Vitamin Concentrations in Human Milk Vary with Time within Feed, Circadian Rhythm, and Single-Dose Supplementation

Abstract

Background: Human milk is the subject of many studies, but procedures for representative sample collection have not been established. Our improved methods for milk micronutrient analysis now enable systematic study of factors that affect its concentrations.Objective: We evaluated the effects of sample collection protocols, variations in circadian rhythms, subject variability, and acute maternal micronutrient supplementation on milk vitamin concentrations.Methods: In the BMQ (Breast-Milk-Quality) study, we recruited 18 healthy women (aged 18-26 y) in Dhaka, Bangladesh, at 2-4 mo of lactation for a 3-d supplementation study. On day 1, no supplements were given; on days 2 and 3, participants consumed ∼1 time and 2 times, respectively, the US-Canadian Recommended Dietary Allowances for vitamins at breakfast (0800-0859). Milk was collected during every feeding from the same breast over 24 h. Milk expressed in the first 2 min (aliquot I) was collected separately from the remainder (aliquot II); a third aliquot (aliquot III) was saved by combining aliquots I and II. Thiamin, riboflavin, niacin, and vitamins B-6, B-12, A, and E and fat were measured in each sample.Results: Significant but small differences (14-18%) between aliquots were found for all vitamins except for vitamins B-6 and B-12. Circadian variance was significant except for fat-adjusted vitamins A and E, with a higher contribution to total variance with supplementation. Between-subject variability accounted for most of the total variance. Afternoon and evening samples best reflected daily vitamin concentrations for all study days. Acute supplementation effects were found for thiamin, riboflavin, and vitamins B-6 and A at 2-4 h postdosing, with 0.1-6.17% passing into milk. Supplementation was reflected in fasting, 24-h postdose samples for riboflavin and vitamin B-6. Maximum amounts of dose-responding vitamins in 1 feeding ranged from 4.7% to 21.8% (day 2) and 8.2% to 35.0% (day 3) of Adequate Intake.Conclusions: In the milk of Bangladeshi mothers, differences in vitamin concentrations between aliquots within feedings and by circadian variance were significant but small. Afternoon and evening collection provided the most-representative samples. Supplementation acutely affects some breast-milk micronutrient concentrations. This trial was registered at clinicaltrials.gov as NCT02756026.

Keywords: acute supplementation effects; circadian variation; human milk; lactation; sample collection; vitamins.

Conflict of interest statement

Author disclosures: D Hampel, S Shahab-Ferdows, MM Islam, JM Peerson, and LH Allen, no conflicts of interest.

Figures

FIGURE 1

Flowchart of study participants in the Breast-Milk-Quality Study. Eighteen apparently healthy Bangladeshi women were enrolled in the study. 1Three aliquots of breast milk were collected from the same breast during every feeding for the duration of the study (I: the first 2 min into the feeding; II: the remainder of the feeding; III: full-breast aliquot obtained by combining I and II). BM, breast milk.

FIGURE 2

Differences between milk aliquots collected within a feeding during a day without maternal supplementation. Values are medians (95% CIs); n = 187 from 18 apparently healthy Bangladeshi mothers at 2–4 mo of lactation. Labeled medians without a common letter differ, P < 0.05. I1, first 2 min of the feeding; II, remainder of the feeding; III, full-breast aliquot obtained by combining I and II. General linear models were used to explore the differences in concentration between aliquots on day 1. All concentrations are μg/L except for B12 (ng/L), Afat and Efat (nmol/L), Fat (g/L), A (μg/100 mL), and E (μg/10 mL). A, vitamin A; Afat, vitamin A adjusted for milk fat; B1, thiamin; B2, riboflavin; B3, niacin (nicotinamide); B6, vitamin B-6; B12, vitamin B-12; E, vitamin E; Efat, vitamin E adjusted for milk fat; Fat, milk fat.

FIGURE 3

Relative daily concentrations of vitamins and fat in breast milk during maternal supplement consumption on days 2 (1 capsule) and 3 (2 capsules). Values are medians (95% CIs); day 1, n = 187; day 2, n = 179; day 3, n = 173 from 18 women. Labeled medians without a common letter differ, P < 0.05. Relative median concentrations (%) are based on median concentrations on day 1 (set to 100% by default) and median concentrations on days 2 and 3. Only the full-breast aliquots (aliquot III) were considered. The mixed-model analysis was used to examine for differences in concentrations by day. A, vitamin A; Afat, vitamin A adjusted for milk fat; B1, thiamin; B2, riboflavin; B3, niacin (nicotinamide); B6, vitamin B-6; B12, vitamin B-12; E, vitamin E; Efat, vitamin E adjusted for milk fat; Fat, milk fat.