RESEARCH ARTICLE Identification of potential serum biomarkers of inflammation and lipid modulation that are altered by fish oil supplementation in healthy volunteers Baukje de Roos 1 , Anouk Geelen 2, 3 , Karen Ross 1 , Garry Rucklidge 1 , Martin Reid 1 , Gary Duncan 1 , Muriel Caslake 4 , Graham Horgan 5 and Ingeborg A. Brouwer 2, 3 1 Department of Vascular Health, Rowett Research Institute, Aberdeen, UK 2 Wageningen Centre for Food Sciences, Wageningen, The Netherlands 3 Division of Human Nutrition, Wageningen University, Wageningen, The Netherlands 4 Department of Vascular Biochemistry, Glasgow Royal Infirmary, Glasgow, UK 5 Biomathematics and Statistics Scotland, Rowett Research Institute, Aberdeen, UK Long chain n-3 polyunsaturated fatty acids (n-3 LCPUFA) lower risk of coronary heart disease (CHD), but mechanisms are not well understood. We used proteomics to identify human serum proteins that are altered by n-3 LCPUFA. Such proteins could identify pathways whereby they affect CHD. Eighty-one healthy volunteers entered a double blind randomised trial to receive 3.5 g of fish oil or 3.5 g of high oleic sunflower oil daily. Serum was collected before and after 6 wk of intervention. Serum was analysed by proteomics using 2-DE. Proteins that were differ- entially regulated were identified by MS. We also analysed serum apolipoprotein A1 (apo A1), high-density lipoprotein (HDL) particle size and haptoglobin. Serum levels of apo A1, apo L1, zinc-a-2-glycoprotein, haptoglobin precursor, a-1-antitrypsin precursor, antithrombin III-like protein, serum amyloid P component and haemopexin were significantly downregulated (all p,0.05) by fish oil compared with high oleic sunflower oil supplementation. Fish oil supple- mentation caused a significant shift towards the larger, more cholesterol-rich HDL 2 particle. The alterations in serum proteins and HDL size imply that fish oil activates anti-inflammatory and lipid modulating mechanisms believed to impede the early onset of CHD. These proteins are potential diagnostic biomarkers to assess the mechanisms whereby fish oil protects against CHD in humans. Received: May 14, 2007 Revised: January 17, 2008 Accepted: January 18, 2008 Keywords: Biomarkers / Fish oil / Inflammation / Lipid metabolism / Serum proteomics Proteomics 2008, 8, 1965–1974 1965 1 Introduction The long chain n-3 polyunsaturated fatty acids (n-3 LCPUFA) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are found in relatively high concentrations in oily fish and the oils derived from them. Numerous epide- miological studies suggest that consumption of fish or n-3 LCPUFA protects against coronary heart disease (CHD) [1– 3]. In addition, some secondary intervention studies have shown significant benefits when n-3 LCPUFA were ad- ministered to patients who had already suffered from myo- Correspondence: Dr. Baukje de Roos, Rowett Research Institute, Division of Vascular Health, Greenburn Road, Bucksburn, Aber- deen AB21 9SB, UK E-mail: b.deroos@rowett.ac.uk Fax: 144-1224-716629 Abbreviations: apo A1, apolipoprotein; CHD, coronary heart dis- ease, DHA, docosahexaenoic acid; EPA, eicosapentaenoic acid; HDL, high density lipoprotein; LCPUFA, long-chain polyunsatu- rated fatty acids; NF-kB, nuclear factor-kB; SAP, serum amyloid P; TNF-Æ, tumour necrosis factor alpha; VLDL, very low density lipoprotein; ZAG, zinc-a-2-glycoprotein DOI 10.1002/pmic.200700457  2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.proteomics-journal.com