Thierry Trenque

Simultaneous determination of amoxicillin and clavulanic acid in human plasma by HPLC with UV detection.

A simple and accurate high-performance liquid chromatographic method with ultraviolet detection at 220 nm has been validated for the simultaneous determination of amoxicillin and clavulanic acid in human plasma. Plasma samples were pretreated by direct deproteinization with methanol. A good chromatographic separation between both compounds was achieved using a reversed phase C8 column and a mobile phase, consisting of acetonitrile-phosphate solution-tetramethyl ammonium chloride solution. The calibration curves were linear over the concentration range of 0.625-20 mg l(-1) for amoxicillin and 0.3125-10 mg l(-1) for clavulanic acid with determination coefficients > 0.998. The method is accurate (bias < 7%) and reproducible (intra- and inter-day R.S.D. < 15%), with a quantitation limit of 0.625 and 0.3125 mg l(-1) for amoxicillin and clavulanic acid, respectively. Analytical recoveries from human plasma ranged from 91 to 102% for both components. This fully validated method, which allows the simultaneous measurement of amoxicillin and clavulanic acid in biological samples, is rapid (total run time < 10 min) and requires only a 100 microl sample. This assay is suitable for biomedical applications and was successfully applied to a pilot pharmacokinetics study in healthy volunteers after a single-oral administration of amoxicillin/clavulanic acid combination (500/125 mg).

Osteonecrosis in Six HIV-Infected Patients Receiving Highly Active Antiretroviral Therapy

OBJECTIVE To report 6 cases of osteonecrosis in HIV-infected patients treated with highly active antiretroviral therapy (HAART) and compare the observed risk factors with those of published cases. CASE SUMMARIES Osteonecrosis was diagnosed between 1999 and 2002 in 6 of 417 HIV-infected patients in our department of infectious diseases. At the time of diagnosis, mean patient age was 42 years, and 5 patients had developed AIDS. Mean CD4+ lymphocyte count was 563.5 cells/mm3 and viral load was undetectable (<50 copies/mL) in 5 patients. The patients’ mean body mass index was 22.5 kg/m 2 . Four had lipodystrophy. All were receiving HAART, including a protease inhibitor in 4 patients; the remaining 2 patients had a history of protease inhibitor treatment. Median time from the first antiretroviral therapy to osteonecrosis diagnosis was 46.5 months. Established risk factors were the use of corticosteroids in 2 patients and dyslipidemia in all patients. All of the patients developed pain and functional impotence of the hip or ankle joints. Osteonecrosis of the hip was bilateral in 4 cases. Three patients required surgical intervention, all of whom had favorable outcomes. DISCUSSION HIV-infected patients are at a higher risk for the development of osteonecrosis and are more likely to be exposed to predisposing factors to its development. The HAART implication as a predisposing factor remains controversial. CONCLUSIONS The pathogenesis of osteonecrosis in HIV-infected individuals may be multifactorial; the reasonable approach for clinicians consists of treating concomitant predisposing conditions that might further cause osteonecrosis.

Simultaneous determination of inulin and p-aminohippuric acid (PAH) in human plasma and urine by high-performance liquid chromatography

Inulin and p-aminohippuric acid (PAH) clearances are used for the estimation of glomerular filtration rate (GFR) and effective renal plasma flow (ERPF). A simple and rapid high-performance liquid chromatography (HPLC) method with UV detection is described for the simultaneous determination of inulin and PAH in the same chromatogram in the plasma and urine of humans. Plasma and urine samples were hydrolyzed with perchloric acid (0.7%) in boiling water. The mobile phase consisted of 0.01 M potassium dihydrogenphosphate with 0.02 M tetramethylammonium chloride and o-phosphoric acid (pH 3)-acetonitrile (94:6, v/v), pumped at a rate of 1.2 ml min-1 on a C8 reversed-phase column. Tannic acid was used as the internal standard and UV detection at 285 nm was employed. The calibration curves were linear over the concentration range of 12.5-100 mg l-1 for inulin and 6.25-50 mg l-1 for PAH with determination coefficients greater than 0.997. The method is accurate (bias < 13%) and reproducible (intra- and inter-day relative standard deviation less than 11%), with a limit of quantitation of 12.5 mg l-1 and 6.25 mg l-1 for inulin and PAH, respectively. Analytical recoveries from urine and plasma were ranged from 81 to 108% for both compounds. This fully validated method, which allows the simultaneous determination of inulin and PAH clearances, is simple, rapid (total run time < 10 min) and requires only a 200 microliters plasma or urine sample.

