Carcinogens, HPLC analysis

Figure 12.11—Comparison of UV andfluorescence detection after chromatographic separation. Aflatoxins, which are carcinogenic contaminants present in certain batches of grain cereals, are controlled by HPLC analysis. It can be seen that the peak intensities in UV detection vary with concentration whereas fluorescence detection is much more sensitive to aflatoxin G2 and B2. (Reproduced by permission of SUPELCO.)...

Analysis of Carcinogens. HPLC methods for the determination of polynuclear aromatic hydrocarbons benzidine and 3,3 -dichlorobenzidine and 4,4 -methylenebis(2-chloroaniline) (MOCA) are discussed. 

Final proof came after chemically detecting the actual carcinogen in milk from bracken-fed cows by HPLC analysis [114]. This point will be resumed later as it makes sense only after a deeper look into bracken bioactive chemistry in the following sections. Direct contact with bracken metabolites may result from other uses. For example, fresh bracken fronds are used in the northern Andes as wrapping of cheese to be smoked, in order to add flavor to the product. Some Wayuu Indians of the Perija Sierra in Western Venezuela bum the dry fronds with a limited air supply inside their huts to fend off mosquitoes. Some communities in Colombia use frond infusions as vermifugues. However, data is still missing to relate these applications to human cancer. But all the well... 

An HPLC technique has been developed [59] for volatile carcinogenic N-nitrosamines which shortens the chromatographic analysis time by a factor of five compared with GLC methods. The nitrosamines are converted into their amine reduction products which are then treated with 2,4-dinitrofluorobenzene (DNFB) in order to form the amines. The amines are then separated and detected at 340-360 nm. The reactions involved are shown in Fig.4.31. 

Air analysis may be performed by U.S.EPA Method TO 13 (U.S.EPA 1988), which is quite similar to the above method. PAH-bound particles and vapors (many compounds may partially volatilize after collection) may be trapped on a filter and adsorbent (XAD-2, Tenax, or polyurethane foam), and then desorbed with a solvent. The solvent extract is then concentrated and analyzed by HPLC (UV/Fluorescence detection), GC-FID, or GC/MS (preferably in SIM mode). Because of very low level of detection required for many carcinogenic PAHs, including benzo(a)pyrene, the method suggests the sampling of a very high volume of air (more than 300,000 L). 

Another species of interest in drinking water analysis is Cr (VI), considerably more toxic than Cr (III) and classified as carcinogenic to humans. Chromium speciation in water has been carried out by HPLC-ICP-MS, using both IEC and RPIPC (see Table 8.3). 

Figure 11.12 Comparison, following a chromatographic separation, of UV and fluorescence detection. Aflatoxins, which are carcinogenic contaminants present in certain hatches of grain cereals, are the subject of analysis by HPLC. Detection by fluorescence is much more sensitive to Gj and B2 than with UV detection (reproduced courtesy of SUPELCO). Below left, schematic of the different components of a LC-detector based upon fluorescence. This detector is able to find rapidely, for each compound eluted, the best coupling of excitation/emission without interrupting the chromatography underway (reproduced courtesy of a document from Agilent Technologies).

HPLC has also been applied to the analysis of trimethylolpropane and pentaerythritols in industrial synthesis solutions, 2-mercaptobenzothiazole in waste dump effluent, polythionates in mining waste water, isocyanates in working atmospheres, carcinogenic amines, aniline in waste water, polyethylene oxide fatty acid surfactants in industrial process waters and phthalate esters in river water. 

Since the discovery of the carcinogenicity of A-nitrosamines and their occurrence in the environment, many separation and detection methods have been developed for their analysis. Polarographic, spectrophotometric, and thin-layer chromatographic methods have been the earliest techniques employed and have been reviewed by several authors.However, the A-nitrosamines are now routinely analyzed by chromatographic procedures. Two major types of chromatography methods are commonly used, namely, GC and HPLC. Other techniques like capillary electrophoresis and capillary supercritical fluid chromatography have been introduced most recently. 

High-molecular-mass PAHs (302 amu or greater) are not normally detected by GC-MS owing to their low volatility. These groups of PAH isomers are present at very low levels yet they have been shown to be much more potent carcinogens. MS detection of liquid effluents allows for the detection of very high molecular mass PAHs such as coronene that are not sufficiently volatile for GC separation. The interface between the HPLC or CE column and the mass spectrometer represents a challenge for the analysis of neutral and nonpolar molecules. Also, the use of water as a mobile phase in reversed-phase LC is problematic due to its low volatility. Heated pneumatic... 

Hecht, S.S., J.D. Adams, and D. Hoffmann HPLC-TEA of tobacco-specific nitrosamines in Environmental carcinogens. Selected methods of analysis. Vol. 6.7V-Nitroso componnds, edited by H. Egan, R. Preussmann, G. Eisenbrand, T. Spiegelhalder, I.K. O Neill, and H. Bartsch, lARC, Lyon, Erance, lARC Sci. Publ. No. 45 (1983)429-436. 

---