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The light-induced turnover of P700 was measured spectrophotometrically in a wide variety of algae and some photosynthetic mutants. Analysis of the postillumination recovery of P700+ revealed that the apparent first-order rate constant for reduction via the cyclic pathway was much lower that that via the noncyclic pathway. After activation of photosystems 1 and 2 the half-time for reduction of P700+ was 5-20 ms, whereas after activation of primarily photosystem 1 a longer half-time of ca. 150 ms was observed. The extent of the photooxidation of P700 was the same in both regimes of illumination. The longer half-time was also noted after inhibition of photosystem 2 by 3-(3,4-dichlorophenyl)-1,1-dimethylurea or mild heat shock and in mutant algae known to lack a functional photosystem 2. No signal was observed in mutants lacking P700 itself but those strains lacking either plastocyanin or cytochrome f were capable of a very slow turnover (reduction t 1/2 greater than 500 ms at room temperature). This very slow turnover was not affected by carbonyl cyanide m-chlorophenylhydrazone or the plastoquinone antagonist, 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone, indicating that the pathway for reduction of P700+ in these mutants is not energy linked and does not utilize the intersystem electron transport chain. The slow, 150 ms, reduction of P700+ due to cyclic flow was not observed when cells were engaged in photosynthesis at high-light intensities. The data are interpreted as evidence for the involvement of the total functional pool of P700 in both electron transport pathways, and we suggest that cyclic electron transport does not contribute to photosynthesis in oxygen-evolving autotrophs.

The kinetics of P -700 were examined spectrophotometrically during the induction of photosynthesis in algae. A pronounced oscillation was observed in the redox level of P -700 upon illumination of dark-adapted cells. The dark adaptation required approximately 1 min. The oscillation may be described as an initial rapid oxidation reaching a peak at approx. 50 ms followed by complete reduction of the pool of P -700. A subsequent slower oxidation resulted in attainment of the final state around 1 s. The main features of the oscillation were qualitatively the same in a wide variety of algae. The modulation in redox level of P -700 required high intensity activation of both photosystems and was eliminated by pre-illumination of the cells with weak short wavelength light but not by longer wavelengths absorbed primarily by Photosystem I. We propose that the P -700 modulation is directly related to the fast redox changes in Photosystem II which occur during the induction of photosynthesis. Cells incubated with methyl viologen did not show the P -700 oscillation confirming the suggestion previously advanced that exhaustion of Photosystem I acceptor and kinetic limitations in the carbon reduction cycle partially control the fast phase of photosynthetic induction.

Although nitrate was not produced in the oxidation of ammonia or hydroxylamine as catalyzed by intact cells, hydroxylamine oxidoreductase from Nitrosomonas europaea catalyzed the aerobic oxidation of hydroxylamine to nitrite and nitrate. The reaction was most rapid in the presence of added electron acceptor. Nitrate was not produced from nitrite. The precursor of nitrate was an N-oxide of oxidation state between hydroxylamine and nitrite. In the presence of diethyldithiocarbamate, hydroxylamine was oxidized to nitrite but nitrate was not produced. For each mol of nitrite produced, approximately 1 mol of diethyldithiocarbamate was oxidized to bis(diethyldithiocarbamoyl)disulfide (Disulfiram). Oxidation of diethyldithiocarbamate required the concomitant oxidation of hydroxylamine. The enzyme did not catalyze the direct oxidation of diethyldithiocarbamate utilizing oxygen, H2O2, nitrite, nitrate or PMS as electron acceptor. The effect of structural analogs of diethyldithiocarbamate suggested that diethyldithiocarbamate was effective because it bound to a metal ion and had an oxidizable SH group. Diethyldithiocarbamate may act by either (a) reducing a reactive form of oxygen generated during the oxidation of hydroxylamine thus preventing the oxidation of an N-compound to nitrate, or (b) reducing an N-containing precursor of nitrate to an N-compound which was subsequently oxidized to nitrite. Methods for the assay of diethyldithiocarbamate are described.