Denitrosation nitrosamines

Ross and Chiarello (1979) have described a method for denitrosation of nitrosamines, such as dlalkylnitrosamines e.g. N-nitrosodimetl laminer etc., and complex aryl-alkyl nitrosamines, e.g. N-nitroso-N-(l-ethylpropyl)-3,4-dimethyl-2,6-dinitroaniline. The products of nitration and amination containing nitrosamlne as a substantial impurity are treated with an aldehyde or ketone in the presence of strong acids such as hydrochloric or hydrobromic acid. Under pressurized conditions at 105-110% for cme to two hours, the nitrosamlne is destroyed. The desired product can be recovered after neutralization of the excess acid. The equation shows the denitrosation of pendimethalin ... 

Type II nitrosamines have two reaction pathways. One pathway involves nucleophilic attack at the carbon of C=0 to generate a tetrahedral intermediate which decomposes to an active diazotate ion (R-N=N-0 ). The other pathway involves the nucleophililc attack on the nitrogen of the nitroso group resulting in denitrosation (Scheme 3.5). The nucleophile can be a biologically prevalent thiol, therefore type II compounds are often used as NO donors for the formation of S-nitrosothiols [67, 68]. 

When thiocyanate ions are added to nitrous acid in water, a pink colouration develops which is believed to be due to the formation of nitrosyl thiocyanate (equation 34), which is too unstable to be isolated but which can be used as a nitrosating agent in aqueous solution. Because the equilibrium constant for ONSCN formation81 is quite large (30 dm6mol 2) at 25 °C, thiocyanate ion is an excellent catalyst for aqueous electrophilic nitrosation. The well established82 series is Cl- < Br < SCN < (NH2)2CS. Thiocyanate ion is also a sufficiently powerful nucleophile to react in acid solution with nitrosamines in a denitrosation process (equation 35), which can only be driven to the right if the nitrosyl thiocyanate is removed by, e.g., reaction with a nitrite trap such as hydrazoic acid. 

A method involving SPE was developed for the determination of ten A-nitroso amino acids in cured meat products. These compounds were derivatized with diazomethane followed by O-acylation of hydroxyl groups with acetic anhydride-pyridine reagent. The methyl esters and their acylated derivatives were separated by GC on a DB-5 fused silica capillary column and quantified with a TEA-CLD specific for the nitric oxide derived from the thermal denitrosation of nitrosamines recovery exceeded 75% at the 10 ppb level579. 

Reactions of the Nitroso Group. Up until recently, the chemistry of N-nitrosamines was restricted to the reaction involving the nitroso group. It was not until the usefulness of N-nitros-amines in synthesis [concept of "Umpolung (3.)] was demonstrated, that the long-known denitrosation of N-nitrosamines became impor-... 

There has been some controversy regarding the site of attack of the proton on the basis of the dipolar structures I and I", it might be inferred that the proton attack should occur on the oxygen atom the fact that N-nitrosamines form 0-complexes and 0-alkylated products would seem to support this view (iT) However, it is currently believed that the N-protonated form is the one that leads to denitrosation, irrespective of whether or not initial protonation occurs on oxygen (18). 

One of the oldest-known reactions of N-nitrosamines is their reduction to 1,1-disubstituted hydrazines discovered by Fischer (19) The most common method to perform this transformation has been zinc dust in acid, generally acetic acid tetrazenes are sometimes formed as by-products (20) and denitrosation can also occur. Several other reducing methods have been investigated reduction with lithium aluminum hydride and catalytic hydrogenation are sometimes useful. Sodium dithionite reduction of benzyl substituted N-nitrosamines in base can result in fragmentation to... 

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. 

The protonation of the nitrosamine is written as N-protonation of the amino-nitrogen atom. This has not been established beyond all doubt, but makes for a reasonable mechanism for the denitrosation process, which must also represent the reverse sequence of steps for nitrosation. 

