Analysis of sterols

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. 

Standard methods of analysis of total sterol content of oils involve saponification of the oil, followed by extraction and isolation of total sterols from the unsaponihable fraction by thin layer chromatography (TLC) (AOCS, 1998). Quantification of individual sterols involves silylation of the sterol fraction and analysis by gas chromatography (GC). Sterols and steryl esters in oils and fats can be analysed by LC-GC after silylation or acylation of the free sterols (Artho et al., 1993). An alternative approach to the analysis of intact steryl esters involves separation of sterols and steryl esters by solid phase 

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In milk fat, cholesterol is associated with Hpoproteins in the milk fat globule. It is also a component of animal membranes and controls rigidity and permeabihty of the membranes. Cholesterol has interesting surface properties and can occur in Hquid crystalline forms. Plants contain sterols such as P-sitosterol [83-46-5] (4b) or stigmasterol [83-48-7] (4c). Their functions in plant metaboHsm are not yet well understood. Analysis of sterols has proven useful for detection of adulteration of edible fats (9). 

The analysis of sterols, sterols esters, erythrodiol and uvaol, and other minor components of oils and fats, is usually carried out by normal-phase HPLC-HRGC by using a loop-type interface and the concurrent eluent evaporation technique, as reported in the papers cited by Mondello et al. (48) (up to 1995) and in more recent papers (49, 50). More recently, reversed-phase LC-GC methods have been... 

Sterol analysis and quantification can be performed using plasma, serum, and cerebrospinal fluid (CSF) samples. Analysis of sterols from tissues and cells requires a pre-extraction work-up. The requirements for the different specimens are as follows ... 

Jeong, T.M., Itoh, T., Tamura, T. and Matsumoto, T. (1974) Analysis of sterol fractions from twenty vegetable oils. Lipids, 9, 921-927. 

The presence of another triterpene alcohol, taraxasterol, has been reported in shea (Itoh et al, 1979 Dencausse, 1995), and also in illipe and sal fat (Soulier et al, 1990). These relatively minor sterols have been little studied or reported and should be included in the analysis of sterolic fractions of confectionery fats. 

In the analysis of sterols and other components of CBAs, in particular the minor components it must be borne in mind that it is selected fractions of the parent fats obtained by crystallization that are used in confectionery and are potential adulterants. These fat fractions are produced with a view to obtaining desired physical properties, which depend on the triacylglycerol composition, and while they do contain measurable levels of some minor components of varying polarity their composition is likely to vary significantly from that of the parent fat. 

Crews, C., Calvet-Sarret, R. and Brereton, P. (1997) The analysis of sterol degradation products to detect vegetable fats in chocolate. J. Am. Oil Chem. Soc., 74(10), 1273—1280. 

Homberg, E. (1991) Analysis of sterols as a means for detection of blended and adulterated fats. Fett Wissen. Technol., 93, 516-517. 

The analysis of minor components is a useful method of detecting adulteration of some pressed oils. For many refined oils, analysis of sterols or tocopherols may be less likely than analysis of bulk components, especially TAGs, to be an effective method of detecting adulteration, but the minor component composition may be helpful when considered with other analytical data. 

Goad, L.J. Akihisa, T. 1997. Analysis of Sterols. Blackie Academic and Professional, Chapman Hall, London, Great Britain. 

Rahier, A. Benveniste, P. 1989. Mass speetral identifieation of phytosterols. In Analysis of Sterols and Other Biologieally Signifieant Steroids (Nes, W.D. Parish, E.J.,Eds.) Academic Press, San Diego, pp. 223-250. 

The analysis of sterols and sterol esters has been proposed as one way to identify oils in blends (311, 312). Johansson and Croon (313) discussed the use of 4-des-methyl-, 4-monomethyl-, and 4,4-dimethylsterols in characterizing different vegetable oils, and the results are summarized in Table (8). The levels of total sterols and sterol classes as well as the relative distribution of the individual sterol members vary between oils. The presence of steradienes can also be used as a marker for the presence of refined oils (314, 315). High oleic acid oils can easily be used to adulterate olive oil. The presence of rapeseed oil in other oils can be detected by the analysis of brassicasterol and its dehydration product, campestatriene. The presence... 

Figure 1. Sterol composition of DM1 resistant and sensitive strains of three fungal pathogens. Details of the methods used for the extraction and analysis of sterols are given In the relevant source papers (36-38). Either cholesterol or cholestanol were used as internal standards.

Molecular Analysis of Sterol 14 a Demethylase Gene from Barley... 

Conventional marine sterols have A -, A - or A Miene nuclei, but the occurrence of A -sterols was reported in Axinella cannabina [36]. Recently a complete analysis of sterols in the same species by Itoh et al. [37] showed the presence of 74 sterols with a variety of nuclei A , A, A , A, Sa-saturated and 5a-methoxy-... 

Fig. 12. Extraction and analysis of sterols from seawater. (1) At this stage, sulphur removal with an activated Cu-column may be necessary. The sample extracts may also be further purified by chromatography on silica gel (see Gagosian and Heinzer, 1979).

