Malic acid production methods

DIFFERENT METHODS OF MALIC ACID PRODUCTION 9.1.1 Malic Acid... 

The conventional method for malic acid production is extraction from fruits. However, this method has no convenient application, owing to its low production capacity (Tsao et al., 1999). Malic acid can be chemically synthesized by the hydration of either maleic or fumaric acid. The enzymatic conversion of fumaric acid mediated by fumarase also permits malic acid production. The conversion of chemically derived fumaric acid to L-malic acid using fumarase (fumaratehydratase, EC 4.2.1.2) can be achieved by biological hydration or the fungal fermentation of simple sugars (Battat et al., 1991 Neufeld et al., 1991 Alberty, 1961). 

Make acid yields coumaUc acid when treated with fuming sulfuric acid (19). Similar treatment of malic acid in the presence of phenol and substituted phenols is a facile method of synthesi2ing coumarins that are substituted in the aromatic nucleus (20,21) (see Coumarin). Similar reactions take place with thiophenol and substituted thiophenols, yielding, among other compounds, a red dye (22) (see Dyes and dye intermediates). Oxidation of an aqueous solution of malic acid with hydrogen peroxide (qv) cataly2ed by ferrous ions yields oxalacetic acid (23). If this oxidation is performed in the presence of chromium, ferric, or titanium ions, or mixtures of these, the product is tartaric acid (24). Chlorals react with malic acid in the presence of sulfuric acid or other acidic catalysts to produce 4-ketodioxolones (25,26). 

Tobacco and its alkaloids have long ceased to have any therapeutic importance, but their extensive use as insecticides and the demand for nicotine for the manufacture of nicotinic acid have stimulated interest in processes of extraction and methods of estimation. On the latter subject there is a voluminous literature, of which critical resumes have been published by various authors.Recent work on this subject has been specially concerned with (1) the development of miero- and semi-miero-methods suitable for estimating nieotine in tobacco smoke and the distribution of nieotine on sprayed garden produce, in treated soils and in tobaeeo leaves,(2) the study of conditions necessary to ensure satisfactory results in using particular processes, " and (3) methods of separation and estimation of nicotine, nomicotine and anabasine in mixtures of these bases. ) In the United States and in Russia considerable interest is being shown in the cultivation of types of tobacco rich in nicotine, in finding new industrial uses for tobacco and its alkaloids, and in possible by-products from tobacco plants such as citric and malic acids, i " Surveys of information on tobacco alkaloids have been published by Jackson, i Marion and Spath and Kuffner. ... 

This process has been operated successfully by the Tanabe Seiyaku Co. in Japan since 1973. Similar processes have since been commercialised by other companies, such as the Kyowa Hakko Co., often using different immobilisation methods such as polyurethane. The same iimnobilized cell approach has also been used by Tanabe since 1974 in their coimnercial process for the production of L-malic acid from fumarate using the hydratase activity of Brevibacterium ammoniagenes cells. 

It is interesting to note that the oxa-analogous Michael addition was reported for the first time in 1878 by Loydl et al. [19] in their work on the synthesis of artificial malic acid, which was five years ahead of the discovery of the actual Michael reaction described first by Komnenos [20], Claisen [21], and later Michael in 1887 [22] as one of the most important methods for C—C bond formation. In continuation of the early work on the oxa-Michael addition [23], the inter- and intramolecular additions of alkoxides to enantiopure Michael acceptors has been investigated, leading to the diastereo- and enantioselective synthesis of the corresponding Michael adducts [24]. The intramolecular reaction has often been used as a key step in natural product synthesis, for example as by Nicolaou et al. in the synthesis of Brevetoxin B in 1989 [25]. The addition of oxygen nucleophiles to nitro-alkenes was described by Barrett et al. [26], Kamimura et al. [27], and Brade and Vasella [28]. 

Tyj"alo—lactic fermentation can be defined as the bacterial conversion of L-malic acid to L-lactic acid and carbon dioxide during storage of new wine. Malic acid is dicarboxylic, but lactic acid is monocarboxylic therefore, the net result of malo-lactic fermentation in wine, aside from the production of carbon dioxide, is a loss in total acidity. In commercial practice, this fermentation is not well understood, and better methods of controlling it are sought. 

Other important applications in the food industry running at a large scale are the production of L-aspartic add with Escherichia coli entrapped in polyacrilamides [6], the immobilization of thermolysin for the production of aspartame [14], The production of L-alanine by Tanabe Seiyaku [7], the production of frudose concen-centrated syrup [3], the production of L-malic acid by the use of Brevibacterium ammoniagenens immobilized in polyacrilamide by entrapment immobilization methods [11] and L-aminoacids production by immobilized aminoacylase [5],... 

The reverse of these reactions viz., the conversion of maleic and fumaric acids, by the addition of hydrogen bromide, into mono-brom succinic acid by the addition of two bromine atoms, into di-brom succinic acid and also by the addition of two hydrogen atoms, into succinic acid itself all show these same relations of maleic and fumaric acids to succinic acid and its bromine substitution products and establish the constitution of these isomeric di-basic unsaturated acids as given. The two acids may also be prepared from malic acid which is, in fact, the chief method by which they are prepared. This reaction will be considered later when malic acid itself is studied. 

