Quantitative Analysis of Bixin

Analytical grade cis-bixin (9-cis-6,6′-diapo-ψ,ψ-carotenedioic acid, 6-methyl ester) was purchased from Spectrum (CAS number: 6983-79-5). LC/MS confirmation of purity (>98% by weight) was performed using electrospray mass spectrometry of bixin [dissolved in tetrahydrofuran and diluted 10-fold in acetonitrile/NH₄OH (0.1 N); ESI-MS (negative ion mode) m/z 393.21 (M - 1)^-] employing a Bruker Apex FT/ICR mass spectrometer, as specified before (Tao et al., 2015 (link)). Polyethylene glycol 400 (PEG400) was from EMD Millipore. Xanthotoxin (8-methoxypsoralen, 8-MOP), and hydrogen peroxide (H₂O₂) were from Sigma. Primary antibodies against NRF2, KEAP1, TRXR1, GCLM, NQO1, HO1, OGG1, GAPDH, and actin were purchased from Santa Cruz Biotechnology. Primary antibody against p62 was from Abnova. Secondary antibodies conjugated with horseradish peroxidase (HRP) were purchased from Sigma. Antibody against 8-oxo-deoxyguanosine (8-oxo-dG) was from Trevigen.

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Rojo de la Vega M., Zhang D.D, & Wondrak G.T. (2018). Topical Bixin Confers NRF2-Dependent Protection Against Photodamage and Hair Graying in Mouse Skin. Frontiers in Pharmacology, 9, 287.

Publication 2018

Apex FT/ICR mass spectrometer Polyethylene glycol 400 (PEG400) hydrogen peroxide (H2O2) Primary antibodies against NRF2 KEAP1 TRXR1 GAPDH actin Secondary antibodies conjugated with horseradish peroxidase (HRP)

8 methoxypsoralen 8 mop 8 oxo dg Acetonitrile Acid Actin Antibody Bixin Ester Gapdh Gclm H2o2 Horseradish peroxidase Keap1 Mass spectrometry Nqo1 Nrf2 Ogg1 Polyethylene glycol 400 Tetrahydrofuran

Corresponding Organization : University of Arizona

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Variable analysis

independent variables

Cis-bixin (9-cis-6,6′-diapo-ψ,ψ-carotenedioic acid, 6-methyl ester)
Polyethylene glycol 400 (PEG400)
Xanthotoxin (8-methoxypsoralen, 8-MOP)
Hydrogen peroxide (H2O2)

dependent variables

Purity of cis-bixin (>98% by weight) as confirmed by LC/MS using electrospray mass spectrometry

control variables

Tetrahydrofuran (used to dissolve bixin)
Acetonitrile/NH4OH (0.1 N) (used to dilute bixin)
Bruker Apex FT/ICR mass spectrometer (used for mass spectrometry analysis)

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