Laakso, P., Analysis of sterols from various food matrices, Eur. J. Lipid Sci. TechnoL, 107,402, 2005. 

Figure 2.10 Reconstructed MRM traces for Bligh/Dyer extract of sterols from mouse brain. Known sterols are labeled, unknown compounds with a sterollike signature are indicated by . MS/MS of the respective steroids can be found at Lipid Maps http //www.lipidmaps.org/data/standards/standards. php lipidclass=LMST. (Reproduced with permission from McDonald, J. G. et al., 2007, Extraction and Analysis of Sterols in Biological Matrices by High-Performance Liquid Chromatography Electrospray Ionization Mass Spectrometry, Methods Enzymol. 432 145-70.)...

McDonald, J. G., Thompson, B. M., McCrum, E. C., and Russell, D. W. 2007. Extraction and analysis of sterols in biological matrices by high performance liquid chromatography electrospray ionization mass spectrometry. Methods Enzymol. 432 145-70. 

The data presented here demonstrate a reliable methodology for monitoring the formation and quantifying the presence of oxidized products of cholesterol. This approach can now be applied towards the analysis of these materials in any type of medium. The synthesis of nona-labeled cholesterol and its use as an internal monitor permit reliable analysis by capillary GC as well as GC-MS. For the analysis of oxides isolated from complex matrices, GC-MS is the desirable analytical method because of its high selectivity. However, the demonstrated applicability of conventional capillary GC provides an inexpensive and more readily available alternative which can be used as an initial screening method. These results provide a basis for new analytical procedures which can be applied to the analysis of sterols of universally recognized medical significance. 

It is also possible to perform GLC on the trimethyl-silyl ethers of the sterols. In that case, the residue is gently heated with a mixture of dimethylformamide, hexamethylsilazane and trimethylchlorosilane to prepare the trimethylsilyl ethers. The gas chromatographic method described provides a qualitative analysis of sterols in oils and fats. However, it can be employed for quantitative analysis within certain limitations (a) the response of each sterol is a function of the type of detector used (b) an accurate quantitative interpretation of the chromatogram... 

GLC is applied to the analysis of fats and fat components in several ways. Typical data for GLC is shown in Table 1. This includes the analysis of FA composition after derivatizing the FAME (or propyl esters for butter fat) and the direct analysis of TAG composition. GLC is used for the analysis of sterols, hydrocarbon contaminants, pesticides, and volatile products produced during fat processing and refining. 

Recently, LC/MS has made significant contributions in the analysis of sterols and steryl esters. Takatsu and Nishi (1993) have employed discharge-assisted LC/TS/MS for the determination of total serum cholesterol. The method incorporates stable isotope dilution using [3,4- C] cholesterol as an internal standard. [MM — H20] ions were monitored by the SIM method. Satisfactory agreement between the analytical result and the certified value of the National Institute of Standards and Technology standard reference material was obtained with a relative standard deviation of 0.6%. The method does not require sterol derivatization. Yang et al. (1992) used FAB/MS to identify cholesteryl sulphate m/z 465) as the [M — H] ion recovered from the appropriate TLC fraction. 

Lerma-Garcia, M. J., Simo-Alfonso, E. R, Mendez, A., Lhberia, J. L., and Herrero-Martinez, J. M. 2011. Classification of extra virgin olive oils according to their genetic variety using linear discriminant analysis of sterol profiles established by ultra-performance liquid chromatography with mass spectrometry detection. Food Res. Int. 44 103-108. 

Al-Ismail, Kh. M., Alsaed, A. K., Ahmad, R., and Al-Dabbas, M. 2010. Detection of olive oil adulteration with some plant ohs by GLC analysis of sterols using polar column. Food Chem. 121 1255-1259. 

Segura-Carretero, A., Carrasco-Pancorbo, A., Cortacero, S., Gori, A., Cerretani, L., and Femdndez-Gutifeez, A. 2008. A simphfied method for HPLC-MS analysis of sterols in vegetable oil. Eur J. Lipid Sci. Technol. 110 1142-1149. 

Hooper, S. N., and Chandler, R. F. Quantitative Gas-Liquid Chromatographic Analysis of Sterols in Biological Samples... 

TLC is often used as a preparative technique to isolate individual lipid fractions for subsequent studies by a variety of analytical procedures. Preparative isolation of the sterol (Chitwood et al., 1985 Shetty et al., 1990) and triacylglyc-erol (Horutz et al., 1993) fractions have been reported. Preparative isolation of sterols may allow these lipids to be analyzed further by argentation TLC (Ditullio et al., 1965 Morris, 1966) or by GLC (Shetty et al., 1990). Moreover, preparative isolation of triacylglycerols will allow for TLC analysis of glyceryl ethers (Snyder, 1971) and HPLC analysis of specific triacylglycerols (Horutz et al., 1993). Experiment 7 is concerned with a preparative procedure for the analysis of sterols by argentation TLC. 

Heftmann (1983) reviewed chromatographic techniques for the analysis of sterols, vitamin D, sapogenins and alkaloids, estrane derivatives, androstane derivatives, corticosteroids, miscellaneous hormones, bile acids, ecdysteroids, and lactone derivatives. 

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