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Among more than 50 provitamin carotenoids, beta-carotene is the most metabolically active source of retinol. Despite diets rich in fruits and vegetables containing beta-carotene, vitamin A deficiency is the leading cause of blindness and childhood mortality in developing countries. In addition, the uncertainty of beta-carotene bioconversion into vitamin A suggests that new data are needed to update the nutritional guidelines in developed countries. Previously, we reported the development of a carotene/retinol plateau isotopic enrichment method (CarRet PIE) for the determination of beta-carotene bioavailability and bioconversion into retinol, which utilizes positive ion atmospheric pressure chemical ionization (APCI) liquid chromatography/mass spectrometry (LC/MS). While seeking to validate the CarRet PIE using a mass balance approach requiring fecal measurements of beta-carotene and retinol, interference was encountered that required substantial modifications of the LC/MS assay. Here we report a new LC/MS assay that is based on the detection of molecular anions of beta-carotene using negative ion APCI with a reversed-phase C30 column for HPLC separation. Sample preparation required saponification to eliminate interfering triglycerides. The limit of detection (LOD) of beta-carotene was 0.25 pmol calculated on the basis of an injection of 20 microL of 0.0125 microM beta-carotene, and the limit of quantitation (LOQ) was 1.0 pmol based on the injection of 20 microL of 0.050 microM beta-carotene. The linear range was 1.1 to 2179 pmol on-column. The wide linear range and low LOD and LOQ of this assay facilitated the sensitive and selective quantitative analysis of beta-carotene in both serum and fecal samples in support of an on-going clinical investigation of beta-carotene bioavailability and bioconversion into vitamin A.

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Background: Glucocorticoid excess promotes visceral obesity and cardiovascular disease. Ligand availability to the glucocorticoid receptor is controlled by isoforms of 11β-hydroxysteroid dehydrogenase (11β-HSD) which converts endogenous cortisone to active cortisol. Aim: To evaluate the expression and activity of 11β-HSD1 in subcutaneous adipose tissue (SC) and visceral adipose tissue (VAT) in prepubertal children with normal weight. Methods: Fourteen patients (11 female/3 male) with a mean age of 6.9 ± 0.9 years and a body mass index (BMI) of 17.4 ± 0.61 underwent elective open abdominal surgery. Results: Expression of 11β-HSD1 mRNA in SC and VAT was similar (0.8 ± 0.15 vs. 0.61 ± 0.12 AU). The activity of this enzyme in SC was significantly lower compared to VAT (1.42 ± 0.39 vs. 2.79 ± 0.61 ng cortisol/g tissue/24 h, p < 0.05). In addition, we observed a significant direct correlation with the expression of 11β-HSD1 in VAT adipose tissue with the patient’s BMI (r = 0.825, p = 0.002). Conclusions: This correlation together with the increased activity of this enzyme in visceral adipose tissue might contribute to decreased hepatic insulin sensitivity due to increased portal cortisol when BMI increases. These observations appear to be particularly important in children born with low birth weight who develop rapid early weight gain.

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