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PubMed2006

Replacing dietary fish oil by vegetable oils has little effect on lipogenesis, lipid transport and tissue lipid uptake in rainbow trout (Oncorhynchus mykiss).

In order to investigate the effects of dietary lipid sources on mechanisms involved in lipid deposition, two groups of rainbow trout were fed from first-feeding to the commercial size of 1 kg (for 62 weeks) with two diets differing only by lipid source: 100% fish oil or 100% blend of vegetable oils (55% rapeseed oil, 30% palm oil, 15% linseed oil). The activities and levels of gene expression of lipogenic enzymes (fatty acid synthetase, glucose-6-phosphate dehydrogenase and malic enzyme) in liver and of lipoprotein lipase in perivisceral adipose tissue, white muscle and liver were determined. Transport of lipid was studied by determining lipid composition of plasma and lipoprotein classes. We also examined the clearance of LDL by assaying the level of LDL receptor gene expression in several tissues. Total replacement of dietary fish oil by the blend of vegetable oils did not affect growth of rainbow trout and did not modify muscle lipid content. Hepatic lipogenesis and lipid uptake in perivisceral adipose tissue, white muscle and liver were also not modified by dietary treatments. Diets containing the blend of vegetable oils induced a decrease in plasma cholesterol and LDL. In trout fed the vegetable oils diet, expression of LDL receptor gene in the liver was down-regulated.

Animal Nutritional Physiological Phenomena↗
PubMed1987

Changing patterns of transcriptional and post-transcriptional control of liver-specific gene expression during rat development.

Genes coding for unique or tissue-specific (differentiated) functions in the liver are induced at different times during development. It has generally been felt that transcriptional control represents the dominant mechanism for regulating expression of these genes. We have determined the relative transcription rates and mRNA steady-state levels for a series of genes specifically or preferentially expressed in rat liver and find examples of transcriptional control (albumin, alpha-fetoprotein, alpha 1-antitrypsin, tyrosine aminotransferase, transferrin, and cytochrome P450, TF-1) and post-transcriptional control (alpha 1-acid glycoprotein, apolipoproteins A-1 and E, malic enzyme, and ATP citrate lyase), as well as "mixed" regulation (ligandin and cytochrome P450, R17). Examples have been identified in which the predominant mode for regulating expression of preferentially expressed genes changes from transcriptional to post-transcriptional at different stages of liver development and some members of multigene families (cytochrome P450s and apolipoprotein genes) also show independent and sometimes contrasting modes of regulation. Therefore, it appears that regulation of specific gene expression in the liver is a dynamic process, far more complex than heretofore suspected, and a much greater contribution of post-transcriptional regulation accounts for changes in expression of genes representing major functions of the liver.

ATP Citrate (pro-S)-Lyase↗
PubMed1991

Differentiation of Salmonella senftenberg into biogroups.

Ninety-six strains of Salmonella senftenberg, isolated between 1984 and 1986 from different parts of England and Wales, were tested for their biochemical reactions and biotyped according to the method of Duguid and others (1975). Nine biogroups were identified on the basis of their metabolism of L-tartrate, D-tartrate, Bitter's xylose and Stern's glycerol. In addition, fumaric, oxalic, succinic, glutaric, malonic, maleic, L-malic, L-aspartic, lactic and formic acids were used but did not increase the discrimination. Three biogroups (7, 2 and 5) accounted for 79 per cent of the cultures examined.

Bacterial Typing Techniques↗
PubMed2000

Evidence that the rate-limiting step for the biosynthesis of arachidonic acid in Mortierella alpina is at the level of the 18:3 to 20:3 elongase.

Mortierella alpina, a fungus used commercially as a source of arachidonic acid, 20:4(n-6), has been examined to see if growth on lipid-based carbon sources leads to repression of either fatty acid biosynthesis and/or fatty acid desaturation and elongation. Changes in the activities of ATP:citrate lyase, isocitrate lyase, carnitine acetyltransferase, malic enzyme, glucose-6-phosphate dehydrogenase and pyruvate kinase when the fungus was grown on fatty-acid-based (Tween) carbon sources were consistent with (i) the cells using the fatty acyl portion of the substrate as the sole carbon source, (ii) pyruvate kinase being the source of pyruvate for biosynthesis under these conditions and (iii) malic enzyme's major function being as a provider of NADPH for lipid biosynthesis. The abolition of fatty acid synthase activity when cells were grown on Tweens indicated the cessation of de novo fatty acid biosynthesis under these conditions. The fatty acyl composition of the lipid accumulated by the fungus grown on Tweens 20, 40 and 80 showed that desaturation and elongation of the substrate lipid still occurred. The absolute amount of arachidonic acid synthesized by Tween-grown cells was the same as for cells grown on glucose. The transformation of incorporated fatty acids into 20:4(n-6) was, it appeared, limited at the elongation of 18:3(n-6) to 20:3(n-6) as, in every case, 18:1, 18:2 and 18:3(n-6) increased in amount in the Tween-grown cells. These data show for the first time that fatty acid synthesis is regulated separately from fatty acid desaturation/elongation and that the latter reactions are not repressed by growth of the fungus on simple fatty acids. Furthermore, the data strongly implicate the elongation of 18:3(n-6) to 20:3(n-6) as the limiting step in arachidonic acid biosynthesis by Mort. alpina.

