Study on the changes in antioxidant and blood sugar-lowering activities of fermented small lentils

Due to the high protein, vitamin, mineral, carbohydrate content and low fat content of lentils, they are increasingly attracting the attention of nutritionists. Additionally, a study indicates that lentil extracts can promote the growth of bifidobacteria, suggesting that lentils have great potential in developing fermented foods that protect the intestinal barrier and improve health. Many studies have shown that lentil phenolic compounds interfere with α-glucosidase and lipid digestion, achieving glucose and lipid homeostasis, while lactic acid fermentation can effectively improve the release of phenolic substances, enhancing the antioxidant and blood sugar-lowering effects of the fermentation products. Antioxidant capacity is usually represented by DPPH free radical scavenging ability and oxygen radical absorbance capacity.

This study used Lactobacillus plantarum TK9, Bacillus subtilis natto, and Lactobacillus paracasei TK1501 to evaluate the changes in antioxidant and blood sugar-lowering activities of lentils under different fermentation modes, aiming to produce products with potential health benefits.

1. Antioxidant activity of lentil extracts under different fermentation methods

It has long been known that antioxidants such as isoflavones and polyphenols in fermented legumes can absorb DPPH free radicals. IC50 is the half-inhibitory concentration, and a lower IC50 value indicates better DPPH absorption. The antioxidant capacity index ORAC is one of the most commonly used methods to assess antioxidant capacity.

As shown in Table 1, after anaerobic or aerobic fermentation with Lactobacillus plantarum TK9 and its co-culture with Bacillus subtilis natto, the IC50 value of DPPH in lentils decreased to varying degrees in both liquid and solid-state fermentation, while ORAC increased at the same time. Therefore, the antioxidant capacity of the anaerobic liquid fermentation product using Lactobacillus plantarum TK9 and Bacillus subtilis natto is the highest. Nevertheless, the products of anaerobic liquid fermentation consistently exhibit lower IC50 and higher ORAC, indicating that anaerobic liquid fermentation is more stable in terms of antioxidant activity. In contrast, the antioxidant activity of the mixed solid-state fermentation product under aerobic conditions is the lowest. When comparing the control group and aerobic mixed probiotics SSF (Table 1), it was found that a high concentration of Bacillus subtilis natto is detrimental to the synthesis of antioxidant substances.

In summary, the results confirm that fermentation is an effective method to enhance the antioxidant content of fermented lentil products due to changes in functional components such as phenolic substances, and co-cultivation improves the overall quality of the final product. Since Bacillus subtilis natto consumes oxygen and produces proteases, it can promote the growth and metabolism of anaerobic probiotics lactic acid bacteria. Even if Bacillus subtilis natto does not have a direct impact on the production of antioxidant substances, its small presence can promote the growth of lactic acid bacteria by altering the co-fermentation environment, thereby increasing the antioxidant content of the final product.

2. α-Glucosidase inhibitory activity of lentil extracts under different fermentation methods

α-Glucosidase is a key enzyme in starch digestion, and inhibiting starch-digesting enzymes can inhibit glucose transport proteins, reduce glucose release rates, and even prevent type II diabetes. Lentils and other legumes contain polyphenols, flavonoids, vitamins, polysaccharides, amino acids, and other substances that can lower blood sugar levels. Changes in the content of polyphenols, flavonoids, and amino acids in lentils after fermentation may have functional consequences. Therefore, the inhibition of α-glucosidase activity is used as a representative of the potential blood sugar-lowering and anti-diabetic effects of fermented lentils.

Figure 1 α-Glucosidase inhibitory activity of lentil extracts at different concentrations: (A) Liquid fermentation of single Lactobacillus plantarum TK9 or co-cultured with Bacillus subtilis natto; (B) Solid-state fermentation of single Lactobacillus plantarum TK9 or co-cultured with Bacillus subtilis natto.

As shown in Figure 1, with the increase in extract concentration, the inhibitory effect on α-glucosidase activity also increased, thus the inhibitory activity of liquid fermentation or solid-state fermentation samples was higher than that of the low concentration control group. At low extract concentrations (less than 10 mg/mL), the α-glucosidase inhibitory activity increased with the increase in extract concentration, although the change in inhibitory activity was small. For lentil extracts from solid-state fermentation (Figure 1B), the inhibition of α-glucosidase activity at low concentrations showed a weak correlation with the increase in concentration. As shown in Figure 1A, the α-glucosidase inhibitory activity in mixed bacterial liquid fermentation increased more at lower extract concentrations.

We further calculated the IC50 value using the relationship between α-glucosidase inhibitory activity and extract concentration, which is more conducive to comparing the inhibitory activities among samples.

As shown in Table 2, the IC50 of liquid samples before fermentation is higher than that of solid samples. Therefore, solid-state fermented lentil samples have better inhibitory activity against α-glucosidase. Thus, due to the mechanical release of degrading enzymes from cells, temperature, or a combination of various factors, some functional components may be destroyed during sample preparation, indicating that solid-state fermentation is more suitable for retaining α-glucosidase inhibitory activity. After fermentation, the IC50 value of liquid fermentation decreased significantly, while the lowest IC50 value was measured in samples co-fermented with Lactobacillus plantarum TK9 and Bacillus subtilis natto in aerobic bottles. Solid-state fermentation showed lower IC50 values under aerobic single fermentation and anaerobic co-fermentation conditions, with IC50 values of 3.955 and 3.985 mg/mL obtained from co-fermentation with Lactobacillus paracasei and Bacillus subtilis natto, respectively. Therefore, we suspect that the products of Lactobacillus plantarum TK9 have a strong ability to inhibit α-glucosidase activity when using Bacillus subtilis natto in anaerobic co-fermentation, and that Lactobacillus paracasei TK1501 can achieve the same effect in solid-state fermentation when the colony count of Bacillus subtilis natto is low. Previous studies have found that flavonols are the only compounds among various phenolics that can inhibit α-glucosidase activity. Based on this, the results of different α-glucosidase inhibitory activities we studied may be related to the changes in flavonols.

3. Conclusion

This study investigated the changes in antioxidant capacity and blood glucose-lowering activity of lentils after fermentation, revealing significant benefits of anaerobic solid-state co-fermentation. In addition to anaerobic solid-state fermentation, co-fermentation exhibited excellent DPPH free radical scavenging ability and ORAC. Furthermore, the lowest IC50 value for α-glucosidase inhibition activity was obtained in anaerobic solid-state co-fermentation, indicating that the corresponding products may be beneficial for patients with type II diabetes. Considering the processing costs before and after liquid fermentation, anaerobic solid-state fermentation may be more suitable for commercial production. Therefore, using lactic acid bacteria and low concentrations of Bacillus subtilis natto for anaerobic solid-state co-fermentation may provide an effective method for producing fermented foods with higher antioxidant capacity and anti-diabetic effects.

Reference: Co‐fermentation of lentils using lactic acid bacteria and Bacillus subtilis natto increases functional and antioxidant components[J]. Journal of Food Science, 2020.

Note: This article is for informational purposes only and is not intended as medical guidance.