Quantitation of photosystem II (PSII) activity in spinach chloroplasts is presented. Rates of PSII electron‐transport were estimated from the concentration of PSII reaction‐centers (Chl/PSII = 380:1 when measured spectrophotometrically in the ultraviolet [ΔA 320 ] and green [ΔA 540–550 ] regions of the spectrum) and from the rate of light utilization by PSII under limiting excitation conditions. Rates of PSII electron‐transport were measured under the same light‐limiting conditions using 2,5‐dimethylbenzoquinone or 2,5‐dichlorobenzoquinone as the PSII artificial electron acceptors. Evaluation is presented on the limitations imposed in the measurement of PSII electron flow to artificial quinones in chloroplasts. Limitations include the static quenching of excitation energy in the pigment bed by added quinones, the fraction of PSII centers (PSII β ) with low affinity to native and added quinones, and the loss of reducing equivalents to molecular oxygen. Such artifacts lowered the yield of steady‐state electron transport in isolated chloroplasts and caused underestimation of PSII electron‐transport capacity. The limitations described could explain the low PSII concentration estimates in higher plant chloroplasts (Chl/PSII = 600 ± 50) resulting from proton flash yield and/or oxygen flash‐yield measurements. It is implied that quantitation of PSII by repetitive flash‐yield methods requires assessment of the slow turnover of electrons by PSII β and, in the presence of added quinones, assessment of the PSII quantum yield.
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