ATP Citrate (pro-S)-Lyase↗
PubMed1975

Regulation of the dicarboxylic acid part of the citric acid cycle in Bacillus subtilis.

The regulation of alpha-ketogluterate dehydrogenase, succinate dehydrogenase, fumarase, malate dehydrogenase, and malic enzyme has been studied in Bacillus subitilis. The levels of these enzymes increase rapidly during late exponential phase in a complex medium and are maximal 1 to 2 h after the onset of sporulation. Regulation of enzyme synthesis has been studied in the wild type and different citric acid cycle mutants by adding various metabolites to the growth medium. Alpha-ketoglutarate dehydrogenase is induced by glutamate or alpha-ketoglutarate; succinate dehydrogenase is repressed by malate; and fumarase and malic enzyme are induced by fumarate and malate, respectively. The addition of glucose leads to repression of the citric acid cycle enzymes whereas the level of malic enzyme is unaffected. Studies on the control of enzyme activities in vitro have shown that alpha-ketoglutarate dehydrogenase and succinate dehydrogenase are inhibited by oxalacetate. Enzyme activities are also influenced by the energy level, expressed as the energy charge of the adenylate pool. Isocitrate dehydrogenase, alpha-ketoglutarate dehydrogenase, succinate dehydrogenase, and malic enzyme are inhibited at high energy charge values, whereas malate dehydrogenase is inhibited at low energy charge. A survey of the regulation of the citric acid cycle in B.subtilis, based on the present work and previously reported results, is presented and discussed.

Adenine Nucleotides↗
PubMed2001

Partial amino acid catabolism leading to the formation of alanine in Periophthalmodon schlosseri (mudskipper): a strategy that facilitates the use of amino acids as an energy source during locomotory activity on land.

When the mudskipper Periophthalmodon schlosseri was exposed to terrestrial conditions under a 12h:12h dark:light regime the fish could be very active, and levels of total free amino acids increased significantly in the muscle and plasma. Alanine levels increased threefold in the muscle, fourfold in the liver and twofold in the plasma. Similar phenomena were not observed in the more aquatic mudskipper, Boleophthalmus boddaerti. From these results, we concluded that P. schlosseri was capable of partial catabolism of certain amino acids to support activity on land. The amino groups of these amino acids were transferred directly or indirectly to pyruvate to form alanine. The resulting carbon chain was fed into the Krebs cycle and partially oxidized to malate, which could replenish pyruvate through the function of malic enzyme. This favourable ATP yield from partial amino acid catabolism was not accompanied by a net release of ammonia. Such an adaptation would be advantageous to P. schlosseri confronted with the problem of ammonia excretion during aerial exposure. Indeed, when P. schlosseri were forced to exercise on land after 24 h of aerial exposure, the alanine level in the muscles increased significantly, with no apparent change in glycogen content. In addition, there was no significant change in the ATP level and energy charge of the muscle. In contrast, when B. boddaerti were exercised on land, glycogen levels in the muscles decreased significantly and lactate levels increased. In addition, muscle energy charge was not maintained and the ATP level decreased significantly. Hence, it was concluded that when P. schlosseri were active on land, they were capable of using certain amino acids as a metabolic fuel, and avoided ammonia toxicity through partial amino acid catabolism. Such a strategy is the most cost-effective way of slowing down internal ammonia build-up without involving energy-expensive ammonia detoxification pathways. Furthermore, an examination of the balance between nitrogenous excretion and accumulation in a 70 g P. schlosseri revealed that degradation of amino acids in general was likely to be suppressed to slow down the build-up of ammonia internally. It is possible that such a strategy may be widely adopted, especially by obligatory air-breathing fishes, to avoid ammonia intoxication during aerial exposure.

