Hydrogen-deuterium exchange of aromatic amines and amides using deuterated trifluoroacetic acid

Abstract

The H-D exchange of aromatic amines and amides, including pharmaceutically relevant compounds such as acetaminophen and diclofenac, was investigated using CF₃COOD as both the sole reaction solvent and source of deuterium label. The described method is amenable to efficient deuterium incorporation for a wide variety of substrates possessing both electron-donating and electron-withdrawing substituents. Best results were seen with less basic anilines and highly activated acetanilides, reflecting the likelihood of different mechanistic pathways.

Introduction

Increasing demand for deuterated compounds, especially due to a renewed interest in the therapeutic prospects of so-called “heavy drugs”,¹ has spurred development of novel methods for hydrogen-deuterium exchange. One group of compounds with significant research interest for H-D exchange is the aromatic amines and amides, due to the presence of these moieties in numerous clinically important pharmaceuticals, including such mainstays as acetaminophen (paracetamol), diclofenac, and acebutolol. The former two compounds are of particular importance with respect to aromatic isotopic exchange due to their metabolism to potentially toxic ring-oxygenated metabolites in the liver by cyctochrome P450 and related enzymes. Several in vitro and in vivo studies performed using deuterated analogs of these compounds have had profound mechanistic implications.² As a result, various syntheses of ring-deuterated variants of acetaminophen³ and diclofenac⁴ have been reported. However, these methods generally require several synthetic steps from expensive deuterated starting materials. Efforts have been made towards exploring the direct aromatic H-D exchange of both of these compounds to varying degrees of success.⁵ In particular, a rapid, direct, and metal-free method for labeling acetaminophen with deuterium has proven elusive.

Acid-catalyzed H-D exchange conditions encompass some of the most powerful and efficient methods for incorporating deuterium into an aromatic ring.⁶ While these methods can employ a wide variety of acidic catalysts, including heterogeneous and Lewis acids, homogeneous Brønsted systems are the most common. Under these conditions, reactivity typically proceeds through an electrophilic aromatic substitution mechanism and the aromatic ring is selectively deuterated at the most electron-rich positions. Commonly-used systems for H-D exchange include a solvent, usually D₂O, as well as a deuterated mineral acid catalyst such as DCl, DBr, or D₂SO₄. However, the use of these harsh reagents has several disadvantages, including unwanted side reactions, low substrate solubility in aqueous solution, and potential substrate decomposition.

The use of deuterated trifluoroacetic acid (CF₃COOD) has several benefits over other acidic H-D exchange methods, including its ease of preparation, simple removal in vacuo, low nucleophilicity, and high solubility properties for a wide variety of substrates. CF₃COOD can be prepared quantitatively from trifluoroacetic anhydride and D₂O in essentially anhydrous fashion. Since the pioneering efforts of Lauer et al.,⁷ CF₃COOD has found use as a highly versatile and effective deuterating agent for hormones,⁸ natural products,⁹ and various other biologically relevant aromatic systems.¹⁰ We considered the viability of this reagent for the deuteration of aromatic amines and amides such as acetaminophen. A handful of examples can be found in the chemical literature, including the CF₃COOD-catalyzed H-D exchange of tryptophan and its derivatives¹¹ along with a single report each regarding H-D exchange of naphthalene-diamines,¹² reserpine¹³ and pteroylglutamic acid.¹⁴ However, to the best of our knowledge, the H-D exchange of other aromatic amines and amides in CF₃COOD has not been studied extensively. Herein we report on the generalized use of CF₃COOD as a source of deuterium and reaction solvent for the preparation of several deuterated aromatic amine and amide derivatives.

Results and Discussion

While investigating palladium-catalyzed H-D exchange of pharmaceutically relevant compounds, we found CF₃COOD be highly effective at deuterating acetaminophen absent any metal catalyst at 110° C (Scheme 1). Since efficient and direct H-D exchange of this compound has only been conducted using the assistance of rhodium salts^5c we decided to investigate this unexpected transformation further. At reflux without any additional catalyst or co-solvent, acetaminophen was deuterated extensively at the aromatic positions ortho to the hydroxyl substituent and more slowly at the positions ortho to the amide. The selectivity of deuteration is opposite that observed in the previously reported metal-catalyzed example and is unprecedented in the direct H-D exchange of acetaminophen.

Scheme 1.

CF₃COOD-catalyzed H-D exchange of acetaminophen.¹⁵

Encouraged by these results, we endeavored to determine the scope of this methodology¹⁶ and subjected simple aniline and acetanilide, as well as aniline derivatives substituted only at nitrogen, to the same CF₃COOD conditions used for acetaminophen (Figure 1). Aniline 1 and N-ethylaniline 2 underwent H-D exchange smoothly under these conditions at the positions ortho and para to the nitrogen atom, a result consistent with a standard EAS mechanism. However, the H-D exchange of N,N-diethylaniline 3 under the same conditions was poor despite the enhanced induction from the alkyl substituents on nitrogen. This low rate of exchange was also observed in previous work on the acid-catalyzed proton exchange of N,N-diethylaniline¹⁷ and attributed to its high basicity and at least partially to steric interference from the ethyl groups. Interestingly, 1-phenylpiperazine 4 was even more reactive towards CF₃COOD–catalyzed H-D exchange than aniline, indicating that dialkylation of the amine is not inherently detrimental towards the exchange process. Acetanilide 5 was similarly reactive towards H-D exchange as aniline, suggesting that acetylation of the nitrogen does not necessarily deactivate the aromatic ring towards reaction with CF₃COOD.

