Ultrasound-assisted extraction of polysaccharides from Ginkgo biloba: Process optimization, composition and anti-inflammatory activity

Abstract

Plant derived polysaccharides can enhance immune function in the human body, effectively prevent diseases, and reduce the probability of bacterial infections. Ginkgo crude polysaccharide (GCP) was obtained from Ginkgo biloba by ultrasonic-assisted hot water extraction. Our data showed that the best extraction conditions of GCP were as follows: extraction temperature 80 °C, ultrasonic time 35 min, extraction time 3 h, and solid‒liquid ratio 1:30. Fourier transform infrared spectrometer (FT-IR) data showed that this polysaccharide might be an acidic polysaccharide with a carboxylic acid ring structure. Further studies implied that GCP was mainly composed of glucose, galacturonic acid, rhamnose, galactose and arabinose, accounting for 39.45 %, 25.01 %, 15.40 %, 11.94 % and 4.25 %, respectively. 0.1, 1 and 10 mg/mL GCP reduced the release of inflammatory factors in RAW264.7 cells via inhibition of the nuclear factor kappa-light-chain-enhancer of activated B (NF-κB) signalling pathway. GCP was separated into five components with different molecular weights by an ultrafiltration membrane. Our data showed that GPa with a molecular weight ≥100 kDa was the main component of GCP. 1 mg/mL GPa, GPb, GPc and GPd had anti-inflammatory activities, and 1 mg/mL GPa had the best anti-inflammatory activities. Our results preliminarily reveal the elements and biological activity of GCP, which will provide a reference for the development of Ginkgo biloba.

Keywords: Ginkgo biloba; Inflammatory factors; Ultrafiltration membrane.

Graphical abstract

Fig. 1

Optimization of extraction processes of polysaccharides from Ginkgo biloba (A) The impact of extraction temperature on the extraction rate of polysaccharides from Ginkgo biloba. (B) The effect of ultrasonic waves on the extraction rate of polysaccharides from Ginkgo biloba. (C) The impact of extraction time on the extraction rate of polysaccharides from Ginkgo biloba. (D) The effect of solid‒liquid ratio on the extraction rate of polysaccharides. Different letters in a curve represent significant differences (P < 0.05). Each experimental group was tested at least 3 times.

Fig. 2

Composition and group analysis of polysaccharides. (A) Fourier transform infrared spectrum of polysaccharides from Ginkgo biloba. (B) Analysis of the monosaccharide composition of GCP by ion chromatography.

Fig. 3

GCP inhibited the release of inflammatory factors in RAW264 7 cells. (A) The RAW264.7 cells were treated with 0.1, 1 or 10 μg/mL LPS for 12 h, and the cell viability rate was analyzed by the method of CCK-8. (B) The RAW264.7 cells were treated with 0.1, 1 and 10 mg/mL with GCP for 12 h, and the cell viability was analyzed by the CCK-8. (C)–(E) The RAW264.7 cells were treated with or without 10 μg/mL LPS, 10 μg/mL LPS plus 0.1 mg/mL, 1 mg/mL or 10 mg/mL GCP for 12 h and the concentration of TNF-α (C), IL -6 (D) or IL -1β (E) was determined by the method ELISA. Different letters mean differences are distinctive (p < 0.05). Each experimental group was done at least 3 times.

Fig. 4

GCP inhibited the release of inflammatory factors in RAW264.7 cells via the NF-κB signalling pathway. (A) RAW264.7 cells were treated with or without 10 μg/mL LPS or 10 μg/mL LPS plus 10 mg/mL GCP for 12 h, and the expression of p65 or phospho-p65 was immunoblotted with antibodies against phospho-p65 and p65. (B) RAW264.7 cells were treated with or without 10 μg/mL LPS or 10 μg/mL LPS plus 10 mg/mL GCP for 12 h, and the expression of IκBα or phospho-IκBα was immunoblotted with antibodies against phospho-IκBα and IκBα. GAPDH was used as negative control.

Fig. 5

The effect of GPs on the LPS-induced release of inflammatory factors. (A) RAW264.7 cells were treated with or without 1 mg/mL GPs, and the rate of cell viability was tested by CCK-8. RAW264.7 cells were treated with or without 1 mg/mL GPs plus 10 μg/mL LPS for 12 h, and the release of inflammatory factors, including TNF-α (B), IL-6 (C) and IL-1β (D), was determined by ELISA. Different letters indicate significant differences (P < 0.05). Each experimental group was tested at least 3 times.

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