Assessment of salt intake to consider salt as a fortification vehicle for thiamine in Cambodia.

Thiamine deficiency is a public health issue in Cambodia. Thiamine fortification of salt has been proposed; however, the salt intake of lactating women, the target population, is currently unknown. We estimated salt intakes among lactating women (<6 months postpartum) using three methods: repeat observed-weighed intake records and 24-h urinary sodium excretions (n = 104), and household salt disappearance (n = 331). Usual salt intake was estimated by adjusting for intraindividual intakes using the National Cancer Institute method, and a thiamine salt fortification scenario was modeled using a modified estimated average requirement (EAR) cut-point method. Unadjusted salt intake from observed intakes was 9.3 (8.3-10.3) g/day, which was not different from estimated salt intake from urinary sodium excretions, 9.0 (8.4-9.7) g/day (P = 0.3). Estimated salt use from household salt disappearance was 11.3 (10.7-11.9) g/person/day. Usual (adjusted) salt intake from all sources was 7.7 (7.4-8.0) g/day. Assuming no stability losses, a modeled fortification dose of 275 mg thiamine/kg salt could increase thiamine intakes from fortified salt to 2.1 (2.0-2.2) mg/day, with even low salt consumers reaching the EAR of 1.2 mg/day from fortified salt alone. These findings, in conjunction with future sensory and stability research, can inform a potential salt fortification program in Cambodia.

Patient-Centric Assessment of Thiamine Status in Dried Blood Volumetric Absorptive Microsamples Using LC-MS/MS Analysis.

Many populations in low- and middle income countries are at a higher risk of thiamine deficiency, mainly due to the lack of dietary diversification and their reliance on staple crops low in thiamine content, such as polished rice. Unfortunately, symptoms of thiamine deficiency are variable and clinical determination of thiamine status is essential for early diagnosis. Currently, the diagnosis of thiamine deficiency in remote regions is hampered due to several drawbacks related to venous blood collection, for example, cold chain transport. Therefore, we here describe the first liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of thiamine diphosphate (TDP) in dried blood, using volumetric absorptive microsampling (VAMS). Moreover, by setting up an additional method in liquid blood, the results in VAMS samples could be compared to liquid blood samples. Both methods, employing a simple one-step extraction and fast (2 min) chromatography, were fully validated based on international guidelines. Accuracy (% bias) was below 6.5% for all QC levels. The total imprecision (% CV) was below 13% for both QCs and native blood samples. The recovery of the VAMS samples was not impacted by the hematocrit, within the hematocrit range of 0.20-0.60. Additionally, we showed improved TDP stability in dried blood compared to liquid blood. VAMS samples were stable for 1 week at 60 °C or at high humidity (80%) and for at least 1 month at room temperature. Finally, we demonstrated the commutability of commercial calibrators with authentic blood samples. The validity and applicability of both methods were demonstrated via their successful application on blood samples from healthy volunteers.

Pharmacokinetic study of benfotiamine and the bioavailability assessment compared to thiamine hydrochloride.

Benfotiamine is a lipid-soluble thiamine precursor which can transform to thiamine in vivo and subsequently be metabolized to thiamine monophosphate (TMP) and thiamine diphosphate (TDP). This study investigated the pharmacokinetic profiles of thiamine and its phosphorylated metabolites after single- and multiple-dose administration of benfotiamine in healthy Chinese volunteers, and assessed the bioavailability of orally benfotiamine administration compared to thiamine hydrochloride. In addition, concentration of hippuric acid in urine which is produced in the transformation process of benfotiamine was determined. The results showed that thiamine and its phosphorylated metabolites exhibited different pharmacokinetic characteristics in plasma, blood and erythrocyte, and one-compartment model provided the best fit for pharmacokinetic profiles of thiamine. The transformation process of benfotiamine to thiamine produced large amount of hippuric acid. No accumulation of hippuric acid was observed after multiple-dose of benfotiamine. Compared to thiamine hydrochloride, the bioavailability of thiamine in plasma and TDP in erythrocyte after oral administration of benfotiamine were 1147.3 ± 490.3% and 195.8 ± 33.8%, respectively. The absorption rate and extent of benfotiamine systemic availability of thiamine were significantly increased indicating higher bioavailability of thiamine from oral dose of benfotiamine compared to oral dose of thiamine hydrochloride.

Serial evaluation of children with ALF with advanced MRI, serum proinflammatory cytokines, thiamine, and cognition assessment.

