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Cut leaves ofAmaranthus hypochondriacuswere placed in one of three solutions of [NaH¹³CO₃] dissolved in KCl water to measure the efflux of xylem‐transported CO₂exiting the leaf in the dark or rates of assimilation of xylem‐transported CO₂* in the light, in real‐time, using a tunable diode laser absorption spectroscope.

In the dark, the efflux of xylem‐transported CO₂increased with increasing rates of transpiration and [¹³CO₂*]; however, rates of¹³C_effluxinA. hypochondriacuswere lower compared to C₃species. In the light,A. hypochondriacusfixed nearly 75% of the xylem‐transported CO₂supplied to the leaf.

Kranz anatomy and biochemistry likely influence the efflux of xylem‐transported CO₂out of cut leaves ofA. hypochondriacusin the dark, as well as the use of xylem‐transported CO₂* for photosynthesis in the light. Thus increasing the carbon use efficiency of Kranz‐type C₄species over C₃species.

Cut leaves ofPopulus deltoidesandBrassica napuswere placed in eitherKCl or one of three [NaH¹³CO₃] solutions dissolved in water to simultaneously measure the assimilation and the efflux of xylem‐transportedCO₂exiting the leaf across light andCO₂response curves in real‐time using a tunable diode laser absorption spectroscope.

The rates of assimilation and efflux of xylem‐transportedCO₂increased with increasing xylem [¹³CO₂*] and transpiration. Under saturating irradiance, rates of assimilation using xylem‐transportedCO₂accounted forc.2.5% of the total assimilation in both species in the highest [¹³CO₂*].

The majority of xylem‐transportedCO₂is assimilated, and efflux is small compared to respiration. Assimilation of xylem‐transportedCO₂comprises a small portion of total photosynthesis, but may be more important whenCO₂is limiting.

Gas exchange necessarily incorporates photosynthesis and (photo)respiration. Each process was expected to respond on different timescales due to differences in metabolite compartmentation, biochemistry and diffusive pathways. We hypothesized that metabolic lags in photorespiration relative to photosynthesis/respiration andCO₂diffusional limitations can be detected by varying the rate of change in [CO₂] duringRACiR assays. We tested these hypotheses through modeling and experiments at ambient and 2% oxygen.

Our data show that photorespiratory delays cause offsets in predictedCO₂compensation points that are dependent on the rate of change in [CO₂]. Diffusional limitations may reduce the rate of change in chloroplastic [CO₂], causing a reduction in apparentRACiR slopes under highCO₂ramp rates.

MultirateRACiRs may prove useful in assessing diffusional limitations to gas exchange and photorespiratory rates.