Adenosine Triphosphate↗
PubMedDate not supplied

[The determination of Krebs cycle carboxylic acids in the ejaculate by chromatography].

The authors suggest a highly informative and unsophisticated method of gas liquid chromatography of lactic, pyruvic, oxalic, butyric, malonic, succinic, glutaric, fumaric, oxalacetic, malic, alpha-ketoglutaric, cis-aconitic, tartaric, citric and isocitric acids in the ejaculate of infertile and fertile men and in testicles of experimental animals. The method is recommended for research and clinical studies of the metabolic processes associated with the functioning of the Krebs' cycle acids in the seminal fluid of infertile patients.

Carboxylic Acids↗
PubMed1970

The regulation of triglyceride synthesis and fatty acid synthesis in rat epididymal adipose tissue.

1. Adipose tissues from rats fed a balanced diet were incubated in the presence of glucose (20mm) with the following additions: insulin, anti-insulin serum, insulin+acetate, insulin+pyruvate, insulin+lactate, insulin+phenazine methosulphate, insulin+oleate+albumin, insulin+adrenaline+albumin, insulin+6-N-2'-O-dibutyryl 3':5'-cyclic AMP+albumin. 2. Measurements were made of the whole tissue concentrations of adenine nucleotides, hexose phosphates, triose phosphates, glycerol 1-phosphate, 3 phosphoglycerate, 6-phosphogluconate, long-chain fatty acyl-CoA, acid-soluble CoA, citrate, isocitrate, malate and 2-oxoglutarate, and of the release into the incubation medium of lactate, pyruvate and glycerol after 1h of incubation. 3. Fluxes of [(14)C]glucose carbon through the major pathways of glucose metabolism were calculated from the yields of (14)C in various products after 2h of incubation. Fluxes of [(14)C]acetate, [(14)C]pyruvate or [(14)C]lactate carbon in the presence of glucose were also determined. 4. Measurements were also made of the whole-tissue concentrations of metabolites in tissues taken directly from Nembutal-anaesthetized rats. 5. Whole tissue mass-action ratios for phosphofructokinase, phosphoglucose isomerase and the combined (aldolasextriose phosphate isomerase) reaction were similar in vivo and in vitro. The reactants of phosphofructokinase appeared to be far from mass-action equilibrium. In vitro, the reactants of hexokinase also appeared to be far from mass-action equilibrium. 6. Correlation of observed changes in glycolytic flux with changes in fructose 6-phosphate concentration suggested that phosphofructokinase may show regulatory behaviour. The enzyme appeared to be activated in the presence of oleate or adrenaline and to be inhibited in the presence of lactate or pyruvate. 7. Evidence is presented that the reactants of lactate dehydrogenase and glycerol 1-phosphate dehydrogenase may be near to mass-action equilibrium in the cytoplasm. 8. No satisfactory correlations could be drawn between the whole-tissue concentrations of long-chain fatty acyl-CoA, citrate and glycerol 1-phosphate and the observed rates of triglyceride and fatty acid synthesis. Under the conditions employed, the concentration of glycerol 1-phosphate appeared to depend mainly on the cytoplasmic [NAD(+)]/[NADH] ratios. 9. Calculated hexose monophosphate pathway flux rates roughly correlated with fatty acid synthesis rates and with whole tissue [6-phosphogluconate]/[glucose 6-phosphate] ratios. The relative rates of production of NADPH for fatty acid synthesis by the hexose monophosphate pathway and by the ;malic enzyme' are discussed. It is suggested that all NADH produced in the cytoplasm may be used in that compartment for reductive synthesis of fatty acids, lactate or glycerol 1-phosphate.

Acetates↗
PubMed2001

Effect of dietary seal and fish oils on lipid metabolism in hamsters.

Eicosapentaenoic and docosahexaenoic acids were distributed mainly in the sn-1 and 3 positions of seal oil triacylglycerols and in the sn-2 position of fish oil triacylglycerols. Seal oil-rich or fish oil-rich fats having constant polyunsaturated (PUFAs)/monounsaturated/saturated fatty acids and n-6/n-3 PUFAs ratios were fed to hamsters for 3 weeks. The control fat contained linoleic acid as the sole PUFA. The concentration of triacylglycerols in the liver was significantly lower in the fish oil group than in the control group. Phospholipid concentration in serum was lower and that in the liver was higher in the seal oil group compared with the fish oil group. The activities of fatty acid synthase (FAS), glucose-6-phosphate dehydrogenase (G6PDH), and the malic enzyme were significantly lower in both the fish and seal oil groups than in the control group. Dietary seal oil more effectively reduced arachidonic acid content in liver phosphatidylcholine and phosphatidylethanolamine and serum phosphatidylcholine than fish oil. These results showed that different intramolecular distribution of n-3 PUFAs influenced glycerolipid metabolism and arachidonic acid content in serum and liver phospholipids of hamsters.

