Human berry intervention study: Microbial metabolites of berry phenolics from faecal samples

Controlled human intervention was carried out to explore the effects of diet rich in berries on subjects with metabolic syndrome. The subjects were randomly assigned in three diet groups: A) ellagitannin rich berries (cloudberry, strawberry, raspberry), B) anthocyanin rich berries (bilberry), and C) control. During the diet faeces samples were taken at five occasions: 1) four weeks before berry intervention, 2) one week before berry intervention, 3) when berries had been consumed for four weeks, 4) when berries had been consumed for seven weeks, and 5) four weeks after the berry intervention ended. The faeces were closed and stored in anaerobic atmosphere, cooled, and frozen at -70 °C in 4 h after defecation. For analysis faeces samples were melted slowly in ice - water bath, and metabolite analysis was carried out from 5% faecal suspension in physiological saline by GC-MS using selective ion mode (SIM) detection. Large amounts of phenolic compounds are likely to enter the colon, where they are metabolised by gut microflora. There is very little information of the bacterial transformations of some groups of phenolic compounds, such as anthocyanins. In order to increase knowledge of the metabolism of phenolic compounds, and the role of gut microflora in the overall metabolism, targeted analysis of the known faecal metabolites of phenolic compounds were carried out. The following 11 compounds were analyzed from faecal samples: 3-hydroxyphenylacetic acid (3-OHPAc), 3-phenylpropionic acid (3-PPr), benzoic acid (BA), 3,4-dihydroxybenzoic acids, 3,4-dihydroxyphenylacetic acid (3,4-diOHPAc), 3,4-dihydroxyphenylpropionic acids (3,4-diOHPPr), 2-hydroxyphenylpropionic acid (2-OHPPr), 3-hydroxyphenylpropionic acid (3-OHPPr), 4-hydroxyphenylpropionic acid (4-OHPPr), 3-hydroxybenzoic acid (3-OHBA) and 4-hydroxybenzoic acid (4-OHBA). Concentrations of these compounds were followed as a course of time of intervention. Statistical evaluation of the results was performed using paired Student's t-test. The comparison showed that ellagitannin rich diet significantly increased concentration of 3,4-dihydroxybenzoic acid (3,4-diOHBA) (p< 0,001) and 3-hydroxybenzoic acid (3-OHBA) (p< 0,001) compared to period before berry diet. Anthocyanin rich diet significantly increased concentration of 3,4-dihydroxybenzoic acid (3,4-diOHBA) (p< 0,001) and 3-hydroxyphenylacetic acid (3-OHPAc) (p< 0,01). The results verified the earlier in vitro data of the metabolism of phenolic compounds.