PhD

In January 1997, the 'Netherlands Foundation for the Advancement of Tropical Research' (WOTRO-NWO) appointed me as a PhD fellow to resume the development of a stable isotope method in a research project entitled: 'Measurement of the bioconversion of ß-carotene to retinol in children in Indonesia'.

The research in this project was carried out within the framework of collaboration between the Division of Human Nutrition and Epidemiology of Wageningen University, the Nutrition Research and Development Centre in Bogor, Indonesia, the Department of Medicinal Chemistry and Pharmacognosy of the College of Pharmacy of the University of Illinois at Chicago, USA, and the Leiden Institute of Chemistry of Leiden University, in Leiden.

This project resulted in a PhD thesis, entitled:

Vitamin A deficiency is a serious health problem in many developing countries. Improved vitamin A status can reduce morbidity and mortality by 23% in developing countries. Vitamin A occurs in food as preformed vitamin A (retinol), present in animal foods and breastmilk, and as provitamin A carotenoids - the major source of vitamin A for a large proportion of the world's population. However, the contribution of plant foods to vitamin A status can only be substantial when not only the consumption and provitamin A content of foods but also the bioefficacy of provitamin A carotenoids in such foods is high. With respect to provitamin A carotenoids, bioefficacy is the product of the fraction of the ingested amount which is absorbed (bioavailability) and the fraction of that which is converted to retinol in the body (bioconversion). Isotopic tracer techniques can supply accurate and precise data on bioavailability, bioconversion and bioefficacy of carotenoids in humans.

A new stable isotope technique - based on reaching plateau isotopic enrichment of ß-carotene and retinol in serum during prolonged intake of multiple low doses of ß-carotene and retinol, each specifically labeled with 10 ¹³C atoms - has been developed. In 2 studies, for periods of up to 10 wk, >100 Indonesian children (aged 7-13 y), consumed daily 2-3 doses of specifically labeled [¹³C₁₀]ß-carotene (<80 µg/dose) and [¹³C₁₀]retinyl palmitate (<80 µg/dose). In the second study, half of the subjects (n = 41) daily also received spinach while the others (n = 36) received orange pumpkin. Either in serum or in feces, the degree of isotopic enrichment of retinol with [¹³C₅]retinol - derived from administered [¹³C₁₀]ß-carotene - and with [¹³C₁₀]retinol, and of ß-carotene with [¹³C₁₀]ß-carotene was measured by high performance liquid chromatography with atmospheric pressure chemical ionization liquid chromatography-mass spectrometry (APCI LC-MS). These techniques have emerged as the most effective and convenient for studying the bioavailability, bioconversion and bioefficacy of provitamin A carotenoids. For interpretation of data, a mathematical model has been developed based on assumptions that can be readily justified in the light of present knowledge on carotenoid metabolism.

From the studies presented, it can be concluded that 2.6 µg ß-carotene in oil has the same vitamin A activity as 1 µg retinol (bioefficacy of ß-carotene in oil, 36%). The bioavailability of ß-carotene in oil was found to be 86%. The bioavailability and bioefficacy of ß-carotene in pumpkin were 1.7 times those of ß-carotene in spinach. Results obtained using data from feces confirmed results obtained using data from serum.

This plateau isotopic enrichment technique can be applied for studying the effect of individual SLAMENGHI factors which affect the bioavailability and bioefficacy of carotenoids in humans (SLAMENGHI is a mnemonic for such factors). The studies presented have provided the most reliable estimates of the bioefficacy of ß-carotene in oil available to date. The estimates are most probably applicable to other children in developing countries. Results from recent studies and from those presented in this thesis indicate that the current guidelines that the vitamin A activity of 1 µg retinol can be supplied by 6 µg (FAO/WHO) or 12 µg (US Institute of Medicine) of ß-carotene in a mixed diet overestimate ß-carotene bioefficacy about 2 to 3 times. This means that approaches other than the promotion of the consumption of fruit and vegetables are required for eliminating vitamin A deficiency.