Spinach is an important dietary source of lutein and β-carotene. Their synthesis is closely linked to chlorophyll synthesis and dependent upon an adequate supply of sulfur and nitrogen. Soils may become sulfur-deficient during winter because microorganisms convert atmospheric SO2 less efficiently to sulfate. The influence of sulfur and nitrogen fertilization on the carotenoid and chlorophyll contents of spinach grown in summer or winter was investigated. Carotenoid and chlorophyll levels were positively correlated. Lutein and β-carotene were 25% higher in summer than in winter. Winter levels were increased by 35-40% by sulfur fertilization in one location but not in the other, with the impact depending upon soil type, growing location, and atmospheric conditions. Carotenoids were little or not affected by nitrogen addition in winter or sulfur addition in summer. It is concluded that sulfur fertilization of spinach in winter may modestly increase carotenoids but high carotenoid levels are best assured with carotenoid-rich cultivars grown in summer.