N-Nitroso Compounds - American Chemical Society

Chemistry of Some N-Nitrosamides CLIVE Ν. BERRY, BRIAN C. CHALLIS, ANDREW D. GRIBBLE, and SUSAN P. JONES

Downloaded by FUDAN UNIV on November 19, 2016 | http://pubs.acs.org Publication Date: December 9, 1981 | doi: 10.1021/bk-1981-0174.ch007

Department of Chemistry, Imperial College, London, SW7 2AZ, England

Mechanisms for the thermal and photolytic decompos ition of N-nitrosamides are b r i e f l y reviewed, and recent results for t h e i r decomposition by acidic and basic catalysts are summarised and discussed. In the presence of acids, decomposition proceeds by concurrent pathways involving either deamination (hydrolysis) or denitrosation of conjugate acid intermediates, whose formation is usually rate l i m iting. Denitrosation predominates at high a c i d i t y and is negligible at pH> 2. Deamination, which generates diazohydroxide a l k y l a t i n g agents, i s therefore considered to be the more likely trans formation under stomach conditions. Decomposition in the presence of bases occurs readily at pH 2-12 by an addition-elimination pathway involving nucl eophilic rather than general base c a t a l y s i s . This reaction, which also generates a diazohydroxide a l k y l a t i n g agent, is considered to be the most important transformation under c e l l u l a r conditions. It can be induced by nucleotides and nucleic acids i n v i t r o , which are then alkylated to a small extent. There i s much evidence to suggest that carcinogenic N-nitrosamines are metabolised by an oxidative process to produce an alkylating agent (_l 2). One potential metabolite i s therefore the corresponding N-nitrosamide resulting from 2-electron oxidation at the eç-carbon atom, and, indeed, such compounds appear to induce tumours at the s i t e of application without metabolic activation (3)· I t follows that the chemical properties of N-nitrosamides are relevant to the etiology of cancer. N-Nitrosamides are much less stable than the parent N-nitrosamines and they can decompose by either thermal, photolytic or acid and base (nucleophilic) catalysed pathways. Thermal decomp o s i t i o n has attracted much attention as a clean method of deaminf

0097-6156/81/0174-0101$05.00/0 © 1 9 8 1 American Chemical Society Scanlan and Tannenbaum; N-Nitroso Compounds ACS Symposium Series; American Chemical Society: Washington, DC, 1981.

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N-NITROSO COMPOUNDS

ation. A good deal i s known about the mechanism of t h i s reaction and the findings have been c r i t i c a l l y reviewed (^ 5)» Thermal decomposition proceeds readily at temperatures from 25-100 C , and involves i n i t i a l rearrangement to a diazoester intermediate I which then rapidly expels nitrogen to give various products as i n f

f


^R^^O-NsN-R» j

^>RC0 R* R(X> H, olefins 2

f

2

(1)

equation 1. Both the ease of rearrangement (3 > 2 > 1 ) and the type of product (formed by carbonium ion or free r a d i c a l substit utions and eliminations) are more dependent on the nature of the R substituent than R. The diazoester intermediate I i s , of course, a putative a l k y l a t i n g agent, but R' invariably reacts intramolecularly with the carboxylate ion on the extrusion of Photolytic decomposition has also been well-investigated, p r i n c i p a l l y by Chow and h i s colleagues ( 6 ) · These reactions proceed on i r r a d i a t i o n at various wavelengths i n both polar and non-polar solvents. The primary process following photoexcitation i s d i s s ociation to a r a d i c a l p a i r I I , and ensuing chemical events involve the amidyl and n i t r i c oxide radicals either as a caged pair or i n 1

0

hv.

R-^-NR» NO*

0

NH(CH ) CH(NO)R 2

5

or

III (2)

II i^^N=CHR +

(NOH)

IV the bulk of the solution (equation 2 ) · The amidyl radicals comm only undergo intramolecular Η-abstraction followed by coupling with the n i t r i c oxide to give a 6 -nitroso product I I I , o r ^ - e l i m ination to give an N-acylimine IV· In poor Η-donor solvents, and for i r r a d i a t i o n at 280 nm, more complex reactions occur for reasons discussed by Chow (6) i n h i s review. Neither thermal nor photolytic decomposition is""likely to be involved i n b i o l o g i c a l transformations of N-nitrosamides, but a different conclusion may apply to the acid and base catalysed decompositions. Decomposition by Acid Catalysts On treatment with gaseous HBr i n organic solvents, N-nitros amides regenerate the parent amide (equation 3) i n quantitative y i e l d ( 7 8 . ) . This implies that N-nitrosamide formation i s revers i b l e and explains why i t i s b e n e f i c i a l to add one mole of base f

Scanlan and Tannenbaum; N-Nitroso Compounds ACS Symposium Series; American Chemical Society: Washington, DC, 1981.

