Pigments in Greek virgin olive oils: occurrence and levels Eleni Psomiadou and Maria Tsimidou* Laboratory of Food Chemistry and Technology, Department of Chemistry, Aristotle University of Thessaloniki, GR-54006 Thessaloniki, Greece Abstract: Fifty-two samples of virgin olive oil from various regions of Greece were examined for the presence and levels of chlorophyll and carotenoid pigments using normal phase liquid chromatogra- phy and spectrometry. Pheophytin a Pheo a) was the main pigment in all the oils examined >10mgkg 1 in more than 70% of samples). Two pheophytin a derivatives peaks A and B), eluted close to Pheo a, may potentially be used to examine handling conditions and length of storage of oil until analysis. Analysis of 25 Greek commercial oils obtained from retail stores veri®ed the above observations. Chlorophyll a may be present in a virgin olive oil just after production. Its absence should not be exclusively attributed to cultivar characteristics or extraction conditions, since pheophytinisation occurs rapidly. Lutein content varied between 0.2 and 3.9mg kg 1 and b-carotene content from 0.4 to 5.1mg kg 1 . The lutein/b-carotene ratio was characteristic <1) for samples from Koroneiki, the major Greek cultivar for oil production. The total Pheo a Pheo a  peak A  peak B)/ total carotenoid lutein  b-carotene) ratio ranged between 2 and 11 owing to prevailing green hues. This ratio may be used as an indicator of oil typicality along with other analytical parameters. # 2001 Society of Chemical Industry Keywords: chlorophyll; pheophytin; lutein; b-carotene; Greek virgin olive oil INTRODUCTION Colourisadecisivefactorthatdirectstheconsumerto preferdarkgreenoliveoilstolightyellowonesor vice versa according to previous experience and habits. Chlorophyllpigmentsareresponsibleforthegreen- ish hues of virgin olive oil. 1±4 The level of chlorophyll pigmentsfoundinoliveoildependsongeneticfactors, the stage of fruit ripeness, the extraction process and storageconditions.Amongchlorophylls,pheophytin a Pheo a)ispredominantlyfoundinthisoilaswellasin other edible plant oils. 5±7 Removal of Mg 2 to form pheophytin is the result of acidic conditions and/or heat. 8 The activity of a magnesium decheletase has also been postulated. 9 Pigment content is reduced with increasing fruit ripeness. Rahmani and Csallany 3 reported higher levels of Pheo a in oils from green olives than in oils from semi-black olives. Pheo b and chlorophyll a Chl a) were detected only in oils from greenfruits,whereasnoChl b wasfound.Theauthors suggested that the total chlorophyll content is mini- mumforoilsfrommatureolives,thusensuringgreater oxidative stability. The extraction process also causes lossesofoilpigments,mainlychlorophylls,owingtoa co-oxidation reaction provoked by hydroperoxide radicals formed by the action of lipoxygenase. 1 Chlorophyllase activity seems to be marginally re- sponsible for the loss of chlorophyll pigments during oil processing. 10 Usuki et al 11 and also Fraser and Frankl 5 stressed the practical importance of knowing the presence and levels of the different chlorophyll derivatives in oils, because of their different contribu- tions to oxidative stability. These pigments act as photosensitisers, promoting oxidation, while anti- oxidant activity was reported in the dark. 8 The major `yellow' pigments of virgin olive oil are luteinand b-carotene. 3,4,12 Carotenoidsareassociated with chlorophylls in all photosynthetic tissues, and their levels are in¯uenced by the factors previously mentioned, though to a lesser extent. 13 Lutein seems to be more vulnerable than b-carotene to acidic conditions. Other carotenes and xanthophylls have also been reported in very low concentrations. 4 The carotenoidlevelsofoliveoilsrangebetweenzeroanda fewmgkg 1 ,dependingontheoriginandcommercial type. Carotenoids, as singlet oxygen quenchers, protect oils from photo-oxidation, whereas their role in autoxidation is associated with the presence of tocopherols. 14,15 The aim of the present study is to characterise the major chlorophyll and carotenoid pigments in virgin oliveoilsofGreekoriginandassesstheirlevels.Toour knowledge,nosuchdataareavailableintheliterature. (Received 25 May 2000; revised version received 4 October 2000; accepted 7 February 2001) * Correspondence to: Maria Tsimidou, Laboratory of Food Chemistry and Technology, Department of Chemistry, Aristotle University of Thessaloniki, GR-54006 Thessaloniki, Greece E-mail: tsimidou@chem.auth.gr Contract/grant sponsor: Foundation of State Scholarships (IKY, Athens, Greece) Contract/grant sponsor: General Secretariat for Research and Technology, Greek Ministry of Development and Elais SA; contract/grant number: EPET II-691; contract/grant number: PAVE 99, No 99BE5 # 2001 Society of Chemical Industry. J Sci Food Agric 0022±5142/2001/$30.00 640 Journal of the Science of Food and Agriculture J Sci Food Agric 81:640±647 online: 2001) DOI: 10.1002/jsfa.859