Abstract
ObjectiveThe study aims at evaluating mid-infrared human milk analyzer (HMA) accuracy and precision, in human milk (HM).Study designRöse-Gottlieb, high-performance anion exchange chromatography-pulsed amperometric detection (HPAEC-PAD), Kjeldahl and amino acid analysis (AA) were selected as references for total fat, lactose and total protein determination.ResultsNo significant difference was observed in lactose content between HMA and HPAEC-PAD. Significant differences were observed in fat and protein content between HMA and reference methods. However, the difference in fat content was lower than 12%, and therefore within the variability declared by supplier. For protein determination, the BCA protein assay was selected. No significant differences were observed in total protein content measured by BCA assay, Kjeldahl and AA methods.ConclusionsHMA was reliable for the quantification of total fat and lactose content, but not for total protein one. The latter was measured by BCA assay, which yielded comparable results to Kjeldahl and AA methods.
Highlights
Human milk (HM) is recognized as the normative standard of nutrition for infants for the first 6 months of life
HM samples (n = 150) were analyzed with the human milk analyzer (HMA) and bicinchoninic acid (BCA) kit for their total protein content (Fig. 3) The average protein content in HM measured with the HMA varied from 1.10 ± 0.22 g 100 mL−1 to 0.83 ± 0.30 g 100 mL−1 and from 1.5 to 1.3 g 100 mL−1 with the BCA assay
Our results for all macronutrients matched the values and trajectories previously reported in literature [21]. The results of this investigation demonstrate that the MIRIS HMA is suitable for the quantification of total fat and lactose in HM, with regard to trueness and precision
Summary
The total protein content of the HM samples was determined using the bicinchoninic acid (BCA) Protein Assay kit (Pierce® BCA protein Assay Kit, http://www.piercenet.com/ instructions/2161296.pdf, Thermo Fisher) according to the instructions provided with the kit. The average lipid content in HM varied from 4.17 ± 0.92 g 100 mL−1 (at 30 days) to 4.65 ± 2.10 g 100 mL−1 (at 120 days), respectively [14] These samples were analyzed for their content in total fatty acids [14] and correlation between total fatty acids measured by gas chromatography and total fat determined by HMA evaluated. In order to evaluate the accuracy of the HMA for the quantification of total protein content, different aliquots of pooled HM samples were analyzed 12 times in duplicate by the HMA (n = 24) and 6 times in duplicate by the Kjeldahl and BCA method (n = 12) (Table S3 Supplementary Material).
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