Short-term stability of amino acids and acylcarnitines in the dried blood spots used to screen newborns for metabolic disorders.

OBJECTIVE: Tandem mass spectrometry is increasingly used in the Middle East in newborn screening for inborn errors of metabolism using dried blood spots. The sensitivity and specificity of this system for analyzing fatty and amino acids, screening for more than 40 metabolic conditions, is known. However, the short term stability of acylcarnitines and amino acids in dried blood spots in extreme heat and humid conditions is not well documented. We examined the short term effect of heat and humidity on the levels of 7 amino acids and 10 acylcarnitines used in newborn screening for inherited metabolic disorders. METHODS: Dried blood spots were exposed with humidity <30% to temperatures of 4, room temperature, 37° C, and 45, and also with humidity >70% at 37° C and 45. Amino acids and acylcarnitines in the dried blood spots were analyzed by tandem mass spectrometry. RESULTS: During the eight days of the study in high temperature and high humidity storage, most acylcarnitines and amino acids lost almost 50% of initial concentration. After eight days' exposure at 37 and 45 with humidity >70%, methionine was determined to be the most sensitive, and phenylalanine and leucine were the least sensitive amino acids. At 37 with humidity >70% C6 was the most sensitive and free carnitine (C0) was the least sensitive acylcarnitine; at 45 with humidity >70% C16 was the most sensitive and C0 was the least sensitive. CONCLUSION: Low humidity and low temperature conditions are required for transportation and storage of dried blood spots.

Selective newborn screening of inborn errors of amino acids, organic acids and fatty acids metabolism in the Kingdom of Bahrain.

Mandatory newborn screening for metabolic disorders has not been implemented in most Middle Eastern countries. Early detection and treatment of inborn errors of metabolism can reduce mortality and minimize morbidity. Preliminary studies conducted in some parts of Middle East suggest that the incidences of inborn errors of metabolism are reported to be higher in the region than anywhere else in the world due to the consanguinity. In this study the incidence of inborn errors of amino acids, organic acids and fatty acids oxidation disorders was investigated from the results of blood spot analysis of 1986 symptomatic children from 1st January 2008 to 31st of December 2011. Out of 1986 newborns screened 25 infants were diagnosed and confirmed with amino acids (n=11), organic acids (n=9) and fatty acids oxidation (n=5) disorders. Overall incidences based on number of live birth between 2008 and 2011 inclusive were 1:6000, 1:8000 and 1:14,000 for amino acids, organic acids and fatty acids oxidation disorders; respectively. Out of 25 infants diagnosed, 21 were the children of first cousin marriages. Results from this study suggest high incidence of inborn errors of amino acids, organic acids and fatty acids oxidation metabolism in Bahrain and significant contribution of consanguinity in inherited metabolic disorders. Mandatory screening for inborn errors of metabolism in Bahrain is highly recommended.

Association of red blood cell 5-methyltetrahydrofolate folate with bone mineral density in postmenopausal Iranian women.

Recent studies have suggested that hyperhomocysteinemia and low plasma folate are associated with fracture and also bone mineral density (BMD) and that they may contribute to the pathogenicity of osteoporosis in postmenopausal women. However, as plasma total homocysteine (tHcy) and plasma folate can be regarded as short-term markers when compared to a long-term variable such as BMD, in this study we tested the hypothesis that low red blood cell 5-methyltetrahydrofolate (RBC 5-MTHFR) as a long-term marker of the folate status may be a better predictor of BMD than plasma 5-MTHF, and its deficiency may contribute to the pathogenicity of osteoporosis in postmenopausal Iranian women. The BMD at the femoral neck and lumbar spine (measured by dual-energy X-ray absorptiometry, DXA) together with anthropometric and biochemical components of the homocysteine re-methylation pathway including plasma tHcy, 5-MTHF and vitamin B12, RBC 5-MTHF and creatinine were determined in 366 postmenopausal women. RBC 5-MTHF was more highly correlated with BMD at the lumbar spine (r=0.21, P=0.001) and femoral neck (r=0.19, P=0.004) than was plasma 5-MTHF (lumbar spine; r=0.14, P=0.03 and femoral neck; r=0.17, P=0.006). Stepwise multiple linear regression analyses revealed that RBC 5-MTHF was one of the predictors of BMD explaining 4.3 and 4.0% variance of BMD at the lumbar spine and femoral neck, respectively, whereas plasma 5-MTHF was excluded in the model and not determined to be a predictor of BMD at both the lumbar spine and femoral neck when adjusted for age, BMI, years since menopause and RBC 5-MTHF. This study suggests that RBC 5-MTHF is a better predictor of BMD than plasma 5-MTHFR when compared to a long-term marker such as BMD, and its deficiency is associated with low BMD that may contribute to the pathogenicity of osteoporosis in postmenopausal women.