OBJECTIVES: This prospective, sequential study was done to understand changes in cerebral edema (CE) on magnetic resonance imaging and magnetic resonance spectroscopy, liver functions, and neurocognitive testing (NCT) in children with acute liver failure (ALF). METHODS: A total of 11 ALF and 8 healthy controls were evaluated with advanced magnetic resonance (MR) imaging, blood proinflammatory cytokines (PCs), thiamine levels, liver functions, and NCT. Reevaluation was done at 43.5 ±â€Š26.9 days (first follow-up, n = 8) and 157.3 ±â€Š52.3 days (second follow-up, n = 6) after discharge. RESULTS: At diagnosis, patients with ALF had vasogenic and cytotoxic CE, raised brain glutamine (23.2 ±â€Š3.4 vs. 15.3 ±â€Š2.7), and serum PCs (tumor necrosis factor [TNF]-α 40.1 ±â€Š8.9 vs. 7.2 ±â€Š2.7  pg/mL, interleukin [IL]-6 29.2 ±â€Š14.4 vs. 4.7 ±â€Š1.2  pg/mL). The mammillary bodies (MBs) were smaller, and brain choline (1.9 ±â€Š0.36 vs. 2.6 ±â€Š0.6) and blood thiamine (55.2 ±â€Š6.7 vs. 81.8 ±â€Š10.2  nmol/L) were lower than controls. At first follow-up, the brain glutamine and CE recovered. Brain choline and MBs volume showed improvement and thiamine levels normalized. Significant reduction in TNF-α and IL-6 was seen. The patients performed poorly on NCT, which normalized at second follow-up. Liver biochemistry and thiamine levels were normal and TNF-α and IL-6 showed further reduction at second follow-up. CONCLUSIONS: Patients with ALF have CE contributed by raised brain glutamine and PCs. MBs are small because of thiamine deficiency and show recovery in follow-up. CE and brain glutamine recover earlier than normalization of NCT and liver functions. Persistence of raised cytokines up to 6 months after insult suggests possible contribution from liver regeneration.

Assessment of thiamin (vitamin B1) and riboflavin (vitamin B2) status in an adult Mediterranean population.

The aim of the present study was to assess the nutritional status for thiamin (vitamin B1) and riboflavin (vitamin B2) in an adult Mediterranean population, in order to identify patterns of intake, groups at risk for deficiency and factors that might influence this risk. A cross-sectional survey was carried out in Andalusia, a western Mediterranean region in southern Spain. Nutrient intakes were studied in a random sample of 3390 subjects (1746 men, 1644 women) who were between 25 and 60 years of age. Blood samples were obtained for biochemical assays in a random subsample of 372 subjects (181 men, 191 women). Food consumption was assessed by 48 h recall. Vitamin B1 and B2 were measured as erythrocyte transketolase and as erythrocyte glutathione reductase activation coefficients, respectively. Energy and vitamin intakes were significantly higher in men than in women. Intakes were below two-thirds of the recommended dietary allowance for vitamin B1 in 7.80 % of the men and 4.50 % of the woman, and were below this level for vitamin B2 in 18.00 % of the men and 11.70 % of the women. Age, educational level, alcohol use and smoking were also associated with differences in the intake of these nutrients. Biochemical analyses showed that vitamin B1 and B2 status was deficient in 6.40 and 5.30 % of the population, respectively. Although factors such as gender, age, level of education, drinking and smoking can have an effect on the risk of inadequate intake of these nutrients, these factors did not affect biochemical indices of nutritional status in the present study.

Simultaneous liquid chromatographic assessment of thiamine, thiamine monophosphate and thiamine diphosphate in human erythrocytes: a study on alcoholics.

An isocratic HPLC procedure for the assessment of thiamine (T), thiamine monophosphate (TMP) and thiamine diphosphate (TDP) in human erythrocytes is described. Several aspects of the procedure make it suitable for both clinical and research purposes: limits of detection and quantification of 1 and 2.5 nmol/l, respectively, recovery of 102% on average (range 93-112%), intra- and inter-day precisions within 5 and 9%, respectively, total elution time 15 min. This analytical methodology was applied to a case-control study on erythrocyte samples from 103 healthy subjects and 36 alcohol-dependent patients at risk of thiamine deficiency. Mean control values obtained were: T=89.6+/-22.7 nmol/l, TMP=4.4+/-6.6 nmol/l and TDP=222.23+/-56.3 nmol/l. T and TDP mean values of alcoholics were significantly lower than those of control cases: T=69.4+/-35.9 nmol/l (P<0.001) and TDP=127.4+/-62.5 nmol/l (P<10(-5)). The diagnostic role of TDP was evaluated and a significant role for thiamine was established in the study of alcohol related problems.

