Aspartame synthesis, commercial

There are a number of reported solid-to-solid reactions where products precipitated as salts rather than as neutral compounds. The thermolysin-catalyzed production of the potassium salt of Z-aspartame [51, 52], the commercialized process of aspartame synthesis, where a salt of cationic D-Phe-OMe and anionic Z-aspartame precipitates [53], and the enzymatic conversion of solid Ca-maleate to solid Ca-D-malate [44] are examples of such behavior. 

Scheme 2. Anic and Enichem s commercial synthesis of aspartame (7) using catalytic asymmetric hydrogenation.

Today, it is well-known that peptides or proteins exhibit various kinds of taste. Our group has been researching on the relationship between taste and structure of peptides, BPIa (Bitter peptide la, Arg-Gly-Pro-Pro-Phe-Ile-Val) (7 as a bitter peptide, Om-p-Ala-HCl (OBA), Om-Tau-HCl as salty peptides(2j, and "Inverted-Aspartame-Type Sweetener" (Ac-Phe-Lys-OH) as a sweet peptide(5). The relationship between taste and chemical structure was partly made clear. Since commercial demand for these flavor peptides is increasing, we need to develop new synthetic methods which can prepare these peptides in large scale. We developed the following two methods (1) protein recombination method as a chemical method, (2) enzymatic synthesis using chemically modified enzyme as a biochemical method. 

Homogeneous asymmetric hydrogenation is a practical synthetic method (27). The DIPAMP-Rh-catalyzed reaction has been used for the commercial production of (S)-DOPA [(5)-3-(3,4-dihydroxy-phenyl) alanine] used to treat Parkinson s disease (Monsanto Co. and VES Isis-Chemie) (Scheme 12) (27, 28). (S)-Phenylalanine, a component of the nonnutritive sweetener aspartame, is also prepared by en-antioselective hydrogenation (Anic S.p.A. and Enichem Synthesis) (29). A cationic PNNP-Rh(nbd) complex appears to be the best catalyst for this purpose (15c) (see Scheme 5 in Chapter 1). 

A number of synthetic peptides are significant commercial products, ranging from the sweet dipeptide aspartame (L-aspartyl-L-phenylalanine methyl ester) to clinically used hormones such as insulin and oxytocin. L-Aspartyl-L-phenylalanine methyl ester (3 Scheme 2) is the methyl ester of the C-terminal dipeptide of gastrin. It was found accidently during the synthesis of gastrin that this synthetic sweetener is about 200 times as sweet as sucrose.f This pleasant sweetness without a bitter aftertaste was the reason that L-aspartyl-L-phenylalanine methyl ester was approved in many countries as a food additive, receiving much attention as a low-calorie sweetener. L-Aspartyl-L-phenylalanine methyl ester can be prepared by various chemical routes and the first enzymatic procedure of commercial interest was described by Isowa et al.h l In the industrial process,L-Asp and DL-Phe were chosen as inexpensive raw materials. L-Asp is available very inexpensively, whereas L-Phe is more expensive than DL-Phe. Z-D-Asp acts as a competitive inhibitor, while D-Phe-OMe... 

Various commercial routes for the production of L-phenyalanine have been developed because of the utilization of this amino acid in the dipeptide sweetener Aspartame. One route that has been actively pursued is the synthesis of L-phenylalanine from trans-cinnamic acid using the enzyme phenylalanine ammonia lyase (105,106). This enzyme catalyzes the reversible, nonoxidative deamination of L-phenylalanine and can be isolated from various plant and microbial sources (107,108). 

Isolation of the amino acids from such wastewaters might be of commercial value. For example, L-phenylalanine has been used as a precursor in the synthesis of a variety of industrial products, for example, the artificial sweetener Aspartame [117]. Purified amino acids can, on the other hand, be used as nutritional supplements in the diet of livestock [118], as weU as in human nutrition [119]. Therefore, there is potential for the extraction of amino acids from dairy and proteinaceous wastewaters for additional income of the producing industries. Application of LMs in treatment of such wastewaters could be of substantial benefit. 

L-aspartic acid ammonia lyase, or aspartase (E.C. 4.3.1.1) is used on a commercial scale by Kyowa Hakko, Mitsubishi, Tanabe and DSM to produce L-aspartic acid, which is used as a building block for the sweetener Aspartame, as a general acidulant and as a chiral building block for synthesis of active ingrediants[1]. The reaction is performed with enzyme preparations from E. coli, Brevibacterium jlavum or other coryneform bacteria either as permeabilized whole cells or as isolated, immobilized enzymes. The process is carried out under an excess of ammonia to drive the reaction equilibrium from fumaric acid (1) in the direction of L-aspartic acid (l-2) (see Scheme 12.6-1) and results in a product of excellent quality (over 99.9% e.e.) at a yield of practically 100%. The process is carried out on a multi-thousand ton scale by the diverse producers of L-aspartic acid. Site directed mutagenesis of aspartase from E. coli by introduction of a Cys430Trp mutation has resulted in significant activation and stabilization of the enzyme P1. 

Aspartame (APM), H-Asp-Phe-OMe, a nonnutritive high-intensity sweetener. APM is about 200 times sweeter than sucrose. It was first approved by the FDA in 1981 as a table-top sweetener and an additive, for example, in dry-based beverages, dry cereals, chewing gum, gelatins, puddings, instant coffee, and tea. APM was discovered accidentally during the recrystaUization of an intermediate of the synthesis of the C-terminal tetrapeptide of gastrin at Searle Co. For commercial synthesis, various... 

As weU as in edible-oil processing, the use of organic solvents is also relevant in the synthesis of amino adds and their derivatives. In producing dipeptides such as L-aspartyl-L-phenylalanine methyl ester, better known as aspartame, from amino acids and derivatives via enzymatic processes in organic solvents such as butyl acetate and 2-methyl-2-butanol, unreacted amino acids should be recycled after the synthesis to make the process more efficient. Reddy et al. tested several commercial membranes and found a polyamide-polyphenylene sulfone composite to be promising for this appHcation. However, further research is stiU needed here to apply other membrane materials, which are more resistant towards solvents like butyl acetate [33]. 

The Eprs of B. subtilis and related bacilli are also very relevant for commercial applications. In particular, these enzymes are employed in the manufacture of detergents, tanning of leather, management of industrial and household wastes, bioprocessing of X-ray or photographic films for the recovery of silver, protein hydrolysate preparation in the food industry, synthesis of aspartame, and other applications [102]. Recently, the fibrinolytic activity of Vpr was discovered. Accordingly, Vpr has the potential to work as a thrombolytic agent in medical applications [103, 104]. 

A commercially viable preparation of the ester of human insulin (35) has been developed using a trypsin-mediated exchange of a threonine residue for the terminal alanyl residue of porcine insulin (Scheme 3.22). This transformation was the first example of an enzymic semi-synthesis of a protein for use in medicine. (See Section 6.9.2 for further details on the production of Aspartame and insulin.)... 

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