D 10 camphorsulfonic acid

Sample preparation and measurement were performed as previously described [4 (link), 27 (link)]. Plasma (40 μL) was extracted with the addition of 400 μL ice-cold methanol containing internal standards (10 mM L-methionine sulfone [Wako, Tokyo, Japan], 100 mM 2-morpholinoethanesulfonic acid [Dojindo Molecular Technologies, Rockville, MD, USA], 100 mM D-10-camphorsulfonic acid [Wako]), 400 μL chloroform, and 120 μL water. After centrifugation at 10,000 × g for 3 min at 4°C, the separated aqueous layer was filtered through a 5 kDa cutoff filter (Millipore, Burlington, MA, USA) to remove protein contamination. The filtrate (300 μL) was lyophilized and dissolved in 20 μL water containing two reference compounds (200 μM each of trimesate [Wako] and 3-aminopyrrolidine [Sigma-Aldrich, St. Louis, MO, USA]) for migration time and then injected into a capillary electrophoresis time-of-flight mass spectrometry system (Agilent Technologies, Santa Clara, CA, USA) [29 (link)–31 (link)]. Among the measured molecules, gastric inhibitory polypeptide (GIP) (active) was measured using an enzyme-linked immunosorbent assay kit. Blood hormones and some metabolites were measured according to methods developed by LSI Medience Co., Ltd. (Tokyo, Japan).

Plasma (40 microliters [μL]) was extracted with the addition of 400 μL of ice-cold methanol containing the internal standards (10 millimolars [mM] l-methionine sulfone [Wako], 100 mM 2-morpholinoethanesulfonic acid [Dojindo], 100 mM D-10-camphorsulfonic acid [Wako]), 400 μL of chloroform, and 120 μL of water. After centrifugation at 10,000 × g for 3 min at 4 °C, the separated aqueous layer was filtered through a 5 kilodalton (kDa) cutoff filter (Millipore) to remove protein contamination. The filtrate (300 μL) was lyophilized and dissolved in 20 μL water containing the 2 types of reference compounds (200 μM each of trimesate [Wako] and 3-aminopyrrolidine [Sigma-Aldrich]) for migration time and then injected into the capillary electrophoresis time-of-flight mass spectrometry (CE-TOFMS) system (Agilent Technologies)^{36 (link)–38 (link)}. Among the measured molecules, GIP (active) was measured using an ELISA kit. Blood hormones and some metabolites were measured according to methods developed by LSI Medience Co., Ltd. The methods used to measure each of these molecules are listed in Supplementary Data 5; among these, the amino acid fractions measured by liquid chromatography–mass spectrometry (LC–MS) are listed in Supplementary Data 6, and the metabolites measured by CE-TOFMS are listed in Supplementary Data 7.