Fat samples

Figure 8.9 is an example of the use of solvent extraction to isolate polychlorinated biphenyls from a fat sample [6]. In this example the matrix is chemically modified to improve the selectivity of the extraction. The fat is first hydrolyzed by refluxing in 1 N ethanolic potassium hydroi prior to the... 

Figure 8.9 Solvent partition schene for the isolation of polychlorinated biphenyls from fat samples.

Extracts of these fat samples were treated with sodium sulfate-concentrated sulfuric acid mixture and fuming acid by the method described by Schechter et al. 5) in order to separate the organic-chlorine compound from the fatty materials. An infrared spectrum from 7 to 15 microns on carbon disulfide solutions of the residues from the fat qualitatively identified the organic-chlorine compound as toxaphene. All the bands of toxaphene in this spectral region were plainly seen in the treated steer extract, whereas none of the absorption bands were visible in the untreated steer extract. 

Few methods for measuring chlordecone in foods are available. Lower recoveries (58-81%) were obtained with GC/ECD for beef, pork, and poultry fat samples using GPC clean-up before analysis (Goodspeed and Chestnut 1991). Precision varied greatly (7.1-47.7% RSD) because of the lower recoveries sensitivity was not reported. 

Pesticide residues were analyzed in 183 milk samples from 165 Finnish women. Heptachlor was found in 12% of the samples heptachlor epoxide was found in 6.6%. Five percent of the samples contained levels of heptachlor epoxide in excess of 0.0005 mg/kg body weight, an acceptable daily intake (Mussalo-Rauhamaa et al. 1988). Fifteen milk and fat specimens from residents of Grand Forks, British Columbia, and 16 milk and 17 fat specimens from residents of Prince George, British Columbia, were analyzed for pesticide residues. Heptachlor epoxide was found in one milk sample and nine fat samples in the Grand Forks group (>0.004 ppm) and in no milk samples and two fat samples in the Prince George group (>0.004 ppm) (Larsen et al. 1971). The residue was not detectable at levels lower than 0.004 ppm because of limitations of the analytical methods and faulty techniques. It is possible that the potential exposure of the residents to heptachlor may also have occurred via food contaminated with heptachlor. 

PCBs are more difficult to extract from full-fat milk than skim milk, suggesting that PCBs are more strongly retained in the full-fat sample matrix. Even at 100 °C, SPME (without saponification) is not an efficient... 

In a residue depletion study in cattle given the normal therapeutic treatment, residues of danofloxacin in liver declined from 372 ppb at 12 h after the last dose to 13 ppb at 5 days after the last dose. Over the same time period, residues at the injection site, kidney, and muscle declined from 669 ppb to less than 10 ppb, from 426 ppb to 5 ppb, and from 112 ppb to less than 10 ppb, respectively. Residues in most fat samples were below 10 ppb. 

Liquid samples such as milk do not normally require application of any pretreatment procedure. Semisolid samples such as muscle, liver, and fat tissues usually require more intensive sample pretreatment for tissue break-up. The most popular approach is grinding the sample in a food chopper or homogenization in a Waring blender to expose residues to the extraction solvent. Fatty tissue samples are usually warmed at 35 C until fat melts (491-493), or sometimes blended with immersion blender (494). A fat sample that has been blended with immersion blender melts to produce yellow oil, whereas oil does not separate... 

Plant oils and animal fat samples (vegetable oils, tallow, lard, fish oils) do not need to be extracted before methylation. A small amount of these samples (10 to 20 mg) can be subjected to the methylation procedures directly. 

Crude oil or fat sample to be analyzed Ascorbic acid (ACS grade)... 

As the levels of tocopherols and tocotrienols in meat samples are usually lower than in oil and fat samples, a larger sample size is needed in the sample preparation. The meat sample is homogenized to weaken the sample matrix. As in Basic Protocol 2, saponification, heating, and liquid/liquid extraction are used to increase the recovery and remove interference compounds. Satisfactory results can be achieved using a reversed-phase HPLC method. 

To prepare fat sample for calibration, melt the calibration sample (if it is not liquid) by heating slightly (<10°C) above its melting point. If the food sample consists of large pieces, grind it coarsely. 

The OSI was primarily developed for oil or fat samples. It is possible to use the method on lipids extracted from foodstuffs however, such lipids may not have the same oxidative stability as the food since the physical structures are different. Additionally, muscle foods oxidize rapidly at elevated temperature due to physical and chemical alterations during cooking thus, this method probably will not be useful with raw meats. 

Melt fat samples in an oven at 100° 2°C until the fats are clear ( 5 min) with no suspended solids. If necessary, pass the fat through a paper filter and then melt again. Mix thoroughly. 

Transfer fat samples to glass NMR tubes, filling the tubes to a height of 30 to 50 mm. Ensure that no fat adheres to the outside of the tubes. 

Melt fat samples as described (see Basic Protocol, step 3). 

The fats used to formulate products such as margarine or shortening do not crystallize at 60°C, but if a fat sample does crystallize at this temperature, a higher temperature can be used. It is better to use as low a temperature as possible to avoid oxidation. 

Saponification causes a significant loss of xanthophylls, even when carried out under relatively mild conditions (ambient temperature for 3 h) (21). In addition, several different saponification procedures have been shown to promote the formation of cis isomers of /3-carotene (74). Since saponification prolongs the analysis and is error prone, it should be carried out only when needed, as in high-fat samples or those containing carotenol esters. 

Digest starchy samples with taka-diastase before saponification. Saponify (hot), extract un-saponifiables with petroleum ether. Silica solid-phase extraction in the sample cleanup mode (high-fat samples only). 

The most suitable solvents for extracting SPA from fats are acetonitrile (113,125,126, 139,142) and water-alcohol mixtures. The fat is usually dissolved in hexane or petroleum ether, and SPA are extracted into acetonitrile (105,110-113,125,126,128). The disadvantages of acetonitrile extraction are that (a) BHT recovery is low and (b) moderately high levels of interfering compounds are coextracted. The advantage of aqueous methanolic extraction of SPA from nonpolar solvent is that the fat is mostly excluded (99,114). Hammond (99) described a methanolic extraction of a melted fat sample, heated to 40-50°C, followed by transfer of the mixture to a deep freeze for a few hours to aid the solidification of any excess fat from methanol. The methanol layer was then decanted and filtered prior to the addition of an internal standard and direct injection. 

Gel permeation chromatography (GPC), also called size-exclusion chromatography, is the most widely used cleanup technique for pesticides in fatty foods. It is the method of choice for rapid cleanup of biological extracts, especially from high-fat samples, to determine pesticide residues, since separation occurs on the basis of molecular size (7). 

Following the succesful development of the cells, a rapid clean-up procedure for fat samples was developed, based on the use of an acid silica column. Using... 

Table 5.1 Reproducibility of the CALUX assay with milk fat samples. Spiked samples were tested singly in three independent test series (adapted from 12)...

In a MAFF study (MAFF, 1995) of pesticides in human fat in the UK, 99% of samples analysed (n = 203) had detectable residues of DDT (as p,p -DDT, o,p-DDT, p,p -TDE and/or p,p -DDE). Since the fat samples were taken at routine autopsy, it is likely that most of the subjects were at least 70 years old and therefore had lived through times when DDT was permitted in the UK. Their residues reflect their lifetime exposure. Twenty-three per cent of the subjects had DDT fat residues between 1 and 9.3 mg.kg 1 which suggests higher exposures. These people might have been exposed directly during DDT s heyday, or might have been fond of oily fish which contains higher residues of OCs than most other foods. 

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