Bromism from Daily Over Intake of Bromide Salt

Bromide intoxication today is an infrequent disease, but preparations containing bromide are still available in nonprescription compounds, on the French market. We report a case with bromide intoxication due to daily over intake (∼20 tablets per day; i.e. total elemental bromide intake ∼ 6 g/day) of calcium bromo-galactogluconate (Calcibronat®) for 1.5 months. A 30-year-old woman with a long history of psychotropic drug abuse was hospitalized in a psychiatric department for neuropsychological manifestations. She presented a seriously disturbed mental status with confusion, disorientation, auditory and visual hallucinations, and loss of short-time memory. A markedly increased serum bromide level of 1717 mg/L (21.5 mEq/L) measured on the first day after her admission confirmed the diagnosis of chronic bromism suspected based on her symptomatology. During her hospitalization, bromide plasma concentrations were measured and monitored using inductively coupled plasma mass spectrometry, a sensitive and very specific method. After withdrawal of the drug, the symptoms improved within 8 days. Serial bromide concentrations gradually declined throughout nearly 2 months of monitoring, until she was discharged from the hospital. We found an elimination half-life of bromide in blood of approximately of 10 days. This case demonstrates that, while today bromism occurs infrequently, it should still be included in the differential diagnosis of neuropsychiatric symptoms.

Gas chromatographic determination of meprobamate in human plasma

A simplified and rapid gas chromatographic method has been developed for the determination of meprobamate in human plasma. The procedure includes a single-step extraction of alkalinized sample with chloroform, and chromatography on a non-polar fused-silica capillary column with flame ionization detection. The method is accurate (97.7 +/- 5.7% at 20 mg/l) and precise (maximum coefficient of variation of 9.5%). It provides an alternative to existing methods and is particularly suitable for toxicological studies.

An unusual case of methyl bromide poisoning.

A nonlethal poisoning case by methyl bromide in a young woman due to leakage of old fire extinguishers is described. The patient developed major action and intention myoclonus the day following exposure. Inorganic bromide concentrations in plasma were determined by inductively coupled plasma mass spectrometry. The initial plasma bromide level was 202 mg/L, 40-fold in excess than the commonly accepted tolerance limit, and decreased slowly to normal levels within 2 months. Although plasma inorganic bromide concentration is known not to be directly correlated to the severity of organic bromide poisoning, its determination was, in the present case, particularly useful to confirm the diagnosis. One year post-exposure, the patient showed no sign of central nervous system toxicity. While such a case of poisoning is particularly rare today, it illustrates, however, that the danger still exists in France although the destruction of fire extinguishers containing methyl bromide has been sought in France for more than 30 years.

Sensitive high-performance liquid chromatographic method for the determination of coumarin in plasma

A high-performance liquid chromatographic method was developed for the determination of coumarin in plasma at low concentrations. The method involves a single-step extraction of the alkalinized sample with hexane and subsequent evaporation of the organic phase in the presence of hydrochloric acid to collect and concentrate the coumarin. Analysis of the acidic phase was performed on a C8 column and coumarin was detected by measuring the UV absorbance at 275 nm. The limit of detection was 0.3 microgram l-1. The assay was used to study the evolution of concentrations of coumarin in one volunteer after oral administration of a single 10-mg dose.