Arginines from Ornithines (Guanidinylation Chemistry)

N^G-Substituted arginine derivatives are advantageously prepared by guanidinylation of the corresponding ornithine precursors (cf. Scheme 12).

NH NH R₁HN NR₂

O NH

NH₂

O

X R₁HN NR₂

- HX

Scheme 12: Conversion of ornithine derivatives into arginine derivatives by amidine group transfer-ring reagents (guanidinylation).

For the amine – guanidine transformation numerous methods have been developed.

The most common reagents for this purpose are of one the following types:

thioureas and S-methylisothioureas, 1H-pyrazole-1-carboxamidines, and triflylguani-dines.

5.1. Thioureas

One of the advantages of the thiourea method is the accessibility of N,N’ -disubstituted thioureas by the reaction of amines with isothiocyanates. Thus, a wide range of different, substituted guanidines is accessible. The reaction of thioureas

with amines is promoted by electrophilic agents, which convert the sulfur atom into a good leaving group. Desulfurization leads to an intermediate carbodiimide which readily reacts with primary or secondary amines to yield the desired guanidines^[28]. The most common desulfurizing agents for this purpose are metal ions (Hg²⁺, Cu²⁺), 1-methyl-2-chloropyridinium iodide (Mukaiyama’s reagent)^[29] or EDC^[30].

The reactivity of thiourea is enhanced by electron withdrawing subtituents at the nitrogen atom(s). Therefore, thioureas substituted with urethane-type protective groups (e.g. Boc, Cbz) are often used as efficient guanidinylating reagents. An additional advantage is that isolation and purification steps are facilitated due to the less polar character of the resulting protected guanidines. The free guanidines subsequently can be obtained using standard deprotection methods^§.

Serious side reactions which can occur are: i) reaction of the amine (or other nucleophilic sites in the substrate) with Mukaiyama’s reagent or EDC (especially when resin bound thioureas are employed) and b) decomposition of the intermediately formed carbodiimide, when unreactive or sterically hindered amines are used.

Scheme 13: Guanidinylation of ornithinamide 23 with thiourea 22 and Mukaiyama’s reagent did not yield the arginine derivative 24.

§ Hydrogen chloride (in organic solvent) does not reliably remove Boc groups from the guanidino nitrogen. However, in TFA/CH2Cl2 1:1 (v/v) cleavage is complete after 2–3 hours.

We probed the applicability of the thiourea/Mukaiyama’s reagent method for the preparation of ω-aminoacyl substituted BIBP 3226 (1) analogs. Thiourea fragment 22 was prepared in three steps and used in combination with 1-methyl-2-chloropyridinium iodide as guanidinylating reagent. Although the reagents were consumed, it was not possible to isolate the desired product 24; in fact, degradation and other side reactions led to a complex mixture of products.

5.2. S-Methylisothioureas

2-Alkylisothioureas have been applied as amidine source for the guanidinylation of amines since the beginning of the 20^th century. Methylation of thiourea with di-methylsulfate or methyliodide yields S-methylisothiourea, a basic compound, which is obtained in crystalline form as isothiouronium sulfate or iodide, respectively. N- and N,N’-protected derivatives of S-methylisothiourea have found widespread application as soluble, easily accessible, and efficient reagents for the preparation of protected guanidines from amines. N,N’-Bis(tert-butoxycarbonyl)-S -methylisothio-urea and N,N’-bis(benzyloxycarbonyl-S-methylisothioureas are commercially available guanidinylation reagents in common use. The reaction can optionally be supported by DMAP^[31] catalysis or addition of stoichiometric amounts of Hg²⁺

salts^[32]. Analogous reactions on solid support are described in literature^[33].

According to this methodology we developed a novel protocol for the preparation of N ^ω-acyl-substituted arginine derivatives, based on the use of N-acyl-N’-tert -butoxycarbonyl-S-methylisothioureas. Therefore, S-methylisothiouronium iodide^**

was mono-Boc-protected with hypostoichiometric amounts of Boc₂O in CH₂Cl₂ in the presence of triethylamine. Subsequently, the N-tert-butoxycarbonyl-S -methyliso-thiourea can be acylated under a broad variety of conditions. Even ω-tert -butoxycarbonylamino acids as acyl fragments were unproblematic. The thus obtained N-acyl-N’-tert-butoxycarbonyl-S-methylisothioureas were allowed to react

** The use of S-methylisothiouronium sulfate — instead of the iodide — leads to poor results due to insufficient solubility of the sulfate.

with the appropriate ornithinamides in the presence of one equivalent of HgCl₂ and two equivalents of triethylamine in DMF at ambient temperature (cf. Scheme 14).

