Synthesis and Reactions of Amino Acids

Synthesis of Amino Acids

Industrial production of Glycine, which lacks any asymmetric center, is mainly by chemical synthesis. D,L-Methionine is chemically synthesized on large scale mainly to meet the animal feed industry. Animals have a D-amino acid oxidase and the required transaminase, which together convert the racemic methionine to the nutritive L-form. The classical process of acid hydrolysis of peptides, as the commercial process, has been restricted only to low volume amino acids. L-amino acids required in large volume are produced by fermentation procedures using engineered bacteria.

Interest in chemical synthesis of amino acids continues unabated, mainly due to the growing demand for D-amino acids and unusual amino acids in optically pure forms. Retro-analysis of the structure leads to several feasible pathways for the synthesis of amino acids. Fig 2.20 depicts a retro-analysis for α-amino acids.

Fig 2.20: Retro-analysis for the synthesis of α-amino acids.

Reduction of double bonds:

The asymmetric centre at α carbon could be introduced through reduction of a double bonds at the α-position. It could be reduction of C = N to C – N (Mode ‘a’) or reduction of C = C to C – C (Mode ‘b’).

Reduction of C=N:

Reduction of the C=N moiety is seen in the biogenesis of amino acids. For example, glutamic acid is biosynthesized from α-ketoglutarate (Fig 2.21). The keto group is first condenced with ammonia to give an imine, which is then, reduces through a NADPH to NADP+ oxidation sequence as shown below.

Fig 2.21: Biogenitic route - α-ketoglutarate to glutamic acid.

In the laboratory, this strategy could be realized through imine or shiff base formation reaction as shown below (Fig 2.22).

Fig 2.22: Chemical synthesis through C=N reduction strategy.

An early example for the asymmetric synthesis of amino acids using mode ‘a’ was reported by Hisley et.al.,(1961) (Fig 2.23). They converted various α-keto-acids to a chiral schiff base using optically active methylbenzylamine. On reduction with palladium hydroxide / C catalyst, the optically active amino acids were synthesized in good yields. Thus, pyruvic acid was converted to D-(-)-alanine in an overall yiels of 78% with high ee% (91%ee). The approach is suitable for several natural L- amino acids and well as D- and L- unnatural amino acids.

Fig 2.23: Hisley’s chiral synthesis of amino acids (1961)

Albertson (1946) introduced the imine moiety as an oxime by nitrosozation of diethyl malonate and reduced. The active methylene was alkylated after protecting the amine. Deprotection/ decarboxylation step yielded the substituted amino acid '(Fig 2.24).

Fig 2.24: Albertson’s nitrosozation route for synthesis of amino acids

Note that the extra carboxylate group serves as an activating group for facititating the alkylation step. One carboxylate is then removed to expose the α-amino acid moiety.

Reduction of the C=C:

Reduction of the C=C (mode ‘b’) is another good method for introducing the alkyl group on to the glycine moiety.

Erlenmeyer synthesis (Erlenmeyer azlactone synthesis)(1893): File:Fig2.2...img3.png File:Fig3.3...img4.png

The oxidation with the two separate groups of the bromine water and the hexane were obtained In this synthesis, N-benzoylglycine (hippuric acid) was condensed with benzaldehyde, in presence of acetic anhydride / sodium acetate. This procedure gave the Media:Knoevenagel condensation and cyclised the acid to an azlactone. The heterocycle was subjected to mild hydrolysis with 1% NaOH . The double bond was reduced with sodium amalgam and the benzoyl protection was removed by acid hydrolysis (Fig 2.25).

Fig 2.25: Erlenmeyer azlactone synthesis (1893)

This versatile procedure has been recently modified for the chiral synthesis of enantiomerically / diastereomerically pure amino acids that are in great demand. A good example is the industrial synthesis of L-Dopa, a drug used in the treatment of Parkinson ’s disease. W.S Knowles shared the 2001 Nobel Price with R. Noyori and K.B. Sharpless for their contributions in the area of asymmetric catalytic reductions (Fig 2.26). Knowles developed several chiral phosphine –metal catalysts for asymmetric reductions (http://nobelprize.org/nobel_prizes/chemistry/laureates/2001/index.html).

Several catalysts are now available for reduction of C=C to expose enantiopure amino acids.

Fig 2.26: Synthesis of L-Dopa by ‘mode b’

Introduction of nitrogen by S_N2 process:

Perkins (1858) displaced the halogen on α-haloacids by direct amination using ammonia. The process worked well with several substituted acetic acids (Fig 2.27).

Fig 2.27: Mode ‘c’ by displacement by ammonia.

for S_N2 displacement of halogens by an – NH₂ moiety. This procedure has been widely adopted for the synthesis of amino acids (Fig 2.28). Gabriel’s phthalimide synthesis (1889) is a general procedure

Fig 2.28: Gabriel synthesis of amino acids

F. Effenberger et.al., displaced the halogen in the haloesters using cyanate as

  F. Effenberger et.al., Angew. Chem. Int. ed. Engl. 18, 474 (1979). 

the nitrogen source. The general scheme and their application to the synthesis of Lysine are shown below (Fig 2.29). The yields are generally good. The cyanate ion is known to displace by S_N2 process, which means an inversion at the asymmetric center when the scheme is applied to enantiometically pure molecules.

