Betaine

Four isonitrogenous (41% crude protein) experimental diets with two levels of lipid (11% and 17% crude lipid) were formulated with the diets containing the higher lipid level supplemented with two levels of betaine (Sinopharm Chemical Reagent Co., Ltd., Shanghai, China). The diets were termed as (1) Control, medium-fat diet (11%), (2) HFD, high-fat diet (17%), (3) HFD+B1, HFD with betaine supplement (10 g/kg dry diet), (4) HFD+B2, HFD with betaine supplement (20 g/kg dry diet) (Table 1). Fishmeal, soybean protein concentrate and soybean meal were used as protein sources, with fish oil, palmitic acid and soybean lecithin used as the main lipid sources. All ingredients were purchased from Ningbo Tech-Bank Feed Co. Ltd., Ningbo, China. The experimental diets were produced according to the methods described in detail previously (5). Briefly, the ground ingredients were mixed in a Hobart type mixer and cold-extruded pellets produced (F-26, Machine factory of South China University of Technology) with pellet strands cut into uniform sizes (2 mm and 4 mm diameter pellets) (G-250, Machine factory of South China University of Technology). Pellets were heated for 30 min at 90°C, and then air-dried to approximately 10% moisture, sealed in vacuum-packed bags and stored at −20°C until used in the feeding trial.

Ginsenoside Ro (98%), oleanolic acid (99%), Stigmasterol (95%), palmatine hydrochloride (98%), betherine (98%), epiberberine (98%), coptisine (98%), astragalin (98%), isoquercitrin (98%), baicalin (98%), wogonin (98%), chrysophanol (98%), physcion (98%) and Geniposide (98%) were sourced from the Chengdu Must Bio-technology Co., Ltd (Chengdu, Sichuan Province, China). Betaine (98%), nonanedioic acid (99.5%), succinic acid (90%), allantoin (98.5%), rutin (95%), β-sitosterol (95%) and 5-hydroxymethyl furaldehyde (99%) were sourced from Sinopharm Chemical Reagent Co., Ltd (Shanghai, China). β-ecdysterone was sourced from J&K Scientific Ltd (Beijing, China).

Citrus peel from orange (Citrus sinensis) and Longissimus thoracis (LT) muscle from pig (Duroc × Landrace × Yorkshire, DLY) were purchased from a supermarket in Changsha City. Toronto Research Chemicals Inc. provided a total of 17 HA and 2 AGE standards. Waters (Milford, MA, USA) was the source of Oasis MCX SPE column. Choline chloride, L-carnitine, betaine, acetamide, ethylene glycol, carbamide, and additional analytical grade chemicals were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China).