Sample hydrolysis carotenoids

Alkaline hydrolysis (saponification) has been used to remove contaminating lipids from fat-rich samples (e.g., pahn oil) and hydrolyze chlorophyll (e.g., green vegetables) and carotenoid esters (e.g., fruits). Xanthophylls, both free and with different degrees of esterification with a mixture of different fatty acids, are typically found in fruits, and saponification allows easier chromatographic separation, identification, and quantification. For this reason, most methods for quantitative carotenoid analysis include a saponification step. 

Food sample pretreatment may consist of either (a) saponification to quantify the free forms (retinol or xanthophylls may occur free or ester-ified in foods) [95,96] or (b) direct extraction to determine the unaltered A vitamers [84,88]. Alkaline hydrolysis is also an expedient to simplify the vitamin A analysis, since retinol is the only form to be quantified nevertheless, due to its sensitivity to light and oxygen, it is important to prevent photo-oxidation by inclusion of a antioxidant (ascorbic acid, hydroquinone, or pyrogallol). A drawback of hot saponification is the generation of artifacts, such as geometric isomers of retinol and carotenoids [97]. 

In contrast, most carotenoids are considerably stable to the action of alkalis, enabling saponihcation as a routine procedure to eliminate fatty matter and chlorophylls, or for hydrolysis of the esters of xanthophylls with fatty acids. Certain carotenoids are, however, alkali-sensitive, notably astaxanthin and in general the carotenoids containing 3-hydroxy-4-oxo-(3 end groups. When analyzing samples containing such carotenoids, contact with alkalis should be avoided. 

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