Assessment of thiamin status in chronic renal failure patients, transplant recipients and hemodialysis patients receiving a multivitamin supplementation.

The thiamin status of patients with chronic renal failure (CRF, n = 14), dialysis patients (DP, n = 24) and patients after renal transplantation (RT, n = 19) was assessed. Thiamin intake was calculated at mean levels of 1.26 mg/d (CRF), 0.83 mg/d (DP) and 1.42 mg/d (RT). Corresponding mean plasma concentrations were 64.2 nmol/l (CRF), 78.3 nmol/l (DP) and 55.1 nmol/l (RT). Thiamin supplements of 1.5 mg or 8.0 mg orally given to patients of the DP-group after each dialysis session showed slightly higher thiamin concentrations in plasma. Transketolase activity coefficients (ETK-AC) were in the same range (1.11...1.19) except for RT-patients who had a slightly but not significantly higher ETK-AC of 1.22. During dialysis treatment (DT), thiamin plasma concentrations dropped to 75 and/or 82% in patients supplemented with 1.5 and/or 8.0 mg. They both reached initial levels again 44 hours later. Despite large inter-individual differences, thiamin concentrations increased in the non-supplemented DP-group. ETK-AC did not change after a 14-day interruption of supplementation and did not deteriorate after a single dialysis session, both in supplemented and non-supplemented patients. A daily thiamin supplementation which complies with the RDA for healthy subjects is indicated in DP and is sufficient to keep thiamin status within the normal range.

Improved measurement of transketolase activity in the assessment of "TPP effect".

The indirect estimation of thiamine levels in human blood by measuring thiamine pyrophosphate effect on erythrocyte transketolase activity is the method of choice in most clinical laboratories. We describe here an optimized, time-saving, and accurate method to determine the thiamine pyrophosphate effect in as many as 16 blood samples simultaneously. The method is based on a multi-point determination using a computer remote-controlled microplate reader. For multiple sample handling, three pooled reaction mixtures are freshly prepared and loaded onto a 96 well microtitre plate. A pre-written software is then initiated to remote-control the system. The data is retrieved and processed to calculate thiamine pyrophosphate effect by a self-written "macro" on a "Quattro-Pro" worksheet database. This method proves to be highly accurate (coefficient of variance: 2.7%), reproducible (coefficient of variance: 4.1%) and economical.

Vitamin B1, B2 and B6 during the course of pregnancy of rural and urban women in northeast Thailand.

The vitamin B1, B2 and B6 status was investigated in a group of rural and urban pregnant women in the northeast of Thailand during the course of pregnancy. The vitamin status of all three vitamins did not change during the course of pregnancy even for the group of rural women. Differences between the group of village women and those females from a high socio-economic urban fraction of the population were only observed for the vitamin B2 status. The B2-dependent enzyme activation coefficient varies between 46.5 up to 56.7% for the rural and between 8.9 to 18.5% for the urban women. Vitamin B6 deficiencies as measured by the activation coefficient varied above 30% up to 40% and more for both groups of women. A functional significance could only be established for vitamin B2 deficiency. The increase of the activation coefficient correlated with an increase in the serum concentration levels of transferrin. Transferrin in turn was inversely correlated with haemoglobin.

Bioavailability assessment of the lipophilic benfotiamine as compared to a water-soluble thiamin derivative.

The bioequivalence of thiamin in 2 therapeutically used preparations was tested in 10 healthy young men. Thiamin was orally administered either as lipophilic benfotiamine or as water-soluble thiamin mononitrate. Biokinetic data, measured as area under the curve and maximal concentration in plasma and hemolysate after ingestion, demonstrated a significantly improved bioavailability from the lipophilic derivative despite an ingested dose of only 40% as compared with the water-soluble salt. A superior cellular efficacy of benfotiamine was also concluded from the short-term stimulation of the thiamin-dependent transketolase activity in erythrocytes.

Clinical chemical methods for the routine assessment of the vitamin status in human populations. Part III: The apoenzyme stimulation tests for vitamin B1, B2 and B6 adapted to the Cobas-Bio analyzer.

