Analysis of 26 amino acids in human plasma by HPLC using AQC as derivatizing agent and its application in metabolic laboratory.

The present study reports the simultaneous analysis of 26 physiological amino acids in plasma along with total cysteine and homocysteine by high-performance liquid chromatography (HPLC) employing 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) as precolumn derivatizing reagent. Separations were carried out using Lichrospher 100 RP-18e (5 µm) 250 × 4.0 mm column connected to 100 CN 4.0 × 4.0 mm guard column on a quaternary HPLC system and run time was 53 min. Linearity of the peak areas for different concentrations ranging from 2.5 to 100 pmol/µL of individual amino acids was determined. A good linearity (R (2) > 0.998) was achieved in the standard mixture for each amino acid. Recovery of amino acids incorporated at the time of derivatization ranged from 95 to 106 %. Using this method we have established the normative data of amino acids in plasma, the profile being comparable to the range reported in literature and identified cases of classical homocystinuria, cobalamin defect/deficiency, non-ketotic hyperglycinemia, hyperprolinemia, ketotic hyperglycinemia, urea cycle defect and maple syrup urine disease.

Hypothermia for moderate or severe neonatal encephalopathy in low-income and middle-income countries (HELIX): a randomised controlled trial in India, Sri Lanka, and Bangladesh.

BACKGROUND: Although therapeutic hypothermia reduces death or disability after neonatal encephalopathy in high-income countries, its safety and efficacy in low-income and middle-income countries is unclear. We aimed to examine whether therapeutic hypothermia alongside optimal supportive intensive care reduces death or moderate or severe disability after neonatal encephalopathy in south Asia. METHODS: We did a multicountry open-label, randomised controlled trial in seven tertiary neonatal intensive care units in India, Sri Lanka, and Bangladesh. We enrolled infants born at or after 36 weeks of gestation with moderate or severe neonatal encephalopathy and a need for continued resuscitation at 5 min of age or an Apgar score of less than 6 at 5 min of age (for babies born in a hospital), or both, or an absence of crying by 5 min of age (for babies born at home). Using a web-based randomisation system, we allocated infants into a group receiving whole body hypothermia (33·5°C) for 72 h using a servo-controlled cooling device, or to usual care (control group), within 6 h of birth. All recruiting sites had facilities for invasive ventilation, cardiovascular support, and access to 3 Tesla MRI scanners and spectroscopy. Masking of the intervention was not possible, but those involved in the magnetic resonance biomarker analysis and neurodevelopmental outcome assessments were masked to the allocation. The primary outcome was a combined endpoint of death or moderate or severe disability at 18-22 months, assessed by the Bayley Scales of Infant and Toddler Development (third edition) and a detailed neurological examination. Analysis was by intention to treat. This trial is registered with ClinicalTrials.gov, NCT02387385. FINDINGS: We screened 2296 infants between Aug 15, 2015, and Feb 15, 2019, of whom 576 infants were eligible for inclusion. After exclusions, we recruited 408 eligible infants and we assigned 202 to the hypothermia group and 206 to the control group. Primary outcome data were available for 195 (97%) of the 202 infants in the hypothermia group and 199 (97%) of the 206 control group infants. 98 (50%) infants in the hypothermia group and 94 (47%) infants in the control group died or had a moderate or severe disability (risk ratio 1·06; 95% CI 0·87-1·30; p=0·55). 84 infants (42%) in the hypothermia group and 63 (31%; p=0·022) infants in the control group died, of whom 72 (36%) and 49 (24%; p=0·0087) died during neonatal hospitalisation. Five serious adverse events were reported: three in the hypothermia group (one hospital readmission relating to pneumonia, one septic arthritis, and one suspected venous thrombosis), and two in the control group (one related to desaturations during MRI and other because of endotracheal tube displacement during transport for MRI). No adverse events were considered causally related to the study intervention. INTERPRETATION: Therapeutic hypothermia did not reduce the combined outcome of death or disability at 18 months after neonatal encephalopathy in low-income and middle-income countries, but significantly increased death alone. Therapeutic hypothermia should not be offered as treatment for neonatal encephalopathy in low-income and middle-income countries, even when tertiary neonatal intensive care facilities are available. FUNDING: National Institute for Health Research, Garfield Weston Foundation, and Bill & Melinda Gates Foundation. TRANSLATIONS: For the Hindi, Malayalam, Telugu, Kannada, Singhalese, Tamil, Marathi and Bangla translations of the abstract see Supplementary Materials section.

Improving Quality of Banked Milk: Utility of Dornic Acid Test.

OBJECTIVE: To identify, if Dornic acid test done on human milk bank sample is as effective as doing routine culture, both pre and post pasteurization. METHODS: The authors analyzed 477 samples, both pre and post pasteurization. Dornic acid measurement was performed by using N/9 NaOH and titrated to get dornicity of the sample. Senstivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) at different Dornic acid levels were calculated. To find diagnostic power of this study, ROC curve was prepared. Dornic acid for paired pre and post pasteurization were noted and analysed for statistical significance. RESULTS: A significant growth was seen in 21% (98/477) samples with coagulase negative staphylococci and gram negative organisms being the major contaminants seen prior to pasteurization. In the index study, Dornic acidity ≥4°D had a sensitivity of 98% but very low specificity of just over 10%. However, 8°D had 92% specificity and acceptable sensitivity of 72% in comparison to gold standard microbiological criteria, making it a good test for analyzing the quality of milk before pasteurization. CONCLUSIONS: Dornic acid test at 8°D has a very high specificity with acceptable sensitivity in comparison to dornicity at 4°D. It can be used as a simple method to select better quality of milk sample prior to pasteurization.