The solution of maleic acid after being decolorized may be used directly for the production of malic acid by hydrolysis, or of succinic acid by hydrogenation. To form malic add the solution is heated in aluminum-bronze autoclaves. This alloy is not suitable for the absorber but may be used here due largely to the absence of oxygen. l or small scale succinic add production the electrolytic reduction method of Norris is more useful, but for large scale operation catalytic hydrogenation has been found to be desirable. 

In 1974 we succeeded in the industrial production of L-malic acid from fumaric acid by Brevibacterium ananoniagenes cells immobilized by the polyacrylamide gel method [9, 10]. The asymmetric reaction catalyzed by the fumarase activity of the cells is shown below. 

If a compound is nonfluorescent, it may be converted to a fluorescent derivative. For example, nonfluorescent steroids may be converted to fluorescent compounds by dehydration with concentrated sulfuric acid. These cyclic alcohols are converted to phenols. Similarly, dibasic acids, such as malic acid, may be reacted with j8-naphthol in concentrated sulfuric acid to form a fluorescing derivative. White and Argauer have developed fluorometric methods for many metals by forming chelates with organic compounds (see Ref. 23). Antibodies may be made to fluoresce by condensing them with fluorescein isocyanate, which reacts with the free amino groups of the proteins. NADH, the reduced form of nicotinamide adenine dmucleotide, fluoresces. It is a product or reactant (cofactor) in many enzyme reactions (see Chapter 24), and its fluorescence serves as the basis of the sensitive assay of enzymes and their substrates. Most amino acids do not fluoresce, but fluorescent derivatives are formed by reaction with dansyl chloride. 

In succession to the L-aspartic acid production, in 1974 we succeeded in the third industrial application, i.e. the production of L-malic acid from fumarlc acid by immobilized microbial cells. L-Mallc acid is an essential compound in cellular metabolism, and is mainly used in pharmaceutical field. L-Malic acid can be produced by fermentative or enzymatic methods from fumarlc acid by the action of fumarase as follows. 

Then the conditions for continuous production of L-mallc acid by a column packed with this bile extract treated immobilized cells was studied. When 1 M sodium fumarate (pH 7.0) is passed through the column at 37 C at flow rate of space velocity=0.2 hr the reaction reaches an equiliblium. From the effluent of the column, L-malic acid can be obtained by ordinary methods. Average yield of pyrogen-free pure L-malic acid from consumed fumaric acid is around 70% of the theoretical. Tanabe Seiyaku Co., Ltd. is operating this production system since 1974, and we are satisfied both with the economical efficiency and with the quality of product. 

Succeeding to the production of L-aspartic acid, we tried to apply this K-carrageenan method to improve the productivity for L-malic acid [19J. 

As shown in Table 4, it is evident that K-carrageenan method is more advantageous than the conventional polyacrylamide method. So, we changed the polyacrylamide method to the K-carrageenan method in 1977. This new method also gives us satisfactory result for industrial production of L-malic acid. 

Methods. The most common procedure for determining malic acid is that of permanganate oxidation in a buffered solution to produce acetaldehyde as developed for musts and wines by Peynaud (1938b, 1946). By slow introduction of very dilute permanganate a recovery as high as 98% to 99.2% was reported, although Joslyn and Oomar (1938) obtained lesser recovery from pure aldehyde solutions. The disadvantage of this procedure is the correction needed for the presence of tartaric acid. Amerine and Winkler (1943) and Amerine (1950-1951) were unable to obtain a constant acetaldehyde production from tartaric acid and preferred to remove most of it as potassium acid tartrate. Some coprecipitation of malic acid probably takes place. 

The malic acid is isolated and incubated with fumarase until no further loss of carbon-bound tritium results. [The enzyme catalyses trans removal of water, as (1.28) t L30).] For the samples of malate derived from (/ )-acetic acid, the minor product L29) loses TOH. For the material derived from (5 )-acetic acid it is the major product 1.33) which loses TOH. So for (/2)-acetic acid a higher retention of tritium (ca. 75%) is observed in the fumarase equilibration and for (5)-acetic acid a lower value (ca. 25%). With the method of analysis established for samples of acetic acid of known chirality it can be used to determine the configuration of samples of acetic acid of unknown chirality derived from biological reactions (for examples see Section 4.2 and 4.4, and [14]). 

However, this is a good example of the dangers which face the unwary in asymmetric synthesis, since it was later discovered that the hydrolysis step is actually accompanied by efficient inversion of absolute configuration at the stereogenic centre. Thus it is the (5)-product from quinine which gives the (/ )-malic acid and vice versa as shown. Regardless of this complication, the method allows convenient access to either enantiomer of the synthetically useful malic acid on a commercial scale. 

Alizade and Simon (1973) studied the stereochemistry of this transformation. Enzymatic methods were used to determine the specific quantities of the stereoisomers. In addition, the fermentation of radioactively labeled glucose and malic acid permitted the study of their products. The heterofermentative cocci (Oenococcus), abundant or exclusive during winemaking, were found to present several properties. They form exclusively D-lactic acid from glucose (Chapter 4) and exclusively L-lactic acid from L-malic acid (Figure 5.4). 

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