The influence of ascorbic acid (AA) intake on plasma cortisol (PC) and tissue AA levels after ACTH treatment was examined using guinea pigs. ACTH produced an 8- to 10-fold rise in PC levels (p < 0.0001) over the 4-hour experimental period compared to gel-injected control animals. The magnitude of rise in PC was similar at both normal (0.50 g/kg diet) and high (10 g/kg diet) AA intakes. A 30–100% higher (p < 0.0001) level of AA in tissues (adrenals, liver, and kidneys) and plasma was observed with the high AA diet. ACTH resulted in a 26–30% lower level of AA in the adrenals (p < 0.0001), but not in other tissues or plasma. PC and adrenal AA responses were unrelated (r = -0.326). These results suggest that ACTH alters AA only in the adrenals, the PC response to and percent decrease in adrenal AA levels with ACTH are not related to AA status, and the absolute level of AA in the adrenals is not critical for steroidogenesis.