Microbiological assays thiamin

Kawasaki (68) briefly reviewed HPLC methods for determining total thiamine alone and in combination with riboflavin. Russell (44) provided a more detailed summary of HPLC methods, published between 1990 and 1994, for thiamine alone and in conjunction with other vitamins. Ball (45) reviewed selected HPLC analyses for thiamine in various foods, as well as other chemical and microbiological assays. 

Methylcobalamin, 516 5-Melhyl-cytosine, deaminatitm, 894 Methylglyoxal, 836 Methylmalonic acid (MMA), 434, 522 MethylmalcHiy. CoA, 434, 517, 518 Mevalonic acid, 327, 328 Mg-ATP complex, 795-796 Micelles, 25,27-29 MLcroaulophagy, 444 Microbiological assays, 508 biotin determination, 541 folate status by, 509 thiamin status, 607 Microcytic anemia, 5H Microsomal ethanol-oxidizing system, 247 Microvilli, 58 Milk... 

Thiamin status has been assessed by direct tests involving the measurement of thiamin levels in the blood or urine. The vitamin can be assayed by the thiochrome method or by microbiological assays. The disadvantage of these methods is that thiamin levels in normal individuals can vary greatly. The test organism used for microbiological assays may be Lactobacillus viridescens or Lactobacillus fermenti. 

Historically, assessment of thiamine status was by animal bioassay (the correction of bradycardia in thiamine-deficient rats) and later by microbiological assays using the fungus Phycomyces hlakesleeanus, yeast fermentation, or bacteria of the Staphylococcus, Streptococcus, or Lactobacillus species. Some bacterial microbiological assays are still in use in the food industry. Early chemical methods were often based upon the production of a fluorophore, thiochrome, when thiamine is oxidized with ferricyanide in alkaline solution, a property that is used in some modern chromatographic methods. 

Several B vitamins, including folic acid, niacin, pyridoxine, and pantothenic acid, are routinely determined using microbiological assays, details of which can be found in the AOAC Official Methods of Analysis. Standard methods for thiamine determination using fluorimetric detection are also detailed in the AOAC methods in addition, LC techniques are now being used routinely for thiamine and other B vitamins, e.g., riboflavin. 

Vitamins may be assayed by means of microbial electrodes constructed from cells whose metabolic activity or growth is dependent on the vitamin assayed. Such microbial strains are well known from the commonly used microbiological assays. For instance, the assessment of thiamine was performed by means of yeast cells immobilized in a calcium al inatte membrane. The difference in oxygen consumption before and after incubation with thiamine in the sample is related to the thiamine content [269]. Similar biosensors have been worked out for nicotinic acid (with Lactobacillus arabinosa) [270], nicotine amide (with B. pumilus and E. coli) [271], and L-ascorbic acid (with Enterobacter agglomerans) [272]. 

Levels of excreted thiamine in urine have been measured using various techniques, including the trichrome method (based on the oxidation of thiamine) (Bessey et al. 1952), microbiological assay using Lactobacillus viridescens (Sauberlich et al. 1979), and high performance liquid chromatography (HPLC) (Roser et al. 1978). Urinary assessment of thiamine status presents several... 

There are various microbiological, chemical and animal assays available for thiamine analysis. Method selection depends on the accuracy and sensitivity required, and on the interferences due to the sample matrix. Although microbiological assays do exist for thiamine, these are rarely used. The highly fluorescent products are the basis of the thiochrome procedure for thiamine quantiflcation. Thiochrome analysis has been used to develop most of the analytical data available on the thiamine content of the food supply. 

The assays have been a powerful tool for thiamine analysis in various foods. However, the analytical process may take up to 72 h and is often plagued by poor reproducibility. It allows for the detection of thiamine amounts between 5 and 50 ng. The aseptic technique should be used throughout the microbiological assay procedure under the same conditions for successful results. 

A combined microbiological assay for thiamine phosphates after separation by PPC was reported to yield a detection limit of approximately 0.02 Lig (60 pmol) of thiamine, which is still at least three orders of magnimde higher than that obtained by high-performance liquid chromatography (HPLC) with fluorescence detection. 

Assays of thiamine in food, fortified food, infant formulas and premixes are based on microbiological and chemical methods. 

MEASUREMENT/ASSAY. The thiamin content of foods is expressed in milligrams or micrograms. It is usually determine by rapid chemical or microbiological methods, which have largely replaced the older bioassay methods in which pigeons, rats, and chicks were used. 

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