Distribution of 5,10-methylenetetrahydrofolate reductase (C667T) polymorphism and its association with red blood cell 5-methyltetrahydrofolate in the healthy Iranians.

BACKGROUND AND AIMS: Homozygosity for the thermolabile variant of 5,10-methylenetetrahydrofolate reductase (C677T) that causes hyperhomocysteinemia has been reported in 5-15% of general populations. This mutation has also been suggested to be positively associated with the risk of vascular disease and neural tube defects. It has also been suggested that present dietary reference values may need to be altered for people heterozygote or homozygote for this mutation as tissue folate status has been reported to be compromised by these genetic variants. The aims of this study were to investigate the distribution of 5,10-methylenetetrahydrofolate reductase (C677T) polymorphism in a population of Shiraz, south west of Iran and to test the hypothesis that folate status is compromised by this mutation in our population. METHODS: In this study age, body mass index, plasma and red blood cell 5-methytetrahydrofolate, plasma total homocysteine and vitamin B12 of 391 healthy Iranians (198 men and 193 women) together with methylenetetrahydrofolate reductase C667T genotypes were determined. The correlates of methylenetetrahydrofolate reductase polymorphism were determined using univariate and multivariate statistical analysis. RESULTS: The frequencies of CC, CT and TT genotypes were 56.2%, 38.7% and 5.1%, respectively. The C and T allele frequencies were determined to be 0.76 and 0.24, respectively and this polymorphism was compatible with Hardy-Weinberg equilibrium (X2=1.54, df=2, P=0.46). Among all the variables examined, red blood cell 5-methyltetrahydrofolate (P=0.007, ANOVA) and plasma 5-methyltetrahydrofolate (P=0.012, ANOVA) were significantly lower in individuals with TT genotype than those with either CC or CT genotype. Plasma total homocysteine was significantly higher in individuals with TT than those with either CC or CT genotype at below the median levels of red blood cell 5-methylterahydrofolate (P=0.03, ANOVA) and plasma 5-methylterahydrofolate (P=0.04, ANOVA). Univariate (r=-0.16, P=0.002) and multivariate analysis (beta=-0.0005, P=0.003) showed that red blood cell 5-methylterahydrofolate was the strongest correlates of methylenetetrahydrofolate reductase genotypes. CONCLUSIONS: Results from this study suggest that methyltetrahydrofoate reductase C667T genotypes are strongly and independently associated with low red blood cell 5-methylterahydrofolate that has been reported to be a more reliable and long-term marker for body's folate status among Iranians. These results may suggest that substantial minority of people in general populations may have increased folate needs and this may place doubts on the validity of assuming "normality" for nutrient requirements in any general population.

Selective screening of amino acid disorders in the south-west of Iran, Shiraz.

A diagnostic study of amino acid concentrations in dried blood spot samples from 1044 symptomatic children revealed high incidences of phenylketonuria, tyrosinaemias, and maple syrup urine disease in the Shiraz region of Iran. Minimum incidences, based on babies born between 1996 and 2001 inclusive, and diagnosed by the end of 2001, are 1:3672, 1:10651 and 1:21 303, respectively.