THE ANALYSIS OF CAROTENOIDS AND RETINOIDS - A REVIEW

In recent years, there has been particular emphasis on obtaining more accurate data on the types and concentrations of various carotenoids and retinoids in foods for various health and nutrition activities. A great deal of attention has thus been given to improved methodologies for the study of these two groups of compounds. This review is aimed at defining the state-of-the-art of analytical methods for the study of carotenoids and retinoids, particularly in foods and blood samples. The first part of the review discusses various preliminary sample treatment procedures, including precautions to be taken during handling of samples. The greater part of the discussion is devoted to reviewing the various methods reported for the separation and quantitation of these two groups of compounds. A pre-requisite in the analysis of carotenoids and retinoids is preliminary sample treatment in order to release, isolate and extract the compounds of interest from the food matrix. Numerous different preliminary treatment procedures have been reported in the literature. In general, however, these procedures follow a few basic steps, upon which various modifications have been made. For convenience, the treatment procedures for plant materials, animal foods and blood samples are separately treated in this review. Because of the unstable nature of the carotenoids and retinoids, especially towards light, heat, oxygen and acids, various precautions have to be taken during saponification, extraction and chromatography steps of the analytical procedure. Precautions against oxidation, such as the use of antioxidants, protection against exposure to direct light, and proper storage of solutions are discussed. After sample extraction, most investigators endeavoured to separate the carotenoids and retinoids from interfering substances in order to obtain more accurate results. A wide variety of separation and detection and quantitation procedures have been used in studies of these compounds. There has been a general trend in the change of these methodologies over the last two decades. An early technique used for the separation of carotenoids in plant materials, mainly in the 1960s, was countercurrent distribution. A few early studies using paper chromatography were also reported. In the 1970s, a few studies of carotenoids and retinoids using gas-liquid chromatography and gel-permeation chromatography were encountered. A number of studies employing thin-layer chromatography (TLC) as a single technique for the separation of carotenoids in plant materials were encountered throughout the 1970s and the early 1980s. TLC was also used by several investigators in combination with other separation techniques. The procedure was less widely used in studies of retinoids. Adsorption (open) column chromatography, utilizing primarily descending, gravity-flow columns, was widely used for the study of carotenoids and retinoids in foods, even in the 1960s. The procedure remained very much in use in the 1970s and 1980s. Commencing from the late 1970s, a new chromatography procedure became more prominent in the literature. High-pressure liquid chromatography has become a widely used procedure for the separation of carotenoids and retinoids in various materials. Owing to the complicated nature of the required analysis, the associated methodologies are still being developed and improved. Used together with adsorption column chromatography and UV-vis spectroscopy, especially during method development stage, HPLC would be the method of choice for arriving at more accurate data on the content and composition of carotenoids and retinoids in foods.