Elix type 2 pure water system

Commercial cocoa powder produced from Forastero cocoa nibs with 10-12% (w/w) fat was supplied by Cargill (Schiphol, Netherlands). Analytical grade NaOH (98%) for alkalization was purchased from Chem-Lab NV (Zedelgem, Belgium) and demineralized water was purified using an Elix type 2 pure water system 20 from Merck Millipore (Overijse, Belgium). UPLC-grade acetonitrile, methanol, formic acid and ammonium formate were purchased from Biosolve (Valkenswaard, The Netherlands), while n-hexane (≥ 97%), acetone (99.8%) and acetic acid (99.8%) were purchased from Acros Organics (Geel, Belgium). Analytical standards were purchased from Extrasynthese (Lyon, France), Sigma-Aldrich (Bornem, Belgium), Santa Cruz Biotechnology (Heidelberg, Germany), or Carl Roth (Karlsruhe, Germany). All other chemicals were of the highest grade commercially available.

Commercial fat-reduced cocoa powder (10-12% w/w) was supplied by Cargill (Schiphol, Netherlands). Analytical grade NaOH (98%) for alkalization was purchased from Chem-Lab NV (Zedelgem, Belgium) and demineralized water was purified using Elix type 2 pure water system 20 from Merck Millipore (Overijse, Belgium). Methanol (≥99.8%) and formic acid (98%) were obtained from Fischer Scientific (Loughborough, UK), while n-hexane (≥97%), acetone (99.8%) and acetic acid (99.8%) were purchased from Acros Organics (Geel, Belgium) . Reference compounds of theobromine, caffeine, epicatechin and catechin (>98% purity) were provided by Sigma Aldrich (Overijse, Belgium). All the other chemicals were of highest grade commercially available.