into the urine in the first 30 min after injection, and low levels of excretion persisted for up to 7 days. The urinary clearance rate of beta-core was 13.7 +/- 1.4 mL/min.m2, accounting for about 8% of the elimination of beta-core from the plasma. The beta-core immunoreactivity in serum and urine was characterized by gel filtration and three independent RIA systems to show that its properties were indistinguishable from those of the injected beta-core. Serum levels of beta-core in pregnant women were less than 0.2 ng/mL, while the amounts excreted in their urine were as much as 5 mg/day. Based on these clearance parameters of beta-core in normal subjects, less than 0.2% of the beta-core excreted in pregnancy urine is derived by urinary clearance of plasma beta-core. Therefore, more than 99% of the beta-core excreted in pregnancy urine is derived from beta-core in a compartment separate from plasma. In particular, these data indicate that there is relatively little placental secretion of beta-core into plasma and that placental secretion does not account for the vast majority of beta-core in pregnancy urine. These findings are consistent with previous data that point to renal parenchymal degradation of hCG and hCG beta as the major source of urinary beta-core in pregnancy.

Glycosylation changes in human chorionic gonadotropin and free alpha subunit as gestation progresses.

Carbohydrate is important to the structure, function, and circulatory survival of the glycoprotein hormones. Human CG (hCG) and the related free alpha-molecule of pregnancy contain four and two asparagine-linked oligosaccharides, respectively. The present study analyzes changes in the glycosylation patterns of hCG and free alpha in early vs. late gestation. Five volunteers provided 24-h urine samples, weekly, throughout their pregnancies. Extracts of early pregnancy (weeks 7-12) and late pregnancy (weeks 28-32) urines were pooled. Early and late samples from each patient were subjected to gel filtration to separate hCG and free alpha, and the populations thus obtained were analyzed by lectin affinity chromatography on Concanavalin A-Sepharose (Con A) and Lens culinaris-agarose (Lch). Using Con A, free alpha and hCG were separated into an unbound fraction (eluted with Con A buffer), a weakly bound fraction (eluted with 10 mmol alpha-methyl-D-glucoside) and a tightly bound fraction (eluted with 500 mmol alpha-methyl-D-mannoside). For free alpha-molecule, a significant decrease in tightly bound Con A forms, was noted from early to late pregnancy with a mean difference of 17.0 +/- 2.4% (P less than 0.01). Concomitantly, in late pregnancy, an increase in Con A unbound forms of free alpha was noted with mean difference of 12.5 +/- 1.7% (P less than 0.01). These changes indicate the presence of more highly branched oligosaccharides on free alpha as gestation advances. No changes were noted in the Con A binding of intact hCG; nearly all hCG bound in both early and late pregnancy. Using Lch, free alpha and hCG were separated into an unbound fraction (eluted with Lch buffer) and a bound fraction (eluted with 500 mmol alpha-methyl-D-mannose). Both free alpha and intact hCG in late pregnancy exhibited increased binding to Lch, with mean differences from early to late pregnancy of 30.2 +/- 4.8% (P less than 0.01) and 11.4 +/- 4.5% (P less than 0.05), respectively. These data indicate increased incorporation of fucose into the carbohydrate moieties in late pregnancy. Taken together, these data derived by analysis using lectin specificity imply the presence of more highly branched, fucosylated oligosaccharides as gestation progresses.

A radioimmunoassay for the core fragment of the human chorionic gonadotropin beta-subunit.

We developed a RIA for the beta-core fragment of hCG that is excreted in the urine of pregnant women and some patients with cancer. We purified beta-core from crude commercial preparations of hCG (of which beta-core is a major constituent) derived from pregnancy urine and used this purified beta-core material to immunize rabbits. One antiserum (RW25) was particularly useful in that a RIA with purified beta-core as both radioligand and reference preparation had high sensitivity for beta-core detection and low cross-reactivity with other hCG-related molecules and glycoprotein hormones. The cross-reactivities of purified hCG (CR125), hCG beta (CR125), and hCG alpha (CR125) preparations were in each instance less than 3 x 10(-3) (wt/wt). The cross-reactivities of purified pituitary glycoprotein hormones were in each instance less than 2 x 10(-4) (wt/wt). Using the RW25 RIA, virtually all beta-core immunoreactivity in pregnancy urine eluted from Sephadex G-100 in a position coincident with that of purified beta-core. Urine from men and nonpregnant women contained very low levels of beta-core immunoreactivity (less than 6.5 micrograms/L), while urine from pregnant women and patients with testicular cancer or other neoplasms had levels of beta-core immunoreactivity ranging as high as 26,000 micrograms/L. We conclude that the improved specificity of our beta-core RIA will facilitate studies of the physiology and cancer biology of beta-core molecules.

Disparity between beta-core levels in pregnancy urine and serum: implications for the origin of urinary beta-core.

We used a highly purified preparation of the naturally occurring core fragment of hCG beta (beta-core) and a new RIA for beta-core to investigate the concentrations and behavior of beta-core in serum and urine. We collected serum and 24-h urine specimens from healthy pregnant women during the first trimester of pregnancy. The concentrations of beta-core in serum were determined by analysis of fractions eluted from Sephadex G-100. Serum concentrations of beta-core immunoreactivity were very low (0.13-1.25 micrograms/L), while large amounts of beta-core were excreted in urine during pregnancy (as much as 4-5 mg/day). Interference with measurement by serum factors did not account for the low levels of beta-core immunoreactivity in pregnancy serum. Based on the known urinary clearance rate of beta-core in healthy nonpregnant subjects, we calculated that urinary clearance of serum beta-core accounts for only about 1% of the beta-core in pregnancy urine. We conclude that during pregnancy, the concentrations of beta-core in plasma are measurable, but extremely low, and that most of the beta-core in urine is derived by mechanisms other than urinary clearance of serum beta-core; most likely, these mechanisms involve nephrogenous production of beta-core from precursor molecules such as hCG and hCG beta.

