Background and objectives: Many studies indicate that the maternal diet is an important factor affecting human milk composition. Human milk composition among lactating women in Latvia, as well as the maternal diet during lactation, has not been sufficiently studied. The aim of this research was to assess dietary habits and macronutrient intake among lactating women in Latvia and to examine the effect of diet on human milk composition. Materials and Methods: Research was conducted between November 2016 and December 2017. Mature human milk samples (n = 61) along with a 72h food diary, a food frequency questionnaire (FFQ), and a questionnaire about maternal and infant characteristics were obtained from voluntary women who were recruited via an invitation published in a social media member group for nursing mothers. Fat content in human milk was determined by LVS ISO 2446:2008, protein content was determined by LVS EN ISO 8968-1:2014, lactose was determined by ISO 22662:2007, and the fatty acid profile was analyzed using gas chromatography. Dietary data were evaluated using the Finnish food composition database Fineli, release 19 (3 March 2018). Results: Median values for fat, protein, and lactose in mature human milk were 4.40%, 1.08%, and 6.52%, respectively. Predominant fatty acids in human milk were oleic acid (C18:1 n9c), palmitic acid (C16:0), and linoleic acid (C18:2 n6c) at 34.60%, 24.00%, and 11.00% of total fatty acids, respectively. The trans elaidic acid (C18:1 n9t) level was <0.10% in all human milk samples. Significant, positive associations (p < 0.05) were found between maternal dietary intake of linoleic, α-linolenic, docosahexaenoic, total cis-monounsaturated, total cis-polyunsaturated, and total n-6 and n-3 polyunsaturated fatty acids, the ratio of n-6/n-3, and the level of these fatty acids in human milk. Total energy and carbohydrate intake among participants were lower, but total fat, saturated fat, and sugar intake were higher than recommended. Protein, linoleic acid, and α-linolenic acid intake were adequate, but docosahexaenoic acid intake was noticeably lower than recommended. Women should be supported with information regarding their nutritional needs during lactation and the possible impact of diet on human milk composition. Conclusion: Macronutrient (fat, protein, and lactose) content in human milk is not affected by maternal diet. Conversely, the human milk fatty acid profile is affected by the immediate diet consumed by the mother. Habitual dietary habits can also impact the fatty acid profile of human milk.

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