Animals↗
PubMed2004

Upregulation of lipogenic enzymes genes expression in white adipose tissue of rats with chronic renal failure is associated with higher level of sterol regulatory element binding protein-1.

Chronic renal failure (CRF) frequently results in hypertriglyceridemia and elevated plasma concentration of very-low-density lipoprotein (VLDL). These abnormalities are thought to be primarily due to depressed lipoprotein lipase and hepatic lipase activities, as well as impaired clearance of plasma lipoproteins. Some results suggest that not only lipoproteins catabolism but also their overproduction might contribute to hypertriglyceridemia in CRF. Because sterol regulatory element binding protein (SREBP) plays an important role in the regulation of lipid homeostasis, increased level of this transcription factor might be involved in modulating lipid metabolism in CRF. The purpose of the present study is to determine whether there is an altered regulation of the SREBP-1 in CRF rats and whether the altered regulation of SREBP-1 is associated with the upregulation of lipogenic enzymes genes expression in CRF rats. In the white adipose tissue (WAT) of CRF rats, marked increases in the microsomal (precursor) and nuclear (mature) forms of SREBP-1 have been found. The increase in SREBP-1 was associated with an increased level of lipogenic enzymes (acetyl-coenzyme A [CoA] carboxylase [ACC], adenosine triphosphate-citrate lyase [ACL], fatty acid synthase [FAS], glucose 6-phosphate dehydrogenase [G6PDH], 6-phosphogluconate dehydrogenase [6PGDH], and malic enzyme [ME]) genes expression. In turn, this was associated with an increased rate of fatty acids synthesis in WAT and a significant increase in plasma triacylglycerol (TAG) and VLDL concentration. Our study indicates that WAT SREBP-1 expression is increased in CRF rats and that SREBP-1 may play an important role in the increased fatty acid synthesis. These results reveal another facet of disturbed lipid metabolism in CRF.

Adipose Tissue↗
PubMedDate not supplied

[The possibilities of GC/MS and HPLC in the analysis of sugars and acids in natural matrices].

A GC/MS procedure is described for the simultaneous quantitation of the minor and major constituents of several natural matrices, including fruits, vegetables, mushrooms, drug/industrial plants and honeys, as their trimethylsilyl derivatives, from one solution, by one injection. Selected minor components have been determined on the basis of their characteristic fragment ions, in the presence of extremely high excess of saccharides of various degree of polymerization. Selective fragmentation of these minor compounds in the ion trap detector provided possibilities for distinguishing them. In the case of honeys the method permitted the simultaneous quantitation of o-phosphoric, malic, shikimic, citric/isocitric, quinic, margaric, oleic and stearic acids, 5-hydroxymethylfurfurol and proline with the extremely high sugar contents of honeys (fructose, glucose, galacturonic acid, inositol, sucrose, trehalose, turanose, maltose, gentiobiose, isomaltose, raffinose, erlose, melezitose, maltotriose, panose, isomaltotriose) and allowed the fast evaluation of sugar and acid constituents of fifteen honeys from various floral and geological origin. Results revealed that (i) the minor components varied in the concentration range of 0.0001 to 0.43%, and, (ii) together with the saccharides of honeys made up the total of identified and determined constituents from 87.8% to 98.5%. Quantitative evaluation of the minor constituents was performed on the basis of their selective fragment ion values with an average reproducibility of 6.7% (RSD).

Biological Factors↗
PubMed1986

Coding nucleotide sequence of rat liver malic enzyme mRNA.

The nucleotide sequence of the mRNA for malic enzyme ((S)-malate NADP+ oxidoreductase (oxaloacetate-decarboxylating, EC 1.1.1.40) from rat liver was determined from three overlapping cDNA clones. Together, these clones contain 2078 nucleotides complementary to rat liver malic enzyme mRNA. The single open reading frame of 1761 nucleotides codes for a 585-amino acid polypeptide with a calculated molecular mass of about 65,460 daltons. The cloned cDNAs contain the complete 3'-noncoding region of 301 nucleotides for the major mRNA species of rat liver and 16 nucleotides of the 5'-noncoding region. Amino acid sequences of seven tryptic peptides (67 amino acids) from the purified protein are distributed through the single open reading frame and show excellent correspondence with the translated nucleotide sequence. The putative NADP-binding site for malic enzyme was identified by amino acid sequence homology with the NADP-binding site of the enoyl reductase domain of fatty acid synthetase.

