SummaryA range of aqueous and non‐aqueous methods for chloroplast preparation were tested to establish suitable methodology for routine analysis of plastid anion content using high performance ion chromatography (HPIC). Aqueous methods of plastid isolation failed to provide a satisfactory procedure for analysis of anions (other than chloride) but three different non‐aqueous methods all gave satisfactory and effective procedures.The simplest non‐aqueous method was used to analyse the anions in chloroplasts from 9 d old barley seedlings which had been grown in clean air or fumigated for the last 1 to 3 d of growth with high, but not unrealistic, concentrations (0.28 ppm) of either SO2 or NO2. Amounts of chloride, phosphate, sulphate, sulphite, nitrate and nitrite were detected in plastid preparation from both clean air and polluted seedlings. NO2 fumigation had no effect on chloride, phosphate, sulphate or sulphite content but increased the levels of nitrite and decreased those of nitrate especially on the second day of treatment. Similar atmospheric concentrations of SO2 had no effect on chloride content but increased the levels of phosphate, sulphate and sulphite as well as having a similar influence as NO2 upon nitrate and nitrite content. Again all these effects of SO2 were greatest on the second day of fumigation.