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Month: February 2011

Measuring Photorespiration with the LI-6400/XT System

Measuring Photorespiration with the LI-6400/XT System


Oxygen is a competitive substrate for ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco), resulting in an inhibitory effect on net photosynthesis.  Higher CO2 to O2 ratios inside the leaf decrease the Rubisco-catalyzed reactions with O2, resulting in less carbon-wasteful reactions in the light (photorespiration). Photorespiration increases on hot, dry days when evaporative stress can cause stomata to close, lowering the intercellular CO2:O2 as CO2 is assimilated. When Rubisco was evolved by plants, earth’s atmosphere had much higher concentrations of CO2 and relatively low O2.  As plants evolved and filled different environmental niches, O2 production increased and atmospheric O2 concentration rose, thereby increasing photorespiration.  C4-type plants such as maize and sugar cane evolved an active CO2-concentrating mechanism that increases the CO2 concentration near the site of the photosynthetic reactions, which suppresses photorespiration.  Scientists and plant breeders are working to transfer this ability of C4-type plants into important C3-type plants such as wheat, rice and soybeans.  One approach to studying and quantifying photorespiration is to suppress it by supplying the leaf with only 2% oxygen (instead of the normal 21% oxygen) and measure the increased rates of photosynthesis.

Experimental Setup

The diagram below shows a simple setup in which a tank of air with only 2% oxygen supplies the incoming air to the LI-6400/LI-6400XT system. To prevent the LI-6400/XT internal pump from fighting against the external tank pressure, the tank supply is fed into the LI-6400/XT air inlet port via a T-fitting. One end of the fitting is vented to the atmosphere (through the flow meter in the diagram below) while the other flows into the LI-6400/XT.  The tank air flow is adjusted so that it slightly exceeds the demand from the LI-6400/XT so that ambient air is not sucked in by the LI-6400/XT pump. Venting to the atmosphere also ensures that the system remains at atmospheric pressure. An inexpensive flow meter can be attached to the vented part of the T-fitting to check and make sure that the air flow is in excess of the demand from the LI-6400/XT pump.

The photograph below shows an actual setup with a tank of gas (air with 2% oxygen)  for supplying air to the LI-6400/XT via a vented T-fitting. A float-type flow meter (taped to the side of the supply tank) is being used to test for excess flow.

This setup was used to test the differences between the response of  Maize (a C4 species) and Bush Beans (a C3 species) to air with a 2% oxygen content and room air with the normal 21% oxygen content.


The figure below shows the Bush Bean leaf (a C3 species) photosynthetic rate in response to varying Ci, the intercellular CO2 concentrations.  At ambient CO2 concentration (Ci ≈250 ppm), photorespiration was reducing the net photosynthetic rate by about 40%.

The figure below shows the photosynthetic rate of a Maize leaf  in response to varying intercellular  CO2 concentration. In Maize (a C4 species), photorespiration is already suppressed by internal mechanisms, and supplying air with only 2% oxygen did not enhance net photosynthesis.


The simple modifications to the LI-6400/XT system’s operating procedure given in this Technical Tip provide one way to investigate photorespiration.  The results of the basic experiment conducted to test this technique confirm that photorespiration accounts for a substantial reduction in the net photosynthetic rates of C3 plants. Efforts by scientists to transfer some of the ability of C4 species to suppress photorespiration into economically important C3 crop plants continue to hold great promise.

Recovering from a bad zero or span; how to undo your doings.

Recovering from a bad zero or span; how to undo your doings.

The LI-7700 is an inherently stable laser-based analyzer that requires minimal user calibration during normal use. User calibrations can still be performed to validate instrument performance, and are required when changing the instrument from its warm to cold temperature operating range. The possibility does exist, however, to perform an incorrect user calibration, which will have undesirable effects on instrument performance.  The instrument software provides multiple ways to help prevent, and even undo, a bad user calibration to ensure high quality data. In this TechTip we detail four simple ways to undo what’s been done when a zero or span calibration goes awry.

1.     Are you sure you want to do this? When setting a zero or span from the Windows interface software, users are given the option of “aborting” or “committing” a zero or span after initiating it.

2.     Back to basics: A factory reset button is present in the Calibration menu that resets the instrument to a non-zero, non-spanned state. While this isn’t the state the instrument actually left the factory in, (as all instruments are shipped with verified zero and span), it will bring instrument response back to a reasonable range and provide users a good starting point for resetting things.

3.     It’s ancient history: All zero and span settings are stored in on-board memory on the LI-7700 and can be accessed through the instrument’s Finder application. On the Cal page, individual settings can be reset to a previous state using the “Rollback” button.

4.     If all else fails, there is always the manual way. From the Cal page in the Finder application, the Advanced button allows users to manually enter the zero and span offsets they want to use. The factory offsets can be found on the calibration sheet shipped with the instrument, or user-defined offsets can be found in the on-board calibration history.