It appears that the NO+ group can be transferred directly only to those species which can behave as reasonably powerful nucleophiles in moderately acid solution. This is undoubtedly the reason why species such as azide ion and amines do not appear to act in this way since they are very extensively protonated in these solutions. Azide ion catalysis has been reported in one case, however (Meyer and Williams, to be published) the denitrosation of N-acetyl-N-nitrosotryptophan, which reacts very rapidly, has been examined at pH 6 where a substantial quantity of free azide ion exists. Presumably the same would be true for some amines at these very low acidities. However, there is experimental evidence from two independent groups which shows that a direct (sometimes called trans-) nitrosation by nitrosamines can occur with aromatic amines. The reactions of N-nitrosodiphenylamine in aqueous acid solution were studied in this context (Challis and Osborne, 1973). The reaction was acid-catalysed for all the substrates studied and was believed to involve the protonated form of the nitrosamine (here written as PhjNNO, H" ). Two separate pathways were detected (a) a direct reaction with N-methylaniline (without the formation of free nitrous acid or a derivative of nitrous acid) as outlined in Scheme 20 and b) an indirect reaction with... 

One interesting feature of denitrosation of nitrosamines is that at high concentrations of the most powerful nucleophiles, the rate of reaction be-... 

Absorption, Distribution, Metabolism, Excretion. Examination of Section 2.6 clearly indicates that oral administration of NDMA has been the preferred route for studying its absorption, distribution, metabolism and excretion. This is not surprising since oral administration is easier to monitor when compared to other routes. The oral route seems to be the most pertinent to study since humans are most likely to be exposed to nitrosamines orally. Toxicokinetic data with regard to dermal and inhalation exposure of NDMA are clearly lacking. Furthermore, dermal and inhalation exposures may lead to different metabolic pathways and patterns of distribution and excretion, which could account for differences in the degree of toxicity exhibited by different routes of exposure. The metabolism of NDMA in isolated microsomal preparations seems to be well understood, but studies with cultured human cells could provide additional useful information. However, exploration of the denitrosation mechanism as an alternative to a-hydroxylation requires more attention. Determination of the urinary excretion of NDMA in control human volunteers and in individuals known to consume foods with high contents of nitrosamines could provide information concerning absorption and excretion of the xenobiotic. 

A-Nitrosamines can be accurately and precisely determined by GC-EPD as their A-diethylthiophosphoryl derivatives after denitrosation. This method is selective and sensitive, allowing cigarette smoke samples to be analyzed directly without pretreatment, except for separation from secondary amines by solvent extraction, and without any interference from other coexisting substances (Eigure 11.9). 

V-Nitrosamines are extremely reactive. They are sensitive to prolonged thermal treatment as well as to photochemical irradiation. Most Af-nitrosamines undergo reactions with inorganic acids, such as HCl, HBr, and HI. In fact, this process is the basis for the denitrosation of such compounds. 

SFE has been demonstrated to be a good extraction technique for A-nitrosamines in rubber products. In addition, SFE allows fast analysis with a reduction in solvent waste, time, and manipulation. Although recoveries are not too good, especially for the smaller A-nitrosamines, SFE could be considered as a useful tool to determine these analytes, considering that through its selectivity it provides quite clean extracts in one step." Reche et al." determined A-nitrosamines in latex products by combining supercritical fluids and chemical derivatization. The addition of a denitrosation reagent into the extractor combined with an adequate liquid trap allows elucidation of the presence of A-nitrosamines as well as their potential precursors. 

Reversed-phase chromatography of A/-nitrosamines has been used in connection with UV, fluorimetric, amperometric, and MS detection. In many cases these methods are based on pre- or postcolumn denitrosation, and derivatization of the denitrosation products. Chemically bonded octadecylsilane, Cig, is the most often reported stationary HPLC phase. Isocratic or gradient elution is usually performed with mixtures of water with acetonitrile, methanol, ethanol, or propanol. In general, the mobile phase is acidified by the addition of acetic acid, ammonium acetate, or phosphate buffers. The HPLC conditions used for the separation and detection of A/-nitrosamines are shown in Table 12.3. 

The most sensitive and selective methods for the detection of nitrosamines are based on their denitrosation followed either by a chemrlummescent determination of the nitric oxide (NO) released or by the determination of the resulting nitrite ion or its amino counterpart, with various techniques (spectrophotometry, fluorescence, chemfluminescence, and amperometry). 

The denitrosation of A-nitrosamines can be achieved by different procedures including the following ... 

The nitrite ion can be detected spectrophotometrically after the separation of nitrosamines by HPLC followed by photolytic or chemical denitrosation. Postcolumn formation of an azo dye by the reaction of nitrite with a Griess-type reagent allows its spectrophotometric detection at 546 The kinetics and mechanisms of the Griess reaction have been extensively... 

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