This designed by our research group, whereby
This cross-sectional analysis is of baseline data from a randomised controlled dietary intervention trial designed to identify the maternal vitamin D intake in pregnancy that would maintain serum 25(OH)D in late gestation at a concentration sufficient to keep umbilical cord 25(OH)D ?25-30 nmol/L 14. The study was a three-arm, parallel, dose-response, double-blind, placebo-controlled randomised trial of vitamin D3 supplementation. The power calculation was based on similar dose-response studies of vitamin D nutritional requirements designed by our research group, whereby 31 participants per arm is adequate to detect a 10 nmol/L difference in 25(OH)D concentrations and provide a 90% power to demonstrate a dose-response relation with slope 1.
5 and alpha equal to 0.05 26-29. This study was conducted throughout the year because it was a pregnancy study, with three assessment points across gestation, unlike our previous trials in non-pregnant groups which were conducted during winter time.
To enable a season-specific analysis and account for a potentially higher dropout rate in late gestation than we would usually see, the sample size was increased to 48 per arm (144 in total). From enrolment, at 10 ?g/d) or calcium (>650 mg/d) prior to randomisation. Study visits took place at the Human Nutrition Studies Unit at the Cork Centre for Vitamin D and Nutrition Research, University College Cork, Cork, Ireland, with the baseline visit at mean ± SD of 14 ± 2 weeks’ gestation. At baseline, trained researchers collected information on general health, lifestyle and, socio-demographics. Habitual calcium and vitamin D intakes were estimated using a specifically designed and validated interviewer-administered quantitative food frequency questionnaire 30. The food frequency questionnaire was interviewer administered to improve precision and the validity coefficient for vitamin D intakes was 0.
92 (95% CI; 0.80–0.97) 30. Height and weight were measured for calculation of BMI (Leicester height measure, CMS Weighing Equipment Ltd., London, UK; digital weighing scales, SECA Ltd., Birmingham, UK).
Non-fasting blood samples were collected, processed to serum and stored at -80°C within 3 hours of collection. The study was conducted in accordance with the Declaration of Helsinki guidelines and ethical approval was obtained from the Clinical Research Ethics Committee of the Cork Teaching Hospitals ECM4(o)04/02/14. The trial is registered at the United States National Institutes of Health Clinical trials Registry (www.clinicaltrials.
gov), ID: NCT02506439. Written informed consent was provided by all participants prior to study commencement. 2.2. Biochemical Analysis 2.2.1.
Serum 25(OH)DMeasurement of 25(OH)D in our laboratory has been detailed previously 5. Briefly, individually quantified 25(OH)D2 and 25(OH)D3 were summed to calculate total 25(OH)D. Serum 25(OH)D2 and 25(OH)D3 concentrations were measured using liquid chromatography-tandem mass spectrometry (LC-MS/MS) on a Waters Acquity UPLC system coupled to an Acquity Triple Quadrupole TQD mass spectrometer detector (Waters, Milford, USA). Method validation used four levels of serum-based NIST (National Institute of Standards and Technology) certified quality assurance material (SRM 972) and quality control materials, purchased from Chromsystems (Munich, Germany), were assayed in parallel to all samples.
NIST calibrators (SRM 2972) were used throughout the analysis. The limit of detection (LoD) for 25(OH)D3 and 25(OH)D2 was 0.31 and 0.44 nmol/L, respectively. The limit of quantitation (LoQ) for 25(OH)D3 and 25(OH)D2 was 1.
03 and 1.43 nmol/L, respectively. The iIntra- and inter- assay coefficient of variations (CVs) for both metabolites were