AIMS Vitamin D and polyunsaturated fatty acids (PUFAs) are derived from partially overlapping sources. Vitamin D is produced in the skin after sun exposure, but is also derived from fatty fish and fish oils. Dietary PUFAs are mainly derived from plant oils that are rich in n-6 PUFAs, but fatty fish provides high amounts of the marine n-3 PUFAs. The Western diet provides an excess of n-6 PUFAs compared to n-3 PUFAs, and the ratios of these may influence human health. Here, we investigated the potential associations of plasma concentrations of vitamin D, marine PUFAs and PUFA ratios. METHODS Plasma concentrations of vitamin D (25(OH)D), marine PUFAs, and PUFA ratios were measured in 372 women from the Norwegian Women and Cancer (NOWAC) Post-Genome Cohort. Covariability was examined in 310 non-users of cod liver oil, using Spearman's rank correlation and linear regression. RESULTS In non-users of cod liver oil, the average concentration of vitamin D was 40.3 nmol/L, and marine PUFA concentration was 0.2 mg/g. PUFA ratios were dominated by the n-6 fatty acids. Vitamin D levels were significantly associated with marine fatty acids and weakly associated with PUFA ratios. CONCLUSIONS Concentrations of vitamin D and marine PUFAs were below recommended levels. The correlation analyses indicated that health-related effects of vitamin D and marine PUFAs respectively may be hard to separate in epidemiological studies. However, measured health effects of PUFA ratios and vitamin D are likely to derive from the influence of the two factors separately. The presented results are the first to show these associations in a nationally representative cohort.