Pharmacokinetics of new 625 mg nelfinavir formulation during pregnancy and postpartum.

OBJECTIVES: Our objective was to evaluate the pharmacokinetics of nelfinavir (NFV) (625 mg tablets) 1250 mg twice daily during pregnancy and postpartum. METHODS: The participants were HIV-1-infected pregnant women enrolled in P1026s and receiving NFV (625 mg tablets) 1250 mg twice daily as part of routine clinical care. Intensive steady-state 12-h NFV pharmacokinetic profiles were performed during pregnancy and postpartum. The target NFV area under the plasma concentration-time curve (AUC(0-12)) was >or=10th percentile NFV AUC(0-12) in non-pregnant historical controls (18.5 microg h/mL). RESULTS: Of 27 patients receiving NFV, pharmacokinetic data were available for four (second trimester), 27 (third trimester) and 22 (postpartum) patients. The NFV maximum concentration (C(max)), 12-h post-dose concentration (C(12)) and AUC(0-12) were significantly lower during the third trimester compared to postpartum (P

Differentiation and characterization of enteroviruses by computer-assisted viral protein fingerprinting.

We have developed and standardized a computerized method for the typing and characterization of enteroviruses with radiolabeled viral protein fingerprints. Enteroviral proteins were radiolabeled with [35S]methionine during growth in cell culture and were then separated by polyacrylamide gel electrophoresis. The dried gel was scanned, and from the resulting computer image (which resembled an autoradiogram) protein patterns were computer extracted and stored in a database. The enterovirus database contained community and prototype strains belonging to 20 different enteroviral serotypes. Each serotype has a discrete protein pattern, and the most important pattern differences for determining each type are in the region of the viral capsid proteins VP1, VP2, and VP3. When the database was challenged with 148 clinical enterovirus strains, 144 (97%) were correctly identified by using the correlation coefficient as a quantitative measure of relatedness between two patterns. This method can identify a type in a single test and represents a practical alternative to virus neutralization because it is less expensive, is much faster (3 rather than 10 days), and does not rely on any virus-specific reagents. The results also show that most of the strains currently isolated from the community have protein patterns different from those of their older prototype strains. Viral protein fingerprinting is an evolving, dynamic system for the typing and characterization of enteroviruses. The method is appropriate for use in clinical virology and reference laboratories for the typing of enteroviruses, for the study of the epidemiology of enteroviruses, and for surveillance of enteroviruses.

Simple high-performance liquid chromatography method for the simultaneous determination of serum caffeine and paraxanthine following rapid sample preparation.

A simple reversed-phase high-performance liquid chromatography (HPLC) method for the simultaneous determination of caffeine and paraxanthine in human serum is described. Serum proteins are precipitated with perchloric acid and the resulting supernatant neutralized for direct injection onto an HPLC column. The method uses a phosphate-methanol mobile phase (85:15, v/v) at pH 4.9 with a flow-rate of 1.75 ml/min and quantitation is by UV absorbance at 274 nm. Elution times are approximately 18 min for caffeine and 8 min for paraxanthine. Theobromine and theophylline have elution times of 5.4 and 9.4 min and do not interfere in the assay. The intra-assay and between-assay means for precision and accuracy for both drugs are: 4.5% C.V. and 3.3% deviation. The sensitivity of the method is 50 ng/ml for each drug.

Prospective comparison of culture vs genome detection for diagnosis of enteroviral meningitis in childhood.

OBJECTIVE: To assess the sensitivity and specificity of a new polymerase chain reaction (PCR) assay with uninterrupted reverse transcription and complementary DNA amplification (RT-PCR) for the diagnosis of enteroviral (EV) meningitis in children. DESIGN: A prospective, cohort study. SETTINGS: Two medical centers: 1 university hospital and 1 children's hospital in San Diego County, California, during a 5-week period. PATIENTS: All pediatric patients younger than 16 years who underwent a lumbar puncture for evaluation of possible meningitis. MAIN OUTCOME MEASURES: The results of cerebrospinal fluid (CSF) RT-PCR were compared with viral cultures and clinical histories. RESULTS: During the 5-week period, 90 patients were entered into the study. Nonpolio EVs were cultured from 10% (9/90) of the patients from the following sites: CSF, 6.7% (6/90) of the patients; stool, 19% (4/21) of the patients; and throat swabs, 5.6% (1/18) of the patients. The EV genome was detected in the CSF by using RT-PCR in 7 of 9 EV culture-positive patients. The sensitivity and specificity of the CSF RT-PCR assay to detect EV meningitis were 77.8% and 100%, respectively. This compared with a sensitivity of 66.7% for detection of EV in CSF by viral culture alone. CONCLUSION: The new RT-PCR assay is a rapid and reliable method for the detection of EV infection in childhood.

Changes in the concentration of 5-methoxytryptophol in the circulation at different phases of the human menstrual cycle.

The pineal indole 5-methoxytryptophol (ML) has been shown to have an antigonadal activity when administered to experimental animals, but data on its normal pattern of secretion have been lacking. Using a new gas chromatography-mass spectrometry assay, the concentration of ML at various phases of the human menstrual cycle has been studied. Daily samples were obtained throughout the month from five women with a normal cycle and two women taking an oral contraceptive. In women with a normal cycle levels of ML were found to be significantly lower in the last third of their cycle; this change was not seen in women taking an oral contraceptive who had low levels throughout the month. The changes in concentration of ML did not correlate with the changes in concentration of gonadotrophins.

Local degradation of luteinizing hormone-releasing hormone (LH-RH) in the rat central nervous system.