Amino Acid Sequence↗
PubMed2000

Dietary conjugated linoleic acid consumption during pregnancy and lactation influences growth and tissue composition in weaned pigs.

We evaluated the effects of conjugated linoleic acid (CLA) on growth performance, tissue fatty acid composition and ex vivo lipogenic enzyme activity in piglets (n = 40) reared on sows fed diets supplemented with CLA or linoleic acid (LA). Weaned offspring of both sow groups were offered either a CLA- or LA-enriched starter diet for 35 d. The starter diets were formulated to contain 2 g CLA (containing 58.9 g CLA/100 g total fatty acids) or LA per 100 g feed. All piglets were slaughtered at 70 d of age and tissue samples of the back fat, omental fat and longissimus dorsi were collected. Irrespective of the dietary fat supplied in the starter period, piglets reared on the CLA sows had greater final body and warm carcass weights (P: < 0.01), and greater feed intake (P: = 0.02) than piglets reared on the LA sows. The dietary effect on the fatty acid composition was similar for the adipose and muscle tissues. Compared with the LA-enriched diets, CLA increased the level of total saturated fatty acids (P: < 0.05), whereas that of monounsaturated fatty acids was decreased (P: < 0.05). Dietary CLA increased glucose-6-phosphate dehydrogenase (P: < 0.01) and malic enzyme activities (P: < 0.06) in the fat tissues, but did not affect fatty acid synthase activity. The shift toward a higher deposition of saturated fatty acids and a lower deposition of monounsaturated fatty acids is the result of down-regulation of Delta9-desaturase activity that was induced by CLA rather than an altered rate of de novo synthesis.

Adipose Tissue↗
PubMed1977

Fatty acid synthesis in testes of fat-deficient and fat-supplemented rats.

Fatty acid synthesis was studied in testes of rats fed a fat-free or fat-supplemented diet. Testes of fat-deficient rats incorporated nearly twice as much intratesticularly injected [1-14C]acetate into total fatty acids (primarily into palmitic acid) as did supplemented rats. To determine the mechanism for the increased synthesis, the activities of the following enzymes were determined in the cytoplasmic fraction of testicular homogenates: fatty acid synthetase, acetyl CoA carboxylase [EC 6.4.1.2], citrate-cleavage [EC 4.1.3.8], malic [EC 1.1.1.38], and the glucose-l-phosphate dehydrogenase [EC 1.1.1.49]: 6-phosphogluconate dehydrogenase pair [EC 1.1.1.44]. Although the activity of fatty acid synthetase did increase in livers from fat-deficient rats, no change was observed in corresponding testes. No difference between the two groups could be demonstrated in testicular activity of citrate-cleavage enzyme, malic enzyme, or the glucose-6-phosphate dehydrogenase: 6-phosphogluconate dehydrogenase pair. However, the activity of cytoplasmic acetyl CoA carboxylase in testes of rats fed the fat-deficient diet was 1.4 times higher than the activity in testes of rats fed the supplemented diet. Fat deficiency did not affect the specific activity of the testicular microsomal elongation system, assayed by incubation with 14C-malonyl CoA. The concentration of unesterified fatty acids was lower in testes of the fat-deficient compared to supplemented rats, indicating that decreased inhibition of acetyl CoA carboxylase in the fat-deficient rats testes might have been responsible for the observed increased de novo synthesis of palmitic acid.

ATP Citrate (pro-S)-Lyase↗
PubMed1987

Determination of organic acids in sugar cane process juice by high-performance liquid chromatography: improved resolution using dual Aminex HPX-87H cation-exchange columns equilibrated to different temperatures.

The application of high-performance liquid chromatography to the quantitative analysis of organic acids in sugar cane process juice is described. Separation of these acids is achieved on Aminex HPX-87H cation-exchange columns. Resolution is improved by connecting two columns in series and equilibrating them at different temperatures. The acids in the sample are first isolated on DEAE-Sephadex A-25 anion-exchange resin equilibrated with 0.5 M isobutyric acid. They are then eluted with 0.5 M sulphuric acid and injected directly onto the HPLC column after filtration through a 0.45-micron membrane. This method allows the simultaneous determination of oxalic, cis-aconitic, citric, phosphoric, malic, trans-aconitic, succinic, glycolic, lactic, formic and acetic acids. Isobutyric acid is also separated on this system and provides a useful guide to the loading capacity of the anion-exchange resin.