Fig 2.29: Effenberger’s cyanate route for introducing nitrogen

Introduction of alkyl groups:

Myers A.G. et.al. achieved a direct C-alkylation on glycine (Mode ‘d’) by using the technique of attaching a chiral auxiliary. They used the cheap and easily available (R,R)-(-)-pseudoephedrine as auxiliary and lithiation / alkylation sequence and developed a general synthesis of optically active amino acids (Fig 2.30). The alkylation reaction proceeded in high yield using a wide variety of electrophiles and with excellent diastereoselectivity. Like the alkylation substrates, the products of the alkylation reaction are frequently crystalline and are readily recrystallized to ≥99% de. This procedure is a highly practical method for the asymmetric synthesis of α-amino acids.

  Myers AG et.al., J. Am.Chem. Soc.,119, 8488 (1995). Organic Syntheses, Coll. Vol. 10, p.12 (2004); Vol. 76, p.57 (1999).

Fig 2.30: Chiral aucilliary route by Myers.

The malonic ester procedure (Fig 2.31) has been used for the synthesis of cysteine and cystine.

Fig 2.31: An amidomalonate route to amino acids

T. Ooi et.al., used a Phase Transfer Catalyst (PTC) to effect the alkylation reaction on activated glycine (Fig 2.32).

Fig 2.32: Phase-transfer catalyst for asymmetric synthesis of amino acids.

   T. Ooi et.al.,O rg.L ett.,9, 3945 (2007);  N.D. Smith et.al., Org. Lett., 7, 255 (2005) 

N.D. Smith et.al., reported an Media:organocupprate catalysed ring cleavage with Media:grignard reagents to give enantiopure α-amino acids (Fig 2.33).

Fig 2.33: Chain elongation procedure for α-amino acids by Smith (2005)

  N.D. Smith et.al., Org. Lett., 7, 255 (2005)

Several other procedures have appeared in recent years, P. Cali et.al., reported C-arylation of diethyl N-Boc-iminomalonate, using a grignard reagent.

   P. Cali et.al., Synthesis, 63 (2002).

This procedure was adopted for multigram synthesis of arylglycines. This is an addition reaction to C=N (Fig 2.34).

Fig 2.34: Arylation sequence on iminomalonate by Cali et.al.,

Functional Group Transformation (FTR) procedures:

Ethyl cyanoacetate or diethyl malonate have been common starting materials in organic synthesis. In Media:Darapsky synthesis (1932) the cyano group becomes the acid moiety of the amino acid. The α-amino group comes from the original ester group after a Media:Curtius rearrangement (Fig 2.35).

Fig 2.35: Functional Group Transformation – Curtius rearrangement on cyanoacetic acid.

In malonic ester based synthesis of amino acids, one of the ester groups undergoes a Curtius or Media:Hoffmann rearrangement (Fig 2.36). Note the partial hydrolysis of the diester to half ester through careful partial hydrolysis using one equivalent of dilute base.

Fig 2.36: FGT seguence - Malonic ester - Curtius rearrangement

Multicomponent condensation routes:

Media:Stecker Synthesis (1850) is a versatile Media:Multicomponent Condensation (MCC) route for the synthesis of α-amino acids. A mixture of an aldehyde (or ketone),

  A. Strecker, Ann. 75, 27 (1850); 91, 349 (1854).

ammonium chloride and potassium cyanide are mixed in a suitable polar solvent. The first condensation product is believed to be an imine, to which a nitrile group is added to give an amino nitrile. The cyano group is hydrolysed to acid with boiling mineral acid (aq. HCl or aq. H₂SO₄) (Fig 2.37). By using optically active methylbenzylamine, K. Harada et.al.,

Fig 2.37: Stecker’s MCC route for amino acids.

were able to achieve asymmetric synthesis of amino acids, using similar protocol.

 K. Harada et.al., Nature, 200, 1201 (1963)

Note that the chiral precursor is completely destroyed in the end through hydrogenolysis. The asymmetric induction was however only poor to moderate (9-58%).

Media:Bucherer Hydandoin synthesis (1934) is the most versatile MCC reaction applicable to aldehydes and hetones. The hydandoins are crystallisable solids. The ring cleavage could be effected by refluxing with acids or bases (Fig 2.38).

Fig 2.38: Media:Bucherer Hydandoin synthesis (1934)

Note that asymmetric ketones would lead to the synthesis of racemic disubstituted amino acids. The mechanism for this reaction has not been established. Methionine has been synthesized by this procedure as shown below (Fig 2.38a).

Fig 2.38a: Methionine by Bucherer Hydandoin synthesis

A recent example of MCC reaction for amino acid synthesis is the Media:Petasis reaction shown below. A proposed mechanism is also shown.

