SUPPLIMENTARY MATERIAL
Analysis of Oxysterols by Electrospray Tandem Mass Spectrometry*
William J. Griffiths1, Yuqin Wang1, Gunvor Alvelius2, Suya Liu2, Karl Bodin2 and Jan Sjövall2
1Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29-39 Brunswick Square, London WC1N 1AX, UK.
2Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-17177 Stockholm, Sweden.
Figure Captions
Figure S-1. MS/MS spectra of the [M]+ ion of GP hydrazones (a) C4-7a,27-diol-3-one (XV, m/z 550.40); (b) [16,16,17(or 20),22,22,23,23-2H7]C4-7a,27-diol-3-one (XVI, m/z 557.44); (c) C4-7a,25-diol-3-one (XIII, m/z 550.40); (d) C4-7b,25-diol-3-one (XIV, m/z 550.40); (e) C4-20R,22R-diol-3-one (XII, m/z 550.40); (f) C4-diol-3-one from brain (m/z 550.40). In the MS/MS spectra the ratio of fragment ions 123.09:135.09:149.11 and 151.09:163.09:177.10 are 10:1:1 and 10:1:1 (a); 10:2:1 and 10:1:1 (b); 10:1:1 and 10:1:1 (c); 10:1:1 and 10:1:1 (d); 7:10:1 and 8:10:1 (e); 4:10:4 and 8:10:4 (f). In (a) - (d) the m/z regions 50 – 121 and 122 – 390 have been magnified by a factor of 280 and 10, respectively; while in (e) and (f) by a factor of 54 and 10 respectively. The y-axis represents relative abundance. All spectra recorded on the Q-TOF Global instrument in “W-mode” at a collision energy 35 eV. 1 - 5 mL of sample (1 ng/mL for steroid standards and 10 mg brain/mL in (f)) was loaded into the nano-ES capillary, and about 1 mL consumed for the acquisition.
Figure S-2. MS/MS spectra of the [M]+ ion of GP hydrazones (a) [16,16,17(or 20),22,22,23,23-2H7]CA4-3-one (XVII, m/z 555.43); (b) [16,16,17(or 20),22,22,23,23-2H7]CA4-7a-ol-3-one (XVIII, m/z 571.42); (c) [16,16,17(or 20),22,22,23,23-2H7]CA4-7b-ol-3-one (XIX, m/z 571.42); (d) CA4-7b-ol-3-one-27-methyl ester (XXI, m/z 585.44). In the MS/MS spectra the ratio of fragment ions 123.09:135.09:149.11 and 151.09:163.09:177.10 are 7:10:2 and 7:10:2 (a); 10:1:1 and 10:1:1 (b); 10:5:3 and 10:1:2 (c); 10:1:1 and 10:1:1 (d). The m/z regions 50 – 121 and 122 – 390 have been magnified by a factor of 280 and 10. The y-axis represents relative abundance. All spectra recorded on the Q-TOF Global instrument in “W-mode” at a collision energy 35 eV. 1 - 5 mL of sample (1 ng/mL) was loaded into the nano-ES capillary, and about 1 mL consumed for the acquisition.
Figure S-3. MS/MS spectra of the [M]+ ion of GP hydrazones (a) BA4-3-one (XXII, m/z 506.34); (b) [2,2,4,23-2H4]BA4-3-one (XXIII, m/z 510.36); (c) [2,2,4,23-2H4]BA4-3-one-24-oylglycine (XXIV, m/z 567.38); (d) BA4-7a-ol-3-one (XXV, m/z 522.33). In (a & d) the ratio of fragment ions 123.09:135.09:149.11 and 151.09:163.09:177.10 are 6:10:2 and 7:10:1 (a); 10:1:1 and 10:1:1 (d); while in (b & c) the ratio of fragment ions 126.11:138.11:152.13 and 154.11:166.11:180.12 are 6:10:2 and 6:10:2 (b); 8:10:1 and 8:10:2 (c). The m/z regions 50 – 121 and 122 – 390 have been magnified by a factor of 280 and 10. The y-axis represents relative abundance. All spectra recorded on the Q-TOF Global instrument in “W-mode” at a collision energy 35 eV. 1 - 5 mL of sample (1 ng/mL) was loaded into the nano-ES capillary, and about 1 mL consumed for the acquisition.
Figure S-4. MS/MS spectra of the [M]+ ion of GP hydrazones (a) C4-3-one (I, m/z 518.41); (b) C4,24-3-one (II, m/z 516.40); (c) C4-24b-ethyl-3-one (III, m/z 546.44); (d) C4,22-24b-ethyl-3-one (IV, m/z 544.43). In the MS/MS spectra the ratio of fragment ions 123.09:135.09:149.11 and 151.09: 163.09: 177.10 are 6:10:2 and 6:10:2 (a); 7:10:2 and 7:10:2 (b); 7:10:2 and 7:10:2 (c); 6:10:1 and 8:10:1 (d). The m/z regions 50 – 121 and 122 – 390 have been magnified by a factor of 280 and 10. The y-axis represents relative abundance. All spectra recorded on the Q-TOF Global instrument in “W-mode” at a collision energy 35 eV. 1 - 5 mL of sample (1 ng/mL) was loaded into the nano-ES capillary, and about 1 mL consumed for the acquisition.