Beta-core fragments are contaminants of the World Health Organization Reference Preparations of human choriogonadotrophin and its alpha-subunit.

Highly purified preparations of human choriogonadotrophin (hCG), hCG alpha and hCG beta, including those preparations which are being distributed by the World Health Organization as International Standards, cross-reacted in a new radioimmunoassay with increased relative specificity for the beta-core fragment of hCG. A major portion of the beta-core immunoreactivity in the hCG and hCG alpha preparations eluted from Sephadex G-100 in a position (approximate apparent molecular size 15,000-18,000) corresponding to that of purified beta-core fragment prepared from pregnancy urine. However, in the case of hCG beta-subunit preparations, virtually all of the beta-core cross-reacting material eluted from Sephadex G-100 in the same fractions as the native hCG beta-subunit. Quantitatively, the cross-reacting beta-core material accounts for less than 1% (w/w) of the total hCG or subunit immunoreactivity, as measured by conventional radioimmunoassays. The presence of the beta-core fragments as discrete molecular components of the hCG and hCG alpha preparations should be borne in mind when these preparations are used to calibrate new radioimmunoassays for hCG-related molecules.

Human chorionic gonadotrophin-like and beta-core-like materials in postmenopausal urine.

In addition to human chorionic gonadotrophin (hCG), the urine of pregnant women contains a small molecular weight form of the hCG-beta subunit known as beta-core. Human CG-like material has been described in tissues, serum and urine of normal man, particularly in postmenopausal women. We examined different urine preparations from postmenopausal women to determine whether beta-core-like material, as well as hCG-like material, could be detected. We studied an acetone extract of a pool of 11 litres of postmenopausal urine, three different commercial preparations of human menopausal gonadotrophins and two commercial preparations of 'pure' FSH. After Sephadex G-100 chromatography of these various postmenopausal urine extracts, fractions were assayed using four assay systems to detect hCG, beta-core, LH and FSH immunoreactivities. Human CG immunoreactivity was readily detected in all urinary extracts and it eluted in a position indistinguishable from that of purified hCG. In addition to this hCG-like material, all urinary extracts, except the commercial 'pure' FSH preparations, contained material which reacted in the beta-core radioimmunoassay. This beta-core immunoreactive material eluted from Sephadex G-100 in a position corresponding to that of purified pregnancy-derived beta-core. We conclude that the urine of postmenopausal women contains material resembling the beta-core molecule found in pregnancy urine. The origin of this beta-core-like material remains to be determined, and its presence will have an impact on the application of urinary beta-core as a tumour marker.

The use of biochemical assays in epidemiologic studies of reproduction.

Recent progress in the assay of urinary hormones has opened new opportunities for epidemiologists to study hormones and health outcomes. This is especially true for studies of female reproduction. The cyclic nature of female reproduction can be fully described only by continuous frequent measurements that, in order to be practical, require easily collected biological specimens. We describe our experience in collecting and analyzing daily urine specimens from 301 healthy women. We conclude that this approach is not only feasible but potentially of great value to epidemiologists for studying fertility, early pregnancy, the effects of toxic exposures on reproduction, and the relationships between reproduction and later risk of chronic diseases.

Measuring early pregnancy loss: laboratory and field methods.

We intensively studied 30 women attempting pregnancy in order to lay groundwork for larger studies of early pregnancy loss. These women collected first morning urine specimens for up to 6 months after discontinuing use of birth control. Urine specimens were successfully collected for 98% of the woman-days in the study. Three assays for human chorionic gonadotropin (hCG) were performed on each urine specimen. An immunoradiometric assay (IRMA) specific to the carboxyterminal peptide of the hCG beta-chain proved to be more sensitive and more specific than two radioimmunoassays (RIAs). Using the IRMA, we found four cases in which hCG rose and fell over successive days, consistent with early pregnancy loss. For three of these four cases, the level of hCG was too low to be detectable with the RIAs. Among the control group of five women with tubal ligations, there was no detectable hCG above threshold with the IRMA. Thus, the enhanced sensitivity and specificity of the IRMA allows very early pregnancy losses to be identified that would otherwise be undetectable. Furthermore, its effectiveness with small quantities of first morning urine makes the IRMA a useful tool for epidemiologic studies.

Serum testosterone response to desialylated human choriogonadotropin in man.

To assess the in vivo steroidogenic activity of desialylated human choriogonadotropin (hCG) in man, highly purified desialylated hCG was administered as a constant intravenous infusion over 6 hours to four normal men at a rate sufficient to maintain substantial levels of desialylated hCG in the serum. The mean percent change of serum testosterone from baseline during the first 6 hours in men given an infusion of desialylated hCG was compared to that in saline-infused controls and that in men given highly purified intact hCG. The mean change of serum testosterone at 6 hours in the group infused with desialylated hCG (129% of baseline) was significantly greater than that of the controls (69% of baseline). Furthermore, the response to desialylated hCG could not be distinguished from that of hCG (125% of baseline). It was concluded that desialylated hCG, when given as a constant intravenous infusion, can elicit a substantial serum testosterone response comparable to that seen with purified hCG, and thus, that desialylated hCG behaves as an agonist of the LH/CG receptors on human Leydig cells.