Acids↗
PubMed2002

Metabolic flux analysis in Synechocystis using isotope distribution from 13C-labeled glucose.

Using the carbon isotope labeling technique, the response of cyanobacterial central carbon metabolism to the change in environmental conditions was investigated. Synechocystis was grown in the heterotrophic and mixotrophic cultures fed with 13C-labeled glucose. The labeling patterns of the amino acids in biomass hydrolysates for both cultures were detected by the two-dimensional 1H-13C correlation nuclear magnetic resonance (2D 1H-13C COSY NMR) spectroscopy and gas chromatography-mass spectrometry (GC-MS) technique. The in vivo intracellular flux distributions were then quantitated from the labeling measurements and metabolite balances using a parameters fitting approach. From the estimated flux distributions, it was found that the pentose phosphate pathway was the major pathway of glucose catabolism in the heterotrophic culture, while in the mixotrophic culture, the flux of CO2 fixation through the Calvin cycle was about two-fold of the glucose input flux. The relative flux through the phosphoenolpyruvate carboxylase was very high in both cultures, and this reaction represented about 25% of the assimilated CO2 in the mixotrophic culture. More importantly, we found a substantial outflow from the tricarboxylic acid cycle to glycolysis pathway carried by the malic enzyme, demonstrating the operation of a C4 pathway in cyanobacterial cells through the PEP carboxylase and malic enzyme. The estimated flux distributions also revealed that the NADPH synthesis was in excess relative to its requirement, and the excess NADPH might be reoxidized in cyanobacterial respiration to provide the energy for cellular requirement. Moreover, the analyzed result also suggested that the activity of the respiratory electron transport chain in cyanobacterial cells was not inhibited by light.

Amino Acids↗
PubMed1979

Properties of mutants of Escherichia coli lacking malic dehydrogenase and their revertants.

Mutants of Escherichia coli lacking malic dehydrogenase activity (mdh) were incapable of growth on acetate", succinate- or malate/mineral medium. Revertants of mdh strains which had regained the ability to grow on C4-dicarboxylic acids could be divided into two distinct classes. One type of revertant had regained the ability to synthesize functional malic dehydrogenase. The other type of revertant still lacked malic dehydrogenase activity but possessed a suppressor mutation which altered the regulation of the synthesis or activity of the C4-dicarboxylic acid transport system, resulting in increased C4-dicarboxylic acid transport activity. This latter class of revertants apparently synthesized oxalacetate from malate via the sequential actions of the NAD-linked malic enzyme, phosphoenolpyruvate synthetase, and phosphoenolpyruvate carboxylase. Evidence has been presented that is consistent with the hypothesis that oxalacetate is the inducer of the C4-dicarboxylic acid transport system. The inability of mutants lacking malic dehydrogenase to grow with a C4-dicarboxylic acid as the carbon source can be attributed to the difficulty such mutants have in synthesizing oxalacetate.

Escherichia coli↗
PubMed2004

Monitoring of the fermentation media of citric acid by the trimethylsilyl derivatives of the organic acids formed.

In this approach, a derivatization method is described for monitoring of organic acids in fermentation media without any separation step. The aqueous phase of fermentation media was evaporated and heated in a silylation reagent to form trimethylsilyl (TMS) derivatives. The silylated compounds are analyzed by 29Si nuclear magnetic resonance (29Si NMR) and gas chromatography-mass spectrometry (GC-MS). 29Si NMR can qualitatively monitor the components produced in the Krebs cycle. Quantification of these compounds is investigated by using selected ion monitoring mode of mass spectrometry. In this mode, mass to charge (m/z) values of their [M - 15]+ ions, which are 465, 275, 247, 221, 335, 251, and 313 of TMS derivatives of citric, alpha-ketoglutaric, succinic, fumaric, l-malic, oxaloacetic, and palmitic (as an internal standard), acids, respectively, are used. The limit of detection and the linear working range for derivatized citric acid were found to be 0.1 mg L(-1) and 10-3 x 10(4) mg L(-1). The relative standard deviation of the method for five replicates was 2.1%. The average recovery efficiency for citric acid added to culture media was approximately 97.2%. Quantitative results of GC-MS are compared with those obtained by an ultraviolet-visible method.

Aspergillus niger↗