Peptidases inactivating luteinizing hormone-releasing hormone (LH-RH) were found to be present in two subcellular fractions prepared from the following areas of the central nervous system of male and female rats: thalamus, hypothalamus, cortex and cerebellum. LH-RH immunoreactivity was also detected in these fractions after peptidase activity had been removed by heat denaturation. These findings support the concept that LH-RH may have a role in modulating neuronal activity within the central nervous system (CNS).

Inactivation of somatostatin by peptidases in different areas of the rat brain.

With the availability of a sensitive and specific radioimmunoassay for growth hormone-release-inhibiting hormone (somatostatin or GH-RIH), it has been possible to investigate the presence of peptidase enzymes capable of inactivating this hypothalamic hormone in the hypothalamus and other brain areas of the rat. It was found that both supernatant and particulate fractions from male rat hypothalami rapidly inactivated somatostatin and that the enzymes involved have an optimum pH of 7.3. Peptidase activity was significantly higher in the supernatant than in the particulate fraction from the hypothalamus, thalamus, cortex and cerebellum. Besides confirming the presence of peptidases inactivating the release-inhibiting hormone in the hypothalamus (the site of somatostatin synthesis and release), the results may indicate that somatostatin has a functional significance outside the hypothalamus-anterior pituitary axis but within the central nervous system.

Local degradation of growth hormone-release-inhibiting hormone (somatostatin) in the central nervous system.

Peptidases inactivating growth hormone-release-inhibiting hormone (somatostatin) were found to be present in two fractions prepared from the thalamus, hypothalamus, cortex and cerebellum of male and female rats. Somatostatin was also detected in significant quantities in fractions from the thalamus, hypothalamus, and cortex, after peptidase activity had been removed by heat denaturation. These results support the suggestion that somatostatin may function as a neurotransmitter or modulator of neuronal activity in the central nervous system.

Leteinizing hormone and luteinizing hormone releasing hormone-like immunoreactivity in the jugular venous blood of sheep at various stages of the oestrous cycle.

Luteinizing hormone and LH-RH-like immunoreactivity were measured in the jugular venous plasma of Clun Forest ewes at various stages of the oestrous cycle. Blood samples were collected through jugular venous cannulae every 2 h for at least 20 days from three ewes during the breeding season. The ewes were checked twice daily for oestrus using a vasectomized ram. Plasma LH peaks of apparent height 112-192 ng NIH-LH-S17 equivalents/ml were detected at oestrus with basal levels of 2-15 ng/ml during most of the remainder of the 17-day oestrous cycle. Peaks of LH-RH-like immunoreactivity occurred at various times of the cycle. The apparent maximal level of these peaks was 220 pg/ml compared with basal levels of less than 10 pg/ml. Further ewes (two for each group) were sampled at 4 min intervals for 12 h, (1) from onset of oestrus, (2) 36-48 h after onset of oestrus or (3) on day 10 of the oestrous cycle. In the ewes sampled at oestrus, peaks of LH-RH-like immunoreactivity were detected before, during and after the preovulatory LH peak. Those detected after the LH peak were unassociated with any further increases in the plasma LH level. In the ewes sampled 36-48 h after onset of oestrus and on day 10 of the cycle, several peaks of LH-RH-like immunoreactivity unassociated with any increases in the LH level were detected. These peaks, and those detected at oestrus, had durations of only one or two samples, and in some cases reached levels of several ng/ml compared with basal levels of less than 10 pg/ml. The significance of these results is discussed.

Effect of neonatal androgen on the activity of peptidases in the rat brain inactivating luteinizing hormone-releasing hormone.

The activity of peptidase enzymes in different brain areas inactivating luteinizing hormone-releasing hormone (LH-RH) was investigated in normal male and female rats treated neonatally with androgen. Of the two fractions containing peptidase activity, changes were found only in the supernatant (soluble/cytoplasmic) fraction and were restricted to the hypothalamus and cortex. In both these brain areas, peptidase activity was considerably higher in androgen-treated than in normal female rats; there was no significant difference between activity in male rats and the androgen-treated female rats. These findings indicate that neonatal androgen produces masculinization of the mechanisms responsible for LH-RH inactivation in the hypothalamus which may reflect changes observed elsewhere in gonadotropin secretion in androgen-treated female rats. The changes in peptidase activity in the cortex may infer that sexual differentiation of the brain under the influence of androgen is not limited to the hypothalamus and can be manifested at extrahypothalamic sites within the central nervous system.

The presence of peptidases in the rabbit hypothalamus capable of inactivating luteinizing hormone-releasing hormone.

The presence of peptidases in the rabbit hypothalamus capable of inactivating luteinizing hormone-releasing hormone (LH-RH) was investigated with the use of a sensitive and specific radioimmunoassay for the releasing hormone. Enzymes acting on LH-RH were found in two fractions from rabbit hypothalami, a particulate fraction and a supernatant fraction, which very rapidly inactivated the decapeptide. Peptidase activity in the particulate fraction from male and female rabbits was approximately the same, but the supernatant fraction from female animals had significantly greater peptidase activity than that from male animals. These findings confirm the hypothesis that rabbit hypothalamus contains peptidases which inactivate LH-RH, and give some indication that the enzymes may be involved in the control by the central nervous system of reproductive function.

Further studies on the nature of the immunoreactive luteinizing hormone-releasing hormone (LH-RH)-like peptide in human urine.

The chemical nature of the immunoreactive LH-RH-like peptide found in human urine has been investigated using ion-exchange chromatography on carboxymethyl (CM)-cellulose and a radioimmunoassay for LH-RH. A single immunoreactive substance was found in urine after LH-RH administration and in urine samples from untreated subjects. This substance did not have the mobility of either the synthetic decapeptide nor the 3-10 octapeptide on CM-cellulose and the evidence suggests that it may be the 2-10 nonapeptide of LH-RH.