Figure 1.

H-D exchange of anilines and acetanilide without ring substituents.¹⁵

To test the effect of various ring substituents, para-substituted substrates were then investigated under the reaction conditions (Figure 2). The reactivity of p-toluidine 6 was similar but not enhanced compared to aniline, despite the presence of an activating methyl substituent on the ring. However, the reactivity of both p-anisidine 7 and p-aminophenol 8 was significantly lower than that of p-toluidine, and deuteration of both of these substrates proceeded with very low selectivity. Interestingly, 4-nitroaniline 9 underwent H-D exchange analogously to p-toluidine despite the very different electronic properties of these aromatic systems. Thus a strong correlation between the basicity of the amine and the rate of H-D exchange can be seen, with more basic anilines reacting somewhat less efficiently and with lower selectivity. Acid-catalyzed H-D exchange of aromatic anilines can proceed via aromatic substitution of either the protonated anilinium ion (slow) or its free-base counterpart (fast) which exists in equilibrium. In strongly acidic solution, electron-rich anilines are more likely to react through the former pathway, and with deactivated anilines, the latter pathway predominates.¹⁸ The observed selectivity differences in this experiment likely reflect these mechanistic differences.

Figure 2.

H-D exchange of para-substituted anilines and acetanilides.¹⁵

Further support of this hypothesis can be drawn from the reactivity of the N-acetylated derivatives of each compound. Acetylation of the amine inhibits protonation at nitrogen, essentially eliminating one of the two possible H-D exchange pathways from consideration. The reactivity of 4′-methylacetanilide 10 was comparable to its non-acetylated counterpart 6, reflecting a minor inductive effect from the methyl substituent. In contrast, the H-D exchange of 4-methoxyacetanilide 11 was very efficient and highly selective compared to p-anisidine 7, and the observed H-D exchange pattern was opposite that of 4′-methylacetanilide. The reactivity and deuteration selectivity of acetaminophen 12 was analogous to that of 4-methoxyacetanilide and similarly improved relative to 4-aminophenol 8. The reactivity of 11 and 12 revealed the strong directing influence of the hydroxy and methoxy functional groups in the absence of protonation at nitrogen. However, the more-deactivated 4-nitroacetanilide 13 barely reacted at all compared to 4-nitroaniline 9. Thus, while optimal H-D exchange results occur with less electron-rich anilines, acetanilides react with greater efficiency with an activating substituent on the aromatic ring.

We then considered the effect of the substitution pattern of the ring on H-D exchange by reacting the other anisidine isomers with CF₃COOD (Figure 3). The influence of the amine group dominated the H-D exchange selectivity of o-anisidine 14, and the positions ortho and para to the amine exchanged rapidly under these conditions while the positions ortho and para to the methoxy group had significantly lower deuterium incorporation. A similar pattern was observed in m-anisidine 15, which underwent rapid exchange throughout the ring except the single position meta to both the amine and methoxy substituents. The effect of an electron-withdrawing substituent nitro substituent in place of the methoxy was pronounced, as 3-nitroaniline 16 exchanged at a much lower rate than m-anisidine.

Figure 3.

H-D exchange of ortho and meta-substituted anilines.¹⁵

The applicability of this method to more complex pharmaceutical compounds was investigated using diclofenac as an example (Scheme 2). Like acetaminophen, diclofenac is an aromatic amine or amide-containing NSAID with wide-ranging therapeutic applications. Upon subjecting the sodium salt of diclofenac to the described experimental conditions, cyclization was observed to the amide derivative 18. This transformation was precedented in another acid-catalyzed H-D exchange experiment of diclofenac, and the original sodium salt can be recovered using a simple base-catalyzed procedure.^5a Significant H-D exchange was observed in the ring annulated to the newly-formed lactam at the positions ortho and para to nitrogen. However, little H-D exchange was observed at the other aromatic positions.

Scheme 2.

H-D exchange of diclofenac.¹⁵

Conclusion

Using deuterated trifluoroacetic acid, the rapid and efficient H-D exchange of a wide variety of aromatic amines and amides was achieved without the need for metal salts or other co-catalysts. Direct H-D exchange of valuable pharmaceutically relevant compounds such as acetaminophen and diclofenac was conducted, engendering the possibility of applying this technique towards deuteration of other biologically active compounds. The exchange reaction generally proceeded according to typical EAS patterns, but was inhibited by strongly basic amines or highly deactivated acetanilides.