  Petasis, N. A.; Akritopoulou, I. (1993). "The boronic acid mannich reaction: A new method for the synthesis 
  of geometrically pure allylamines". Tetrahedron Lett. 34: 583–586. Petasis, N. A.; Zavialov, I. A. (1997). 
  "A New and Practical Synthesis of -Amino Acids from Alkenyl Boronic Acids". J. Am. Chem. Soc. 119: 445–446. 

Fig 2.39: Petasis reaction in amino acid synthesis

This reaction has been applied for the synthesis of amino acids. Note that,

  M. Follmann, F. Gaul, T Schäfer, S. Kopec, P. Hamley, Synlett, 2005, 1009-1011

unlike the other two MCCs discussed above, the Patasis reaction does not involve any toxic materials either as starting materials or by products Media:(Green Chemistry). This procedure enables chemists to quickly assemble even complex amino acids (Fig 2.40).

Fig 2.40: Complex amino acids using Patasis reaction

Through heterocycles: The hydantoin and azloctone procedures could be classified under synthesis using heterocycles. Another useful route is the alkylation of diketopiperizine, followed by hydrolysis as shown below (Fig 2.41).

Fig 2.41: Amino acids from diketopiperizine.

In 2003, E. Bunuel et.al. modified this synthesis to a very efficient asymmetric synthesis of amino acids starting from a chiral diketopiperizine as shown below (Fig 2.42).

Fig 2.42: Bunuel’s (2003) aminoi acid synthesis using chiral diketipiperizine.

  E. Bunuel et.al. Org. iomol. Chem., 2531 (2003).

General properties of amino acids:

Amino acids are generally colourless, crystalline solids with high melting points. Some amino acids melt with decomposition. The common amino acids are generally very soluble in water and insoluble in apolar solvents. These properties indicate that the molecule is very polar. As discussed earlier, amino acids are zwitterionic in nature. The titration curves discussed earlier indicate the nature of the amino acid and indicate the isoelectric point of the amino acid. The IR spectrum shows a characteristic hydrogen bonding pattern at 3130 – 3000 cm ⁻ 1 and a carboxylate ion peak at1600 – 1500 cm ⁻ 1.

Reactions of amino acids:

The presence of acid and base functions in the same molecule poses some special problems in dealing with amino acids. Nonetheless, the general chemistry resembles the chemistry of amines and acids.

Reactions as amines:

The amine group readily protonates and forms salts with mineral acids. The amino group could be acylated with common acylation reagents like acid chlorides, acid anhydrides and esters (Fig 2.43).

Fig 2.43: Acylation reactions at the amine moiety

Treating with nitrous acid (diazotisation) gives a hydroxyl group in the place of the amino group. The reaction occurs stereospecifically, with retention in configuration. This has provided an efficient process for the synthesis of optically pure α-hydroxyacids from the readily available chiral pool of amino acids. Nitrosyl chloride or bromide converts the amino group to the halide (Fig 2.44).

Fig 2.44: Diazotisation at the amine moiety.

For peptide chemistry, the most important reaction is the synthesis of chloroformates to give -NHCOOR group (Fig 2.45). This will be discussed in detail in the next section.

Fig 2.45: Carboalkoxyl protection at the amine moiety

Reactions of carboxylic acid:

The COOH group gives acid chlorides with thionyl chloride and phosphorous pentachloride. Since an amine group is also present, this procedure is normally applied only after the amine group is masked (protected) (Fig 2.46). This is commonly called N-protection. Acyl fluorides are also well documented. The chemistry of these active groups would be discussed later.

Fig 2.46: Formation of acid chloride on N-protected amino acids

The common esterification procedures could be directly applied to selectively protect the acid group as an ester (Fig 2.47). This is commonly known in peptide chemistry as C-protection.

Fig 2.47: Esterification of the COOH moiety

With lithium Aluminium hydride, the acid or ester groups are reduced to alcohol to give β-aminoalcohols (Fig 2.48). This procedure provides optically active β-aminols needed in drug chemistry.

Fig 2.48: Aminols from amino acids

Heavy metals such as copper salts readily chelate with both the reactive groups (Fig 2.49). They are used for separation of select amino acids in complex mixtures.

Fig 2.49: Copper chelates of amino acids

On heating with acetic anhydride and excess pyridine, the acid is converted to methylketone and the amine is acylated. This reaction discovered in 1928 is called Dakin-West reaction (Fig 2.50).

Fig 2.50: Dakin-West reaction

When formalin solution (aqueous solution of formaldehyde monomer) is added to glycine, methylene glycine is formed. The mechanism of this reaction is not known. This product is very acidic. It could be used for estimation of glycine by titration. This procedure is called Media:Sorenson titration (Fig 2.51).

Fig 2.51: An activated acid for Sorenson titration.

The esters of amino acids (as well as ester dipeptides) readily dimerise to diketopiperizines (Fig 2.52). This is a major side reaction, which should be averted during peptide syntheses.

Fig 2.52: Formation of diketopiperizines