Malic acid + alanine

Important one-centre compounds including glyceraldehyde, lactic and malic acids, alanine and butan-2-ol... 

This research group applied the same approach to another red wine type, Rioja (Lopez-Rituerto et ah, 2009). In this study, PCA, performed on the entire fermentation time course of 207 days, demonstrated increases in ethanol, succinic, lactic, and acetic acids, while the alanine and malic acid concentrations decreased. Metabolite changes occurring during alcoholic fermentation were evaluated by performing PCA of the first 7 days of the... 

FIGURE 4.21 H NMR spectra (400 MHz) of time course evolution of red wine in alcoholic and malolactic fermentations for grape red must (pH 3). Peaks 1, ethanol 2, ethanol satellites 3, lactic acid 4, acetic acid 5, succinic acid 6, malic acid 7, 2,3-butanediol 8, proline 9, alanine. (From Avenoza et at, 2006.)... 

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],... 

A similar process is also used for the production of L-malic acid from fumarate, in this case using a hydratase enzyme derived from Brevibacterium ammoniagenes. Another variation of the Tanabe technology involves the synthesis of L-alanine from L-aspartic acid through the use of immobilized whole cells (P dacunae) containing aspartate-decarboxylase. 

It is also possible to convert nonchiral readily available industrial organic chemicals into valuable chiral natural-analogue products. This is demonstrated by the conversion of achiral fumaric acid to L(-)-malic acid with fumarase as the active enzyme. The same compound is converted to the amino acid L(-h)-aspartic acid by Escherichia bacteria that contain the enzyme aspartase. If pseudomonas bacteria are added, another amino acid L-alanine is formed (Eq. 9.10). 

Three immobilized enzyme or microbial cell systems currently used industrially in synthesis of chiral amino acids plus one presently under development are described. L-amino acids are produced by enzymatic hydrolysis of DL-acylamino acid with aminoacylase immobilized by ionic binding to DEAE-Sephadex. Escherichia coli cells immobilized by K-carrageenan crosslinked with glutaraldehyde and hexamethylenediamine are used to convert fumaric acid and cimmonia to L-aspartic acid and Brevibacterium flavum cells similarly immobilized are used to hydrate fumaric acid to L-malic acid. The decarboxylation of L-aspcirtic acid by immobilized Pseudomonas dacunhae to L-alanine is currently under investigation. 

An in-depth stndy of the interactions between amino acids and tartaric and malic acids focused on alanine, arginine, and proline, present in the highest concentrations in wine, as well as on amino acids with alcohol fnnctions, i.e. serine and threonine (Dartiguenave et al, 2000). 

Immobilized enzymes found their most Important application In biochemical analysis and medicine (diagnostics, therapy). Immobilized enzymes can also be used for the production of certain compounds via blotrans-formatlon (e.g. malic acid, aspartic acid, alanine, etc.) In most cases they have been replaced by Immobilized cells. 

As outlined earlier in this chapter, the initial carboxylation product of CAM is oxalacetate. The oxalacetate is immediately reduced to malate or aminated to aspartate. It is malate or, more properly in CAM, malic acid which tends to accumulate. More than likely, through decarboxylation of malate, pyruvate is formed, some of which is aminated to alanine. Hence, the initial stable products of CAM are firstly malic acid, and then aspartate and alanine (Fig. 3.12). 

Fig. 3.12. Main reactions of CAM resulting in the primary product, malic acid, and secondary products aspartate, alanine, and pyruvate. Tertiary and other products are formed by the usual reactions of intermediary metabolism...

Another excellent example of the application of NMR techniques in the structure determination of natural products is the structural resolution of rhizobactin, a siderophore isolated from Rhizobium meliloti (354). The proton homonuclear 2D-correlated spectrum revealed the four separate coupled units, whereas the heteronuclear 2D-correlated spectrum established the assignments (Fig. 2.43). Together, these spectra revealed that rhizobactin is composed of one unit each of ethylene diamine, alanine, lysine, and L-malic acid. The sequencing of... 

Within less than 1 min the radioactivity of carbon dioxide can be picked up in sugar phosphates, phosphoglyceraldehyde, phospho-pyruvic acid, phosphoglyceric acid (PGA), amino acids (particularly alanine and aspartic acid) and organic acids (particularly malic acid) (Fig. 5.5). By reducing the period of exposure to CO to a few seconds it was possible to show that the first stable intermediate product of photosynthesis was PGA (Fig. 5.6) and that virtually all the C was located in the carboxyl carbon (marked ) of this compound. 

Two other types of C4 pathways are recognized. In type-2 plants, Atriplex spongiosa) and type-3 Panicum maximum) plants, malate is replaced by aspartate as the major C4 acid transported to the bundle sheath cells. After transport, aspartate is converted to OAA by transamination. In type-2 plants, OAA is reduced to malate, which in turn is decarboxylated by NAD-malic enzyme in the bundle sheath cell mitochondria to give NADH, CO2 and pyruvate. In type-3 plants, OAA is decarboxylated in the cytosol by PEP carboxykinase in the presence of ATP, yielding PEP, CO2 and ADP. The return of carbon to the mesophyll cells for regeneration of the CO2 acceptor occurs as pyruvate (or alanine to maintain nitrogen balance) in type-2 and as PEP (or again perhaps as alanine) in type-3. These variations in the C4 pathway are summarized in Table I (see also Ref. 14). 

Fig. 42.9. Metabolism of the carbon skeletons of BCAA in skeletal muscle. 1. The first step in the metabolism of BCAA is transamination (TA). 2. Carbon from valine and isoleucine enters the TCA cycle as succinyl CoA and is converted to pyruvate by decarboxylating malate dehydrogenase (malic enzyme). 3. The oxidative pathways generate NADH and FAD(2H) even before the carbon skeleton enters the TCA cycle. The rate-limiting enzyme in the oxidative pathways is the a-keto acid dehydrogenase complex. The carbon skeleton also can be converted to glutamate and alanine, shown in blue.

The basic construction of the mathematical model using simplified metabolic networks to describe the reactions of the citric acid cycle and associated transamination reactions between pyruvate and alanine, oxalacetate and aspartate and a-ketoglutarate and glutamate, and the use of the FACSIMILE program (Chance et al., 1977) to solve the rather large number of simultaneous differential equations generated by the model was the same as previously described (Chance etal., 1983). For the present experiments the model was expanded to include an input flux at the level of succinate to represent propionate metabolism to succinyl-CoA, and a dilution of the aspartate pool to represent net proteolysis. These input fluxes required an output flux of carbon from the citric acid cycle in order to maintain a steady state carbon balance, for which the conversion of malate to pyruvate via malic enzyme was chosen. The model calculates the unknown flux parameters to provide a minimum least squares fit of the C fractional enrichments of specific carbon atoms of metabolic intermediates as measured by C NMR spectroscopy. 

Fig. 3.1. Autoradiogram of two dimensional paper chromatogram of 60 minute dark fixation by Bryophyllum calycinum. 1, alanine 2, glutamine 3, asparagine 4, glycine 5, serine 6, glutamate 7, aspartate 8, citrate 9, isocitrate 10, malate 11, fumarate 12, succinate. The main labeled compound is malate. Other metabolically related organic and amino acids also appear. The only products after a 6-s exposure are malic and aspartic acids (data of P.Saltman et al., Plant Physiol. 32, 197-200, 1957, by permission)...

---