Choline bitartrate

Betaine, choline bitartrate, and GAA were purchased from Sigma-Aldrich (St. Louis, MO). Spinach powder was kindly supplied by KAGOME Co. Ltd. (Tochigi, Japan). All the other chemicals were purchased from Wako Pure Chemical Industries (Osaka, Japan) or Sigma-Aldrich and were of analytical grade. Milk casein was purchased from Nacalai Tesque (Kyoto, Japan) and soybean protein isolate (SPI) was obtained from Fuji Oil (Izumisano, Japan). Mineral mixture (AIN-93G), vitamin mixture (AIN-93), and cellulose powder were purchased from Oriental Yeast (Tokyo, Japan), and the other ingredients of the diet were purchased from Wako or Sigma-Aldrich.

The hypercaloric feed was formulated based on the AIN-93G diet and according to Cheng et al. [45 (link)]
The ingredients: cellulose, carboxymethyl cellulose, L-cystine, choline bitartrate, and butylhydroquinone were purchased from Sigma Aldrich (Burlington, MA, USA). Starch, clarified anhydrous butter, maltodextrin, sucrose, and soybean oil were purchased from a local feed formulating ingredient distributor. Vitamin and mineral premix were purchased from Dyets Inc. (Bethlehem, PA, USA) and casein was purchased from Hilmar Ingredients (Hilmar, CA, USA). The distribution in percentage is shown in Table 3, and macronutrient distribution compared to the standard diet is presented in Table 4. The hypercaloric diet was prepared by mixing all ingredients and baking at 100 °C for 1 h.

WD (45% fat and 16% sucrose by kCal, based on Research Diets D12451) was prepared in-house as described in Healy et al.^{48 (link)}. Ingredients were sourced from local suppliers including sucrose (JL Stewart, GRAD25B, Australia), corn starch (JL Stewart, CFLR25W, Australia), wheat bran (JL Stewart, BRAN10UF, Australia), casein (Ross Cootee, Australia), methionine (Sigma, M9500, Australia), gelatine (JL Stewart, GEL2, Australia), choline bitartrate (Sigma, C1629, Australia), lard (JL Stewart, LARD15, Australia), safflower oil (Protec, Australia), trace minerals (MP Biomedicals, 0296026401, Australia), AIN-93M mineral mix (MP Biomedicals, 0296040102, Australia), and AIN-93-VX vitamin mix (MP Biomedicals, 0296040201, Australia).