Analytical methods nitrosamines

By using modem production methods it is possible to reduce the amounts of 1,4-dioxane to a level that is barely detectable with the best current analytical methods. Free ethylene oxide is now below detectable levels. Furthermore, volatile and nonvolatile nitrosamines ( NDELA ) both seem to be below detection limits of ppb in the alkanolamide-based sulfosuccinates. A good overview of modern analytical methods for the detection of 1,4-dioxane and ethylene oxide as well as nitrosamines and formaldehyde is given in Ref. 60. 

Reliable analytical methods are available for determination of many volatile nitrosamines at concentrations of 0.1 to 10 ppb in a variety of environmental and biological samples. Most methods employ distillation, extraction, an optional cleanup step, concentration, and final separation by gas chromatography (GC). Use of the highly specific Thermal Energy Analyzer (TEA) as a GC detector affords simplification of sample handling and cleanup without sacrifice of selectivity or sensitivity. Mass spectrometry (MS) is usually employed to confirm the identity of nitrosamines. Utilization of the mass spectrometer s capability to provide quantitative data affords additional confirmatory evidence and quantitative confirmation should be a required criterion of environmental sample analysis. Artifactual formation of nitrosamines continues to be a problem, especially at low levels (0.1 to 1 ppb), and precautions must be taken, such as addition of sulfamic acid or other nitrosation inhibitors. The efficacy of measures for prevention of artifactual nitrosamine formation should be evaluated in each type of sample examined. 

Radioisotope-labeled nitrosamines have proven valuable in development of analytical methods and for demonstrating efficiency of recovery of nitrosamines from tobacco products and smoke (37-39). The very high specific activity required for low part-per-billion determinations has discouraged most analysts from using this approach. Unless a radiochromatographic detector with adequate sensitivity is available, samples must be counted independently of the final chromatographic determination, and one of the advantages of internal standardization, correction for variation in volume injected, is lost. 

Low resolution MS yields specificity comparable to that of high resolution MS, if a relatively pure sample is delivered to the ion source. Either high resolution GC or additional sample purification is required. To obtain sufficient specificity, it is necessary to demonstrate that the intensities of the major peaks in the mass spectrum are in the correct proportions. Usually 10 to 50 ng of sample is required to establish identity unambiguously. Use of preparative GC for purification of nitrosamines detected by the TEA ( ) is readily adaptable to any nitrosamine present in a complex mixture and requires a minimum of analytical method development when new types of samples are examined. 

The prototype of the tobacco specific nitrosamines, NNN, has been detected in both tobacco smoke and unburned tobacco. Various analytical methods have been used including gas chromatography (GLC) (13,14,15,16) combined GLC-mass spectrometry (17), thin layer chromatography (18) high pressure liquid chromatography (HPLC) (19,20), and combined HPLC-thermal energy analysis (21). 

Aromatic A-nitrosamines on treatment with acid can undergo the well-known Fisher-Hepp rearrangement (32). For example, A-nitroso-A-methylaniline (NMA) on treatment with concentrated HC1 in ethanol rearranges to A-methyl-4-nitrosoaniline. It has been mentioned that A-nitrosamines are slowly denitrosated by treatment, especially on heating, with inorganic acids. Such denitrosation, however, proceeds smoothly and much faster in anhydrous medium, e.g., upon treatment with HBr in glacial acetic acid (35). Analysis of the resulting secondary amines or the nitrous acid provides the basis for several analytical methods to be discussed later. 

Pylypiw HM Jr. 1987. The development and application of analytical methods for the study of volatile and non-volatile N-nitrosamines (Pharmacokinetics). Disser Abstr Inter 48 1028-B. 

GC coupled to TEA detection is the most suitable analytical method to determine volatile nitrosamines. However, a large number of A/-nitrosamines are not generally adequate for direct analysis by GC, either because of low volatility or thermal instability. In these cases, HPLC seems to be the method of choice for the analysis of volatile and nonvolatile nitrosamines. The correct choice of stationary phase and type of composition of the mobile phase depends on the detection method used. 

