High performance liquid chromatographic assessment of carotenoids’ stability during thermal food processing

Carotenoids are among of the most important bioactive compounds in food products due to their antioxidant, coloring, provitamin A and many health-related properties. Unfortunately, they are some of the most sensitive substances to heat, light, oxygen and acidic environments; hence, common food processing techniques cause important degradations of carotenoids, decreasing both the nutritional value and the appearance of carotenoid-containing foods. For assessing the stability of carotenoids during some thermal food processing procedures, the fruits of Cucurbita maxima Duch. (pumpkin) were selected, the main reason of this choice being both the amount and the diversity of carotenoids they contain, this study including both carotenes and xanthophylls. An optimized high performance liquid chromatography (HPLC) method was developed in order to properly quantify the involved carotenoids. Samples from both raw and processed fruits were extracted with acetone and diethyl ether; liquid-liquid extractions were then performed to transfer carotenoids in diethyl ether, being followed by saponification’s with methanolic potassium hydroxide. HPLC separations were achieved in less than 20 minutes, using a Nucleosil 120-3C₁₈ column, with a gradient involving acetonitrile : water (9 : 1) and ethyl acetate; detection was accomplished using a Waters photodiode array detector. The identification of carotenoids was achieved on the basis of their visible spectral characteristics, retention times amd co-chromatography with standards; for quantification the external standard method was used. A differential degradation of carotenoids was observed, depending on the processing method: during the thermal processing, both xanthophylls and carotenes were affected, zeaxanthin and violaxanthin being the most sensitive carotenoids, while cucurbitaxanthin A and lutein and were the most stable ones. The decrease with almost one third of the provitamin A activity during thermal processing was mainly due to the degradation of beta-carotene.