The programming of the Cobas-Bio centrifugal analyzer for kinetic tests of transketolase, glutathione reductase and aspartate amino-transferase is described. The results obtained in a population of 200 healthy people of both sexes are reported.

Thiamine status in urban adolescents: effects of race.

To evaluate thiamine status in an urban adolescent population, we performed two investigations. In Study I, we compared whole blood thiamine levels in 101 healthy adolescents from varied racial backgrounds with those that had been obtained previously in 146 healthy white adults from a different geographic locale. Blood thiamine values were significantly lower in the adolescents as a group, but the differences were entirely due to the lower levels in the black adolescents. To explore further these differences (Study II), we compared thiamine status in 34 adolescents with that of their parents using measures of both whole blood thiamine content and of erythrocyte transketolase activity. White adolescents had significantly higher total whole blood thiamine values than black adolescents, and white parents had significantly higher thiamine values than black parents by both total whole blood assay and level of transketolase activity. There were no differences in thiamine status between adolescents and parents of the same race. Racial composition is an important variable to consider in population surveys of thiamine status.

Assessment of vitamin B1, B2 and B6 status by coenzyme activation of red cell enzymes using a centrifugal analyser.

We describe a fully automated method for the assessment of vitamin B1, B2 and B6 status using a centrifugal analyser. The activation of the red cell enzymes transketolase, glutathione reductase and aspartate aminotransferase) by their respective coenzymes were measured in freshly prepared haemolysate. The enzyme catalytic activities in the sample were measured with (maximal activity) and without (basal activity) the coenzyme, and the percentage activation was calculated. The between run precision for red cell transketolase, glutathione reductase and aspartate aminotransferase were 8.5%, 10.3% and 9.5% respectively. When whole blood was stored at room temperature for 6 hours, red cell aspartate aminotransferase activity significantly decreased (n = 10, p less than 0.05). There were no significant changes in the activities of the other two enzymes. For a group of 30 healthy young subjects, the mean (standard deviation) values for the percentage activation of transketolase, glutathione reductase and aspartate aminotransferase were 11.9% (7.3), 35.1% (19.1) and 85.3% (18.0), respectively. The vitamin status of a group of 86 pregnant women was assessed by this method; 2.3%, 8.1% and 8.1%, respectively, of the pregnant women showed a higher percentage activation for transketolase, glutathione reductase and aspartate aminotransferase than that found in the young subjects. Both groups correlated well with respect to the basal activity and the percentage activation of each enzyme. Basal activity was inversely proportional to the percentage activation. It is therefore suggested that the basal activity can be used as a second criterion in the assessment of vitamin status.

Adaptation of coenzyme stimulation assays for the nutritional assessment of vitamins B1, B2 and B6 using the Cobas Bio centrifugal analyser.

Adaptation of coenzyme stimulation assays for the nutritional assessment of thiamine, riboflavin and pyridoxine on the Cobas Bio centrifugal analyser are described. Whole blood was collected into acid-citrate dextrose, which preserves the erythrocytes, prior to assay for several days. Washed erythrocytes stored at -70 degrees C and subsequently thawed, showed altered enzyme activities. The methods offer improved precision over existing procedures and take advantage of the high throughput capabilities of the instrumentation.

Assessment of the thiamine nutritional status. An evaluation of erythrocyte transketolase activity, the stimulated erythrocyte transketolase activity, and the thiamine pyrophosphate effect.

The most widely accepted approach to estimation of thiamine nutrition has been the measurement of the erythrocyte transketolase activity (ETKA), the ETKA stimulated in vitro with thiamine pyrophosphate (TPP) (which is suggested to be designated ETKAS) and the percentage increase of ETKA after stimulation with TPP in vitro, called the thiamine pyrophosphate effect (TPPE). In spite of 30 years of experience in the determination of these variables doubtfulness still exists of how to assess them. On the basis of a study of a group of alcoholics and a group of non-alcoholics a system of evaluation of ETKA, ETKAS and TPPE is proposed.

Clinical chemical methods for the routine assessment of the vitamin status in human populations. Part II: The water-soluble vitamins B1, B2 and B6.

Simple and proven techniques for the assay of the coenzyme stimulation of the erythrocyte enzymes transketolase, glutathione reductase and glutamate oxaloacetate aminotransferase are described. The results found in a cohort of 150 blood donors and their comparison with the values for total thiamine in whole blood and pyridoxal phosphate in erythrocytes are presented.