A considerable number of CE separation methods exist for a wide variety of analytes. However, nitrosamines separation and determination by CE requires additional development for its practical use. " " For the separation of hydrophilic, low molecular weight, neutral, and polar compounds such as nitrosamines, it is necessary to develop CE techniques for enhancing the selectivity. The main reason is that these compounds do not interact strongly with the commonly used surfactants (e.g., sodium dodecyl sulfate, SDS) or other buffer modifiers such as cyclodextrins in electrokinetic chromatography. The separation depends on several factors which must be optimized to reach... 

The majority of the analytical methods for the detection of //-nitrosamines have employed GC or liquid chromatography in conjunction with a TEA. The disadvantage of these techniques, however, is that a subsequent confirmation is needed to ensure that the method does not give a false-positive response. 

N-Nitrosamines are widely distributed in various human environments. The concern was initially focused on their widespread occurrence in food and consumer products, as beer, meats cured with nitrite, smoked fish, tobacco and tobacco smoke, rubber products including baby bottle nipples and pacifiers, cosmetics, drug formulations, or herbicides formulations. Much data of their occurrence have been obtained by inadequate analytical methods and must await confirmation. Considerable progress has been made in the development of adequate and specific methods for trace analysis of nitrosamines, and reliable information is expected in the near future. 

Such mechanistic studies are important as they provide information crucial to development of water treatment technologies that minimize formation of nitrosamines while still providing safe disinfection of drinking water. Clearly still more needs to be done on this issue, and development of analytical methods for NDMA precursors and reaction intermediates will continue to play a key role. 

Botanical extracts such as lanolin. Aloe vera, Centella asiatica or glycyrrhetic acid are studied. Vitamins such as A, C, B5 and E have also been reported. Studies on the following compounds used as ingredients can also be found heterocyclic compounds, organic acids, thiocompounds, alkanolamines, amines, amino acids, alcohols, peroxides, amides, polymers, peptides and ceramides. Analytical methods to determine toxic compounds present as impurities or as by-products such as nitrosamines, phenols, pesticides, polychlorinated biphenyls, etc. have also been proposed in literature. 

A review of nitrosamines in cosmetics (Havery and Chou, 1994a) and others in sunscreens (Havery and Chou, 1994b) summarizes the most important aspects of analytical methods, occurrence, concentration levels and removal strategies. 

Beer Samples. The beer samples were examined as part of the American Society of Brewing Chemists (ASBC) and Association of Official Analytical Chemists (AOAC) collaborative studies of NDMA in beer. Duplicate samples were analyzed by the column extraction procedure and the ASBC distillation procedure (35). The AOAC procedure (36) was similar, except that a larger sample (50 vs. 25 g) was examined and sulfamic acid was added to minimize artifactual formation of nitrosamines. Both methods utilize N-nitrosodipropylamine (NDPA) as an internal standard. 

Available detection and confirmation methods are adequate for establishing identity and amounts of nitrosamines in most environmental and biological samples. The validity of analytical results, especially at levels in the part-per-billion and lower range, depends upon the experience and skill of the analyst in preventing or detecting contamination or artifactual formation of nitrosamines. 

U.S. EPA Method TO7 describes the determination of N-nitrosodimethylamine in ambient air (U.S. EPA, 1986). The method is similar to the NIOSH method discussed above and uses Thermosorb/N as adsorbent. The air flow is 2 L/min and the sample volume recommended is 300 L air. The analyte is desorbed with methylene chloride and determined by GC/MS or an alternate selective GC system, such as TEA, HECD, or thermoionic nitrogen-selective detector. The latter detector and the TEA are more sensitive and selective than the other detectors. Therefore, the interference from other substances is minimal. Other nitrosamines in air may be determined in the same way. 

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