Olive sterol

Several applications involve the removal of large amounts of triglicerides, including the determination of wax esters in olive oil (39), sterols and other minor components in oils and fats (40, 41), PCBs in fish (42), lactones in food products (43, 44), pesticides (45), and mineral oil products in food (46,47). Grob et al. (47) studied the capacity of silica gel HPLC columns for retaining fats, and concluded that the capacity of such columns is proportional to their size, although the fractions of the volumes that are then transferred to the GC system grow proportionally with the column capacity. For these reasons, 2-3 mm i.d. LC columns are to be preferred for LC-GC applications. 

R Lanuzza, G. Micali and G. Calabro, On-line HPLC-HRGC coupling and simultaneous transfer of two different LC fractions determination of aliphatic alcohols and sterols in olive oil , 7. High Resolut. Chromatogr. 19 444-448 (1996). 

Olive husks sc-co2 n.i. 0.4 n.i. unsaturated fatty acids sterols... 

Paquot, C., and H. Kaller. Evolutions of the sterols and the triterpenoid alcohols during the ripening of the fruits of the olive tree. C R Acad Sci SerC 1976 282 1041. 

There are cases where HPLC separation is performed not in order to quantify the alcohols but as a technique for the purification of the analytes to be subjected to further instrumental analysis. This is the case, for example, with the identification and determination of the structure of an abscisic acid in starfruit extract (Averrhoa carambola L.). The separation and purification of the analytes was carried out also with HPLC using a mobile phase of diethyl ether, whereas the structure was elucidated by H and UC-NMR (6). In a similar way, to separate the sterols and alkanols from the unsaponifiable matter from olive oils on a silica column, a gradient composed of hexane/diethyl ether was chosen in an offline system (7), whereas an online HPLC-HRGC system uses as its mobile phase hexane/isopropanol (8). 

Recently Amelio et al. (7) described a method which may find routine applications and which makes use of SPE for the separation and clean-up of the unsaponifiable from olive oil, from which the aliphatic alcohols are separated by means of HPLC (besides sterols and the two triterpenic dialcohols erythrodiol and uvaol). The alkanols are then derivatized and analyzed by means of HRGC. The use of an autosampler and a fraction collector for use with HPLC permits a considerable automatization of the analysis. (Reprinted from Ref. 1, p. 581, by courtesy of Marcel Dekker Inc.)... 

Fig. 5 HPLC of extravirgin olive oil unsaponifiable and collected fractions. A = fraction containing alkanols (6 and 9 min) S = fraction containing sterols (10 and 18 min). (Reprinted from Ref. 7 with the kind permission of Elsevier Science—NL, Sara Burgerhartstraat 25, 1055 KV Amsterdam, The Netherlands.)...

M Amelio, R Rizzo, F Varazini. Determination of sterols, erythrodiol, uvaol and alkanols in olive oils using combined solid-phase extraction, high-performance liquid chromatographic and high-resolution gas chromatographic techniques. J Chromatogr 606 179-185, 1992. 

K Grob, M Lanfranchi, C Mariani. Evaluation of olive oils through the fatty alcohols, the sterols and their esters by coupled LC-GC. J Am Oil Chem Soc 67(10) 626-634, 1990. 

Major flaxseed sterols are stigmasterol, camp sterol, and 5-5 avenasterol (Daun et al., 2003). Obtusifoliol, gramisterol, and citrostadienol constituted 45%, 22%, and 12%, respectively, of the total 4a-monomethylsterol in flaxseed (Kamm et al., 2001). Squalene content of flaxseed oil was reported as 4 mg/100 g oil, which was significantly lower than olive, corn, and rice bran oils. Squalene content is an intermediate compound of biosynthesis of plant sterols, which may have protective effects on lipid quality. Squalene could act as a peroxy radical scavengers in high polyunsaturated fatty acid oil (Dessi et al., 2002). 

Three other GC analyses now used in authentication, largely for olive and other oils which should not be refined or solvent extracted, are the determination of waxes, aliphatic alcohols, triterpene alcohols (uvaol and erythrodiol), and stigmastadiene and other sterol-dehydration products (EEC, 1991). These analyses are used at present not to detect adulteration with other oils, but with solvent-extracted or refined oils. However, it is possible that, with solvent-extracted oils, wax, aliphatic alcohol and terpene alcohol compositions, could prove useful in differentiating or detecting different oils. 

It is claimed (Youk et al., 1999) that olive, sunflower and peanut oils contain mainly esterified sterols, while soyabean and sesame oils contain mainly free sterols. This does not appear to have been utilized previously, but could be useful with mixtures of the two classes. Although it is possible to de-sterolize oils, and remove characteristic sterols, this usually forms other sterols that can be detected (Biedermann and Grob, 1996 Lanuzza and Micali, 1997 Mariani and Venturini, 1997). If it is suspected that this has occurred, then the presence of other suspect components should be investigated. 

There are a number of minor oils, all of high value, most of which are marketed mainly either for medical purposes or for their flavour. Olive, evening primrose, borage, fish oils and cocoa butter are described elsewhere. Others include hazelnut, walnut, macadamia, almond, apricot, pumpkin, poppy-seed and rice bran oils. The process of testing for authenticity of these oils should be approached in the same way as for the bulk oils above, i.e. fatty acid profile, sterols, tocopherols and triglyceride composition. However, there is little generally available published material on the ranges of values to be expected... 

Biedermann, M. and Grab, K. (1996) Detection of desterolized sunflower oil in olive oil through isomerized A7-sterols. Z. Lebensm. Unters. Forsch., 202, 199-204. 

Sterols, free and esterified, are a relatively abundant fraction of the minor components of olive oil. The regulation requires a concentration >1000mg/kg for virgin, refined and ordinary olive oil, whilst the content must be higher for the other types of olive oil (see Table 2.2, Annex I). 

These two pentacyclic triterpenes are found concentrated in the skin of the olive fruit. Extraction processes employing only physical means results in an olive oil with a low concentration of the two triterpenic diols, their highest limit having been fixed at 4.5% of the overall sterol concentration for oils 1-6. Olive-residue oils obtained by organic solvent extraction of pomace contain large amounts of uvaol and erythrodiol, their official limits respectively being >4.5 and >12% for olive-residue and olive-residue refined oil (see Table 2.2). 

Paganuzzi, V. (1983) Analysis of the sterol fraction of olive oil used for covering canned fish. Riv. Ital. Sost. Grasse, 60, 116-124. 

In some cases where extreme refining conditions have been used, complete removal of the sterols, both free and esterified, may occur (Grob et al., 1994). By removing the sterols from an oil in this way, it is possible to prepare an adulterant that is undetectable by sterol analysis. As with adulteration of virgin olive oil with refined olive oil, this type of adulteration may be detected by analysis of sterol degradation products. 

In olive oil, the main sterol is (l-sitosterol and this dehydrates to a mixture of 2,4- and 3,5-stigmastadienes on heating to high temperatures or in the presence of a bleaching earth (Grob and Bronz, 1994 Cert and Moreda, 1998). 

Cert et al. (1994) reported that bleaching caused the formation of the highest levels of stigmasta-3,5-diene and that bleaching temperature and earth type had an effect on sterol dehydration. These authors also reported that heating olive oil... 

Table 7.2 Authentication of mono varietal virgin olive oils comparative results of fuzzy logic algorithms (Calvente and Aparicio, 1995) and the supervised chemometric procedure of linear discriminant analysis. Chemical compounds used linolenic acid, 24-methylen-cycloarthanol sterol and copaene hydrocarbon...

---