Kale (Brassica oleracea var. acephala) is a vegetable common in most cultures but is less studied as a functional food compared to other cruciferous vegetables, such as broccoli. We investigated the effect of supplementing a high-fat diet (HFD) with kale (HFKV) in C57BL/6J mice. We particularly explored its role in metabolic parameters, gut bacterial composition and diversity using 16S rRNA sequencing, systematically compared changes under each phylum and predicted the functional potential of the altered bacterial community using PICRUSt2. Like other cruciferous vegetables, kale attenuated HFD-induced inflammation. In addition, kale modulated HFD-induced changes in cecal microbiota composition. The HFD lowered bacterial diversity, increased the Firmicutes: Bacteroidetes (F/B) ratio and altered composition. Specifically, it lowered Actinobacteria and Bacteroidetes (Bacteroidia, Rikenellaceae and Prevotellaceae) but increased Firmicutes (mainly class Bacilli). Kale supplementation lowered the F/B ratio, increased both alpha and beta diversity and reduced class Bacilli and Erysipelotrichi but had no effect on Clostridia. Within Actinobacteria, HFKV particularly increased Coriobacteriales/Coriobacteriaceae about four-fold compared to the HFD (p < 0.05). Among Bacteroidia, HFKV increased the species Bacteroides thetaiotaomicron by over two-fold (p = 0.05) compared to the HFD. This species produces plant polysaccharide digesting enzymes. Compared to the HFD, kale supplementation enhanced several bacterial metabolic functions, including glycan degradation, thiamine metabolism and xenobiotic metabolism. Our findings provide evidence that kale is a functional food that modulates the microbiota and changes in inflammation phenotype.

Like humans, outbred Sprague-Dawley CD rats exhibit a polygenic pattern of inheritance of the obese phenotype and not all individuals exposed to a high calorie intake develop obesity. We hypothesized that differences in gut microbiota composition account for phenotype differences between obese prone (OP) and obese resistant (OR) rats. We studied the gut microbiota composition of OPand OR rats after a high fat (HF) diet and how they respond to fermentation of resistant starch (RS). In phase 1 of the study 28 OP and 28 OR rats were fed a HF diet. In order to determine causal role of microbiota on phenotypes, In phase 2, a microbiota transplant between the two phenotypes was performed before switching all rats to a HF diet supplemented with 20% RS. We determined microbiota composition by 16S sequencing and predicted microbiota function by PICRUSt2. Despite a similar calorie intake, in phase 2 OP rats gained more weight and accumulated more abdominal fat in both phase 1 and 2 compared to OR rats (P < 0.001; n = 6). The OP rats fermented RS more robustly compared with OR rats with an increase in total bacteria, short chain fatty acids, and increased weight of the cecum, but microbiota of OP rats had much lower alpha diversity and evenness. The microbiota of OP rats, had higher amounts of bacteria from order Bacteroidales, specifically family Muribaculaceae (S24-7), which is known to possess several starch degrading enzymes and was reported in previous studies to increase with fermentation of RS. The OR rats fermented RS less but had higher bacterial diversity and evenness and had significantly higher bacterial counts from phylum Firmicutes particularly order Clostridiales, genus Clostridium and an uncultured bacterium of the genus Akkermansia. The microbiota of OR rats had enhanced bacterial chemotaxis, phosphotransferase system (PTS), and fatty acid biosynthesis compared to OP rats whose microbiota had higher glycan degradation and LPS biosynthesis pathways. The microbiota transplant did not change obesity phenotype or microbiota composition. In conclusion, a higher alpha-diversity and evenness of the microbiota and higher proportions of Clostridiales and Akkermansia in OR rats were associated with a better metabolic phenotype with lower body fat. However, robust RS fermentation caused a lower diversity and evenness and did not result in a leaner phenotype.

The properties of kale as a functional food are well established. We sought to determine how fermentation further enhances these properties. We tested different fermentation conditions: (i) spontaneous fermentation with naturally occurring bacteria, (ii) spontaneous fermentation with 2% salt, (iii) Lactococcus lactis , (iv) Lactobacillus acidophilus , (v) mixture of L. lactis and L. acidophilus , (vi) mixture of L. lactis , L. acidophilus , and Clostridium butyricum. We quantified selected bioactive components using high‐performance liquid chromatography (HPLC) and antinutritional factors using a gravimetric method and spectrophotometry. We then determined (i) the antioxidant capacity of the vegetable, (ii) anti‐inflammation capacity, and (iii) the surface microbiota composition by 16S sequencing. All fermentation methods imparted some benefits. However, fermentation with mixed culture of L. lactis and L. acidophilus was most effective in increasing polyphenols and sulforaphane accessibility, increasing antioxidant activity, and reducing antinutritional factors. Specifically, fermentation with L. lactis and L. acidophilus increased total polyphenols from 8.5 to 10.7 mgGAE/g (milligrams of gallium acid equivalent per gram) and sulforaphane from 960.8 to 1777 μg/g (microgram per gram) but decreased the antinutritional factors oxalate and tannin. Total oxalate was reduced by 49%, while tannin was reduced by 55%–65%. The antioxidant capacity was enhanced but not the anti‐inflammation potential. Both unfermented and fermented kale protected equally against lipopolysaccharide (LPS)‐induced inflammation in RAW 264.7 macrophages and prevented increases in inducible nitric oxide synthase (iNOS), tumor necrosis factor‐alpha (TNF‐α), interleukin‐1 beta (IL‐1β), and interleukin‐6 messenger RNA (IL‐6 mRNA) expression by 84.3%, 62%, 68%, and 85.5%, respectively. Unfermented and naturally fermented kale had high proportions of sulfur reducing Desulfubrio and Proteobacteria usually associated with inflammation. Fermenting with L. lactis and/or L. acidophilus changed the bacterial proportions, reducing the Proteobacteria while increasing the genera Lactobacilli and Lactococcus. In summary, fermentation enhances the well‐known beneficial impacts of kale. Fermentation with mixed cultures of L. lactis and L. acidophilus imparts higher benefits compared to the single cultures or fermentation with native bacteria present in the vegetable.