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SUMMARY: Several substances have been tested for inhibitory effects on carotenoid biosynthesis in Flavobacterium R1519, an organism which normally produces zeaxanthin (β,β-carotene-3,3ʹ-diol) as essentially its only carotenoid. Diphenylamine, several mixed-function oxidase inhibitors (SKF 525A, SKF 3301A, SKF 7997A3, metopirone base, diphenylpropylacetic acid, piperonyl butoxide), several alkaloids (atropine, brucine, hyoscyamine, pilocarpine, quinidine, quinine, scopolamine, strychnine), some substitued pyridines (pyridine, 3-picoline, 3-ethylpyridine, 5-ethyl-2-methylpyridine) and morpholine showed no specific inhibitory effects. In cultures grown in the presence of CaCl2, β-carotene (10%) and β-cryptoxanthin (10%) partly replaced zeaxanthin. Nicotine specifically inhibited zeaxanthin synthesis. Lycopene (in 5 mM-nicotine) and rubixanthin (in 1 mM-nicotine) accumulated as the new main pigments; [2-14C]mevalonic acid was efficiently incorporated into both. Nicotine thus inhibited the cyclization reaction in carotenoid biosynthesis in Flavobacterium R1519 as in other organisms; in zeaxanthin biosynthesis cyclization appeared to precede hydroxylation. Similar inhibitory effects were produced by (+) and (-) nicotine and by nor-nicotine, but not by the closely related anabasine, myosmine, β-nicotyrine and nor-nicotyrine.