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(usually NaOAc) when synthesizing N-nitrosamides from amides and n i t r o s y l gases ( 7 ) · Denitrosation~should also occur i n aqueous acid together with hydrolysis reactions characteristic of regular amides. These expectations are borne out by recent studies (9«10)

R ^ ^ l /


N

+ HBr

R-*^NHR

t

+ NOBr

(3)

N 6

which show that t y p i c a l N-nitrosamides undergo concurrent denitro sation and deamination i n aqueous HpSO^ and HCIO^ (Scheme 1 ) . The extent of each reaction depends on Doth the structure of the

> w R»

+

H 0 (fast)^

RC0 H 2

+ R'-NsN-OH

slow VII R*0H + N

2

+

H 0 (slow)^

Λ

R-^^r H R' X

RCONHR' + HNO

p

fast

VI Scheme 1. Concurrent acid catalysed denitrosation and deamination (hydrolysis) of N-nitrosamides N-nitrosamide and the solvent a c i d i t y , but generally denitrosation i s more strongly acid-catalysed: hence, i t i s a minor reaction ( RCO" + R»CH N=N-0H ?

f

R CH=N=N + H 0 2

(4)

interest to establish both the mechanism of these reactions and t h e i r propensity under c e l l u l a r conditions. We have shown (10) that base (dr nucleophilic) catalysed hydrolysis contributes to the decomposition of N-nitroso-2-pyrrolidone at 25°C i n C l HCCO Η and C1H CCO Η buffers at pH 1-3, and


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N-NITROSO COMPOUNDS

i s the sole decomposition pathway i n HCO H and CH_C0pH buffers at pH 3 · 7 - 5 Λ · Evidence to t h i s effect for CH C0 H buffers i s the correlation of pseudo f i r s t order rate coefficients (Rate = k [Substrate]) with [CH CO"] but not [CH_CO H] shown i n Figure"?, and the absence of HNOp as a reaction proauct» We have also shown (18) that other bases stronger than CH_C0! (pK. £·75) catalyse the decomposition of N-nitroso-2-pyrrolidone at 25 C. With the exception of imidazole, these reactions follow uncomplicated second order kinetics (Rate = kp[Substrate][Base]) and only products of deamination (hydrolysisT are obtained. Gen e r a l l y , kp values increase with the base strength of the catalyst and f i t tne Br/rfnsted relationship withj|=: 0.66· However, the ab sence of s i g n i f i c a n t catalysis by s t e r i c a l l y hindered bases (eg. 2,6-lutidine), the strong catalysis by imidazole r e l a t i v e to HP07 (k (Imidazole)/k (HP0p = 83) and by hydroxide ion r e l a t i v e to p

2


1

2

2

imidazole (k (H0~)/kp(Imidazole) = 4,300), and the observation of second order c a t a l y t i c terms j n imidazole (Rate = [Substrate] (kp[Imidazole] + k_[Imidazole] ) suggests that decomposition by some ( i f not a l l ) catalysts involves nucleophilic attack at the carbonyl C-atom as shown i n Scheme 2· 2

Scheme 2· Imidazole catalysed decomposition of N-nitroso-2pyrrolidone The tendency for N-nitrosamides to undergo hydrolysis by a nucleophilic catalysed*"pathway has been confirmed by studies of N-alkylnitroso acetamides ( 1 9 ) · Results summarised i n Table I for N-n-butyl-N-nitroso acetamide show that i t s decomposition i s also subject to s t e r i c constraints (2,6-lutidine^pyridine) and i s best effected by strong nucleophiles (eg, imidazole, t h i o l s ) irrespect ive of t h e i r base strength (pK.)· Further, the second order dep endence on [Imidazole] i s more clearly defined for the decomposit-


BERRY ET AL.

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107


7.

Figure 2.

