Quantitative analysis of the principle soy isoflavones genistein, daidzein and glycitein, and their primary conjugated metabolites in human plasma and urine using reversed-phase high-performance liquid chromatography with ultraviolet detection.

Soy isoflavones are becoming of increasing interest as nutritional agents which can be used to combat osteoporosis and hyperlipidemia, and are also being considered as potential cancer chemopreventive compounds. However, prior to their formulation and distribution as therapeutic agents, thorough pharmacokinetic and toxicological assessment needs to be completed in men and women in a variety of health conditions in order to ensure their therapeutic efficacy and safety. At this time, studies of purified soy isoflavones are possible, and are being designed to fully evaluate the pharmacological utility of these preparations. In support of these studies, quantitative analysis of soy isoflavones in biological fluids can be accomplished with a wide variety of methods and analytical instrumentation. However, the relatively ubiquitous presence of high-performance liquid chromatography with ultraviolet detection (HPLC-UV) in most analytical laboratories, the relative ease of its operation, and the lesser expense of this instrumentation as compared to more sophisticated techniques such as liquid chromatography-mass spectrometry, offers some distinct advantages for its use in pharmacokinetic studies. In this manuscript, the development and validation of an HPLC-UV method for the quantitation of the principal soy isoflavones, genistein, daidzein, and glycitein, and their primary metabolites, in human plasma and urine is described. This analytical approach allows for pharmacologically relevant concentrations of the analytes and their principle metabolites to be detected, and has been validated in close agreement with the US Food and Drug Administration's guidelines for the validation of methods to be used in support of pharmacokinetic studies.

Effects of sex and gonadectomy on cocaine metabolism in the rat.

The purpose of the current study is to determine whether sex differences in metabolism of cocaine (COC) exist that could contribute to the greater behavioral sensitivity of females to COC administration. To investigate this question, concentrations of COC and its two principle metabolites benzoylecgonine (BE) and ecgonine methyl ester (EME) were measured by gas chromatography/mass spectroscopy in brain and plasma collected from male and female rats that were sacrificed between 5 and 90 min after injection COC (15 mg/kg i.p.). COC concentrations did not differ in plasma or brain tissue of males and females, but sex-specific patterns of metabolite distribution were detected. BE was 2-fold higher in plasma and brain of males than females, whereas EME was much higher in brain and plasma of females. The influence of gonadal hormones on COC metabolite patterns were determined using gonadectomized and prepubertal rats. Castration of male or female rats did not alter brain or plasma COC, but did decrease BE concentrations. Seven-day-old pups injected with 15 mg/kg of COC had higher blood and brain COC than adults and relatively low levels of metabolites. No sex differences were found for COC, BE, or EME in brain or plasma of pups. These findings indicate that although gonadal steroids influence COC metabolism, these effects do not explain sex differences in COC-induced behaviors.

Isolation and identification of novel metabolites of gemfibrozil in rat urine.

Gemfibrozil (GEM) is a clofibrate analog used to treat moderate to severe hypertriglyceridemias. In lab animals, GEM causes peroxisome proliferation, an effect that has been associated with hepatocarcinogenesis in rats. In humans, hepatobiliary disorders, but not carcinogenesis, have been associated with GEM therapy. In the present study [14C]GEM was administered orally to rats at a dose of 2000 mg/kg. At various time points, radioactivity in urine was analyzed by liquid scintillation spectrometry, high-pressure liquid chromatography, liquid chromatography/mass spectrometryn, gas chromatography/mass spectroscopy, and nuclear magnetic resonance. Nine metabolites of GEM were identified, some that have not been reported previously. Although the majority of metabolites were glucuronidated, some nonglucuronidated metabolites were identified in urine, including a diol metabolite (both ring methyls hydroxylated), and the product of its further metabolism, the acid-alcohol derivative (ortho ring methyl hydroxylated, meta ring methyl completely oxidized to the acid). Hydroxylation of the aromatic ring also was a common pathway for GEM metabolism, leading to the production of two phenolic metabolites, only one of which was detected in the urine in the nonconjugated or free form. Also of interest was the finding that both acyl and ether glucuronides were produced, including both glucuronide forms of the same metabolite (e. g., 1-O-GlcUA, 5'-COOH-GEM, and 5'-COO-GlcUA-GEM); the positions and functionality of the glucuronide conjugates were identified using base hydrolysis or glucuronidase treatment, in combination with liquid chromatography/MSn and nuclear magnetic resonance.

Computer-assisted measurement in diabetic patients with and without foot ulceration.

Computer-assisted measurement of sensibility was done at four anatomical sites of the foot of patients with diabetes with (19) and without (40) foot ulceration. Cutaneous pressure threshold measurements of patients with diabetes were compared with measurements on 30 patients without diabetes who had nonulcerated feet. Both groups had mean one-point static and moving touch, and two-point discrimination thresholds significantly different from the general population. Computer-assisted sensibility testing demonstrated significantly higher pressure thresholds for one-point static and moving touch and two-point discrimination in the ulcerated compared with the nonulcerated foot of patients with diabetes. This measurement technique is valid for documenting diminished sensation in the foot.

Use of Cryo/Cuff for the control of postoperative pain and edema.

Presented as a new modality, the Cryo/Cuff system incorporates a combination of hydrostatic pressure and cold to reduce post-traumatic pain and edema. It was hypothesized that this combination would also significantly reduce postoperative pain and edema. A sample population of 25 participants who were scheduled to have bilateral foot surgery was selected. Baseline circumference was determined preoperatively. Instruction and demonstration of the apparatus was given to the participants; an ice cap was used on the left foot as a control. The system is designed as a boot to be worn over the foot and ankle. Attached to the boot by a tube is a reservoir containing water and ice. Elevation of the reservoir may provide up to 40 mm. of mercury pressure. Study data gathered incorporated both evaluation by calculation of standard deviation above or below circumference mean as well as two subjective modes of patient assessment. Eighty percent of study participants experienced a reduction of pain and swelling at the first and subsequent postoperative visits. The Cryo/Cuff system appears to be an effective modality in the facilitation of the postoperative recovery process.

Treatment of painful diabetic neuropathy with capsaicin 0.075%.

An 8-week, double-blind, vehicle-controlled study was conducted to determine the effectiveness of topical capsaicin 0.075% cream in relieving pain associated with diabetic neuropathy. Patients were selected who experienced moderate to very severe pain, which interfered with sleep or activities on a daily basis, and who were unresponsive or intolerant to conventional therapy. The results after 8 weeks showed a statistically significant difference in favor of the capsaicin-treated patients, with 90% of these patients improved. The results of this study indicate that topical capsaicin 0.075% cream is safe and effective in managing painful diabetic neuropathy.