Guanidinylation proceeds smoothly and the products were obtained in good yield and excellent purity. This reaction was the most versatile and satisfactory method for the preparation of N^G-acyl-substituted analogs of BIBP 3226 (1) and BIIE 0246 (2) among all tested procedures.

Scheme 14: Preparation of Acylguanidines from N-acyl-N’-tert-butoxycarbonyl-S-methylisothio-ureas. Conditions: i: HgCl2, NEt3, DMF, r.t. ii: TFA/CH2Cl2 1:1 (v/v).

5.3. 1H-Pyrazole-1-carboxamidines

In 1953 Scott et al.^[34] found that the reaction of amines with 3,5-dimethyl-1H -pyrazole-1-carboxamidine (25a, cf. Fig. 5) nitrate resulted in the formation of gu-anidinium nitrates. Four decades later, Bernatowicz et al.^[35] suggested 1H -pyrazole-1-carboxamidine hydrochloride as convenient reagent for the conversion of amines into guanidines.

Fig. 5: 1H-Pyrazole-1-carboxamidine derivatives as guanidinylation reagents.

As in case of the thioureas and isothioureas, diacylation considerably enhances reactivity^[36]. N,N’-Bis(tert-butoxycarbonyl)- and N,N’-bis(benzyloxycarbonyl)-1H -pyrazole-1-carboxamidine (25c,d) are commercially available, popular

guanidinyl-ation reagents, frequently used in peptide synthesis. Reactivity can be further improved by introduction of an electron-withdrawing nitro substituent in 4-position of the 1H-pyrazole heterocycle (25e^[37]).

By contrast, N-monoacylated derivatives are unreactive^[36]. Therefore, we had to use N-acyl-N’-tert-butoxycarbonyl-substituted 1H-pyrazole-1-carboxamidine derivatives (27, cf. Scheme 15) for the synthesis of N^G-acylated argininamides. Unfortunately, acylation of the monoacylated intermediate (26) happens only under forcing conditions. NaH^[36], LiH^[38], or LiHMDS^[39] were used as strong bases to generate the corresponding anion, which was acylated by anhydrides or acyl chlorides.

Alternatively, per-tert-butoxycarbonylation of N-monoacylated 1H -pyrazole-1-carb-oxamidines can be performed using Boc₂O/DMAP (cat.).

NH₂

Scheme 15: Preparation of N-tert-butoxycarbonyl-N’-acyl-1H-pyrazole-1-carboxamidines. Condi-tions: i: DIPEA, R-C(O)X (acyl chloride, anhydride, or activated carboxylate); ii: Boc2O/DMAP (10 mol-%); iii: Mg(ClO4)2, THF, 50 °C.

Treatment with Mg(ClO₄)₂ in THF at 50 °C selectively removes one Boc group from the disubstituted nitrogen atom yielding the N-acyl-N’-tert-butoxycarbonyl-1H -pyr-azole-1-carboxamidines (27)^[40]. By analogy with this procedure we prepared some of the N^G-acylated analogs of BIIE 0246 and BIBP 3226 (see Chapter 6 and 7).

Though the guanidinylation efficacy was satisfactory, the cumbersome preparation makes reagents of chemotype 27 less attractive. Especially, the presence of additional functional groups in the acyl fragment, as in our ω-(tert -butoxycarbonyl-amino)alkanoyl spacers, turned out to be problematic.

5.4. 1-Triflylguanidines

Recently, Feichtinger et al. introduced a novel type of guanidinylation reagents, the dialkoxycarbonyl protected trifluormethansulfonyl guanidines^{[41, 42]}. In a comparative study 28a proved to be superior to most other guanidinylation reagents^[42].

Schmuck et al. successfully prepared guanidinocarbonyl pyrroles as artificial receptors for carboxylate ions by activation of N-(1H-pyrrole-2-carbonyl)-N’-tert -butoxy-carbonyl guanidine with trifluormethanesulfonic anhydride and subsequent guanidinylation of the amine component^[43]. We adapted this approach for the synthesis of N^G-substituted 1 without isolation of the N,N’-diacyl-N-triflylguanidine. In our hands the method was successful for the synthesis of a 4-fluorobenzoyl substituted analog of BIBP 3226, but failed for the introduction of a 5-tert-butoxycarbonylaminopentanoyl spacer.