The Assay of Thiamine in Food

Thiamine can accumulate in all cells of the body but there is no single specific storage site per se. The body does not store the vitamin and thus a daily supply is needed. Animal assays are used for determining the availability of thiamine in a food source. They are based on the effect of thiamine on the growth and evolution of diseases related to thiamine-lacking effects. Thiamine deficiency is associated with anorexia, tissue wasting, impaired cardiac function, weakness and neurological abnormalities, all of which are classical symptoms of beriberi. 

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. 

Coverage includes B vitamins and folate in the context of a historical background, disease, cardiovascular effects and the importance of vitamins in biochemistry as illustrated by a single vitamin. Thereafter there are chapters on the chemistry and biochemistry of thiamine, riboflavin, niacin, pantothenic acid, pyridoxine, biotin, folate and cobalamin. Methodical aspects include characterization and assays of B vitamins and folate in foods of all kinds, dietary supplements, biological fluids and tissues. The techniques cover solid-phase extraction, spectrofluorimetry, mass spectrometry, HPLC, enzymatic assay, biosensor and chemiluminescence. In terms of fimction and effects or... 

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. 

Thiamine, or vitamin Bi, is a water-soluble compound which is rapidly broken down by moist heat in neutral or alkaline solutions into its constituent pyrimidine and thiazole rings. The ready destructability of thiamine is important in human nutrition, since much may be lost in the preparation of food. Some of the biochemical methods used in evaluating thiamine nutrition are based on reactions with the thiazole and pyrimidine portions of the thiamine molecule. The thiochrome method is widely used in assaying biological materials for thiamine, while determination of the urinary excretion of pyramine (a pyrimidine-like compound) has been used to assist in assessment of nutritional status. 

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. 

Destruction vs. Formation. When looking for chemical markers of sterility, one is tempted to look for compounds that are destroyed at sterilizing temperatures for tire simple reason that the chenucal identity and the assay method is already known to the investigator. Several examples were listed by Kim and Taub (7). This approach has a limitation, because a typical chenucal reaction in foods is much slower than bacterial destruction at high temperatures and one has to be able to measure a small loss of the compoimd. For example, tire D-value (time required to reduce the concentration by 90%) for destruction of thiamin is 244 min at 122°C (8). The D-value for destruction of B. stearothermophilus is about 1 min at the same temperature. The D-value and k, the rate constant for a first-order reaction, are related by eq. (1). 

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