Preparation of safflower fermentation solution and study on its biological activity

Abstract

Introduction: Safflower, a traditional Chinese medicine, is rich in chemical components including flavonoids, polysaccharides, and alkaloids. It exhibits pharmacological effects such as antioxidant, anti-inflammatory, anti-tumor, and anti-thrombosis properties, making it a valuable resource in the medical field. Furthermore, due to its antioxidant and anti-inflammatory effects, safflower is increasingly being utilized in the cosmetics industry.

Methods: In this study, yeast was employed to ferment safflower, and the optimal fermentation conditions were established through single-factor experiments and response surface methodology. Subsequently, the antioxidant and anti-inflammatory efficacy of the safflower fermentation solution was assessed using both cellular and zebrafish models. Finally, the safety of the safflower fermentation solution was evaluated through a cosmetic eye irritation test.

Results: From a total of 20 yeast strains, YF-5 was identified as the dominant strain for safflower fermentation. By optimizing the fermentation conditions, it was established that the optimal parameters for YF-5 fermentation of safflower are as follows: a fermentation temperature of 36.55°C, a material-to-liquid ratio of 1:20.46, a fructose concentration of 6.20%, a fermentation duration of 72 h, and an inoculum volume of 4%. The biological activities of safflower, including its antioxidant and anti-inflammatory properties, were enhanced through yeast fermentation. In HaCaT cell and zebrafish oxidative damage assays, safflower fermentation solution inhibits the production of malondialdehyde (MDA) and increases superoxide dismutase (SOD) activity as well as total antioxidant capacity (T-AOC). In the RAW264.7 cell inflammatory damage assays, a 20% safflower fermentation solution was found to inhibit the release of TNF-α and NO in the inflammatory model, with inhibition rates of 30.94 and 28.86%, respectively. In the zebrafish inflammatory damage assays, the quantity of fluorescent neutral proteins in the 5% safflower fermentation solution was 0.7 times that observed in the dexamethasone (0.1 mg/mL) positive control group, indicating that its anti-inflammatory activity is comparable to that of dexamethasone (0.1 mg/mL). In the chicken embryo chorionic membrane experiment, it was observed that the safflower fermentation solution did not cause significant damage to the blood vessels of the chorionic allantoic membrane (CAM). This finding demonstrates that the safflower fermentation solution possesses a certain degree of safety.

Discussion: Safflower fermentation solution has antioxidant and anti-inflammatory bioactivities, and it has passed cosmetic safety evaluations. It can be used as a new natural cosmetic ingredient added to cosmetic products.

Keywords: anti-inflammatory; antioxidant; fermentation; safflower; yeast.

Figure 1

Screening of dominant strains. (A) The total polysaccharide content map of safflower fermented by different strains. (B) Total flavonoids content of safflower fermented by different strains. (C) Hydroxyl radical scavenging rate of each fermentation group. (Compared with the control group, ^*p < 0.05).

Figure 2

One-way experiment. (A) Types of carbon sources. (B) Fructose concentration. (C) Inoculum volume of bacterial solution. (D) Fermentation time. (E) Fermentation temperature. (F) Solid–liquid ratio. (Compared with the control group, ^*p < 0.05 and ^**p < 0.01).

Figure 3

Results of the Plackett–Burman and Box–Behnken experiment. (A) Contours and response surface plots of the effects of fructose concentration and solid–liquid ratio on the total flavonoid content in safflower fermentation broth. (B) Contours and response surface plots of the effects of fructose concentration and fermentation temperature on the total flavonoid content in safflower fermentation broth. (C) The effects of solid–liquid ratio and fermentation temperature on the content of total flavonoids in safflower fermentation broth were analyzed by contour map and response surface map.

Figure 4

Effect of different concentrations of safflower fermentation solution on HaCaT cells. (A) The effect of different concentrations of hydrogen peroxide and vitamin C (25 μg/mL) on the viability of HaCaT cells. (B) The effect of different concentrations of safflower fermentation liquid on the viability of HaCaT cells. (C) Total antioxidant effects of different concentrations of safflower fermentation liquid on oxidatively damaged HaCaT cells. (D) The effect of different concentrations of safflower fermentation liquid on the MDA content in oxidatively damaged HaCaT cells. (E) The effect of different concentrations of safflower fermentation liquid on the SOD activity in oxidatively damaged HaCaT cells. (Compared to the normal group, ^*p < 0.05 and ^**p < 0.01).

Figure 5

Effect of safflower fermentation solution on the growth of embryos of zebrafish of the AB lineage. (A) The effects of different concentrations of safflower fermentation broth on the hatching rate, mortality rate, and deformity rate of zebrafish embryos. Effect of safflower fermentation solution on total antioxidant capacity, MDA content, and SOD activity in an embryonic model of oxidative stress zebrafish. (B) The effect of different concentrations of safflower fermentation liquid on the total antioxidant capacity in zebrafish. (C) The effect of different concentrations of safflower fermentation liquid on the MDA content in zebrafish. (D) The effect of different concentrations of safflower fermentation liquid on the SOD activity in zebrafish. (Compared to the normal group, ^*p < 0.05).

Figure 6

Investigation of safe concentration of safflower fermentation solution against RAW264.7. (A) The effect of different concentrations of safflower fermentation liquid on the viability of RAW264.7 cells. (B) The effect of different concentrations of safflower fermentation liquid on the TNF-α content. (C) The effect of different concentrations of safflower fermentation liquid on the NO content. (Compared to the normal group, ^*p < 0.05). Effect of safflower fermentation solution on neutrophil migration in zebrafish in vivo. (D) The number of migrating fluorescent neutrophil proteins in zebrafish tails by different concentrations of safflower fermentation broths. (E) The effect of different concentrations of safflower fermentation broth on the migration of fluorescent neutrophils in the tail of zebrafish. (Compared to normal group, ^*p < 0.05, ^**p < 0.01, and ^***p < 0.001).

Figure 7

Safety evaluation of safflower fermentation solution. (A) Different concentrations of the reference substance control CAM diagram. (B) CAM diagram of 100% safflower fermentation broth.