Variation in the rate of deamination of N-n-butyl-N-nitrosoacetamide with [CH CO H] and [CH COf] at 25°C. s

g

s


108

ΛΓ-NITROSO COMPOUNDS

Table I Rate Coefficients for the Decomposition of N-n-Butyl-N-nitrosoacetamide at 25°C. — Ç —

Catalyst


C H

1θ\

HOOT CH S HO" Cysteine thiolate Glycine Methionine Lysine Arginine d

d

s 0.114 0.14

4.75 5.17 6.77 6.95 7.05 7.21 9.72 15.75 1.8/8.3/10.8 2.34/9.60 2.12/9.28 2.16/9.18/10.79 1.82/8.99/12.48

C 0

3 2 Pyridine 2,6-Lutidine Ν-Methylimidazole Imidazole

-1 -1

No catalysis 15.7 39.7 1.35

438 000 378,000 34,400 0.14 No catalysis f

2.75 1.17

ion of N-n-butyl-N-nitroso acetamide than N-nitroso-2-pyrrolidone and i s found to be pH dependent (Figure 3 ) . The imidazole cata lysed reaction follows a complex rate expression (equation 5) consistent with the detailed mechanism of Scheme 3 · The occurrence of second order [Imidazole] terms relates to the existence of two tetrahedral intermediates (probably IX and X) i n prototropic equi librium. At pH c a . 6 , intermediate IX which can decompose spontan eously to products predominates, and second order [Imidazole] terms are not observed. At pH 7 however, reaction v i a intermed iate X becomes increasingly important. Since i t s decomposition to f

Rate

=

k.[Im] { k - + Κ· k'[Im] + Κ k-[H0"]} [Substrate] — ^ ILZi HLZ£ J— k + k^ + K klXlm] + Κ k [H0"] —*-1 —3 X ""2 X —2

(5)

1

Λ

where ^

= k· + kJjCHO"] and

o

+

= Κ[Ηθ"][ΙιηΗ]/[Ιιη] χ

products i s f a c i l i t a t e d by general acid catalysts (including the imidazolium i o n ) , second order [Imidazole] terms are observed. This mechanistic interpretation i s supported by the i s o l a t i o n of N-acetylimidazole as a major product and the observation of only f i r s t order catalysis by N-methylimidazole irrespective of the pH (19).

Thus, N-nitrosamides also undergo deamination (hydrolysis) by an addition-elimination pathway involving nucleophilic rather than


Chemistry

of

N-Nitrosoamides


BERRY ET AL.

[IMIDAZOLE]

Figure 3.

M

Imidazole catalyzed deamination of N-n-butyl-N-nitrosoacetamide 25°C.


at

iV-NITROSO COMPOUNDS


110


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the general base catalysis usually observed for regular amides. This difference can be related to the enhanced leaving a b i l i t y of the N-nitrosamino fragment. The reaction proceeds over a wide range of pH (ca.2-12) and i s probably the most important decompos i t i o n pathway under c e l l u l a r conditions. S i g n i f i c a n t l y , i t also generates diazohydroxide a l k y l a t i n g agents such as VIII and X I .


Decomposition by Nucleotides and Nucleic Acids One goal of our current investigation i s to show that genet i c a l l y important c e l l u l a r constituents can i n i t i a t e the release of an a l k y l a t i n g agent from the N-nitrosamide. This might explain how highly reactive diazohydroxide metabolites effect the a l k y l a t ion of nucleic acids within the c e l l nucleus. Preliminary evid ence (20) summarised i n Table I I shows that the decomposition of Table I I Rate Coefficients for the Decomposition of N-n-Butyl-N-nitroso acetamide by Nucleotide-5'-monophosphates i n 0.1M Phosphate Buffers at 25°C.

Catalyst HP0= Guanosine Adenosine Cytidine Ribose 2 -Deoxyguanosine 2 -Deoxyadenosine 2 -Deoxycytidine 2 -Deoxyribose 1 1 1

1

10 k 5

pH

7.23 7.26 7.34 7.31 7.10 7.09 7.08 7.0

1

o

M" s"

1

51 60 38.5 35Λ

14.5 18 13Λ

14.5

N-n-butyl-N-nitroso acetamide i n phosphate buffer at 25°C. i s catalysed by nucleotide components. Surprisingly, the strongest catalysis applies to guanosine- and adenosine-5'-monophosphates, but i t should be noted that a l l the k- values cited i n Table I I are pH dependent. The decomposition of N-n-butyl-N-nitroso acet amide i n phosphate buffer at 25°C. i s also catalysed by c a l f - l i v e r RNA (Figure 4) to an extent comparable with the c a t a l y t i c rate co e f f i c i e n t s given i n Table I I ( 2 0 ) . Examination of the products from reaction of guanosine- and adenosine-5 -monophosphates with radiolabelled N-nitroso acetamides i n phosphate buffer shows lowl e v e l (

N-Nitroso Compounds - American Chemical Society

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