Optimization of Geranium wilfordii Maxim. polysaccharide extraction process and its alleviation of chronic aflatoxin b1 toxicity in duckling

The present study was aimed at optimizing the extraction process of Geranium wilfordii Maxim. polysaccharide (GWP) and investigating its protective effects against chronic aflatoxin B 1 (AFB 1 ) poisoning in ducklings. The extraction parameters for GWP were optimized via response surface methodology (RSM), followed by characterization of its monosaccharide composition using FTIR and PMP-HPLC methods. A chronic AFB 1 poisoning model was established in ducklings, and the animals were assigned to four groups: blank control (BC), AFB 1 , high-dose GWP (GWP-H), and low-dose GWP (GWP-L), for a 21-day experimental period. Growth performance, serum biochemical indices (AST, ALT, and TP), gut microbiota (16S rRNA sequencing), tight junction protein expression, hepatic antioxidant status, histopathological changes in the liver and jejunum, and the regulation of the TLR4/NF-κB/NLRP3 signaling pathway were comprehensively evaluated. In addition, serum pharmacology analysis was performed in primary duck hepatocytes (PDHs) in the presence of the TLR4 agonist RS09 TFA to further validate the underlying mechanisms. The optimal extraction conditions for GWP were determined to be an extraction temperature of 100 °C, a solid-to-liquid ratio of 1:18 (g/mL), and three extraction cycles. PMP-HPLC analysis showed that GWP was a heteropolysaccharide composed of mannose, ribose, rhamnose, glucuronic acid, galacturonic acid, glucose, xylose, and arabinose, with molar ratios of 7.17:1:5.14:2.15:11.69:40.29:20.09:11.82, respectively. In the animal experiment, AFB 1 -induced chronic toxicity in ducklings was significantly alleviated by GWP treatment, as evidenced by the restoration of gut microbiota homeostasis, mitigation of intestinal barrier injury, attenuation of hepatic oxidative stress, and suppression of the TLR4/NF-κB/NLRP3 signaling pathway in both the jejunum and liver. In vitro serum pharmacology experiments found that, AFB 1 group serum significantly decreased PDHs viability and markedly increased TLR4 mRNA expression; GWP-H goup and GWP-L goup serum reversed these trends, whereas co-treatment with RS09 TFA and GWP group serum attenuated their ameliorative effects on cell viability and the regulation effect on TLR4 expression. The extraction method for GWP was optimized in this study, and its potential to alleviate AFB 1 -induced liver and jejunal injury in ducklings was confirmed. These protective effects may involve the restoration of intestinal and hepatic homeostasis and the inhibition of abnormal TLR4 activation, thereby providing a theoretical basis for its further clinical application. • The extraction method for GWP was optimized using response surface methodology. • Growth retardation along with intestinal and hepatic injuries induced by chronic AFB 1 exposure in ducklings were alleviated following GWP administration. • Damage to the intestinal barrier and hepatic oxidative stress caused by chronic AFB 1 exposure were ameliorated after GWP intervention. • Both jejunal and hepatic injuries resulting from chronic AFB 1 exposure were mitigated through suppression of the TLR4/NF-κB/NLRP3 signaling pathway hyperactivation by GWP.