Radioimmunoassays for spirapril and its active metabolite spiraprilate: performance and application.

Radioimmunoassays for a nonsulfhydryl angiotensin converting enzyme inhibitor prodrug--spirapril--and its active metabolite--spiraprilate--are described. Nonextraction equilibrium assays using antibodies with a high specificity for spirapril or spiraprilate were used, with charcoal separation of bound and free tracer. Within-assay reproducibility (CV%) was less than 20% in the concentration range 0.5-40 micrograms/L for both analytes and the comparable value for between-assay reproducibility was less than 25%. Results for external quality control samples were in good agreement with the expected values of 0-250 micrograms/L (spirapril, r = 0.997) and 0-300 micrograms/L (spiraprilate, r = 0.999). Overall, samples circulated to four laboratories gave good agreement for measured values, including one center using gas chromatography-mass spectrometry analysis for the two compounds. Data are presented to show the suitability of these two assays to the measurement of spirapril and spiraprilate in clinical samples from assays to the measurement of spirapril and spiraprilate in clinical samples from dose-ranging and bioequivalence studies. Results are also shown relating drug plasma concentration data to a measurement of the pharmacodynamic effects of spiraprilate, namely inhibition of angiotensin converting enzyme activity. It is concluded that these assays have the sensitivity for use in studies to model the relationship between the pharmacokinetics and pharmacodynamics of the two compounds.

Urinary creatinine as an index of urinary excretion of estrogen in cows prepartum and postpartum.

Urine was collected from 55 cows via indwelling urinary catheters for periods of 12 h on different days (28 days prepartum to 60 days postpartum). Excretion of urinary creatinine (mg/h per kg bodyweight) among Holsteins increased from .94 on day 28 prepartum to 1.14 on day .5 postpartum and then decreased to .82 on days 30 to 45 of lactation. Excretion of creatinine among 12-h collections of urine on different days did not differ for groups of cows within periods prepartum and postpartum, and coefficients of variation within cows were 12 to 13%. Ratio of urinary estradiol (-17 alpha) to urinary creatinine was correlated (.93) more highly with its excretion based on volume of urine excreted than was its urinary concentration (.79) within periods prepartum and postpartum. Ratios of urinary metabolites to urinary creatinine rather than their concentrations should be used to express rates of excretion in cows' urine when urine excreted per hour is unknown. Moreover, daily rates of excretion of estradiol in urine can be estimated [ng estradiol/day = A X ng/mg urinary creatinine x kg bodyweight x 24 h, where A is average excretion of urinary creatinine (mg/h per kg bodyweight)] for respective days prepartum and postpartum.

Plasma levels of progesterone in pseudopregnant rabbits actively immunized with a progesterone-protein conjugate.

PIP: The effects of active immunization with a progesterone protein conjugate on plasma progesterone levels in rabbits injected with a luteinizing dose of human chorionic gonadotropin (HCG) were determined. Mature female New Zealand rabbits were used. As antigen, a hemisuccinate of llalpha-hydroxyprogesterone was linked with bovine serum ablumin using the acetic anhydride method. Nuclear magnetic resonance analysis revealed that 24 moles of the hemisuccinate were linked with each mole of the protein. The conjugate was lyophilizied and stored at 4 degrees C. For immunization, an emulsion was prepared using equal volumes of Freund's complete adjuvant and .15 m NaCl solution of the immunogen. 7 rabbits received intradermal injections and 7 received weekly sc injections for 6 weeks. Sera were collected weekly for 6 weeks. Animals with high levels of antibody production were given iv injections of 1 mg of conjugate to boost antibody titers. Pseudopregnancy was induced in 14 rabbits by iv injection of 100 IU HCG. 5 weeks after the immunization schedule was finished animals were made pseudopregnant again and daily plasma progesterone was determined. Increased levels of antibodies to progesterone were found in all immunized animals but not in controls. Plasma progesterone levels in treated animals increased to a peak on Day 9 of pseudopregnancy. On Days 0 through 10, 17, and 18 progesterone values in immunized animals were significantly (p less than .05) higher than those of controls. It is concluded that immunization of rabbits by this method lead to abnormally high levels of peripheral progesterone in pseudopregnancy. I n a subsequent study it was found that despite the high plasma progesterone concentration produced in immunized animals, these females could be successfully mated.^ieng