Vincamine

BK powder (100 mg) was suspended in 4 mL of methanol/purified water (75 : 25, v/v). The suspension was shaken and sonicated for 5 min each. After centrifugation at 1700 g for 5 min at room temperature, the supernatant was collected as the first extract solution. Then, 4 mL of methanol/purified water (50 : 50, v/v) was added to the residue, and the second extraction solution obtained by the procedure described above was mixed with the first extract solution. The mixed extract solution undiluted or diluted 10- or 100-fold with methanol was injected into the LC-MS/MS system after pooling with a respective internal standard solution, niflumic acid (Sigma-Aldrich), or vincamine (Tokyo Chemical Industry Co.). Two systems were used for analysis of the constituents: system 1, an API4000 triple quadrupole mass spectrometer (AB SCIEX, Tokyo, Japan) equipped with an Agilent 1100 system (Agilent Technologies, Inc., Tokyo, Japan); system 2, a TripleQuad6500 (AB SCIEX) equipped with an Agilent 1290 system (Agilent Technologies, Inc.). Analytical conditions are summarized in the Supplementary Tables S1 and S2 (available online at http://dx.doi.org/10.1155/2015/853846).

GM (Lot No. GME001) was supplied by the Botanical Raw Materials Research Department of Tsumura & Co (Ibaraki, Japan). DMSO (a solvent for the test substance) was purchased from Wako Pure Chemical Industries, Ltd. (Osaka, Japan), vincamine (IS for measuring GM and HM-1/2) from Tokyo Chemical Industry Co., Ltd. (Tokyo, Japan), and 2,5-dihydroxybenzoic acid (a matrix for MALDI-MSI analysis) from Bruker Daltonics (Bellerica, MA, USA). Other chemicals were purchased from commercial sources.