Combined Calorimeter and Spectrophotometer for Observing Biological Reactions

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Abstract
A stopped‐flow fast reaction apparatus is described which permits chemical reactions with rate constants of 50 sec−1, or less, to be followed both spectrophotometrically and calorimetrically. The thermal detector is a single copper‐constantan thermocouple, 5μ thick, with a response time of 3 msec. Using a bandpass of 0 to 80 cps, the signal‐to‐noise ratio is 1:1 for a heat change of 100 μcal/ml. The apparatus is housed in a 30×30‐cm aluminum cylinder which is temperature stable to ±0.00001°C; reactants can be brought to within 0.001°C of equilibrium in less than 1 h. Acrylic light pipes are employed in the optical detection system to conduct a monochromatic light beam to the observation tube and to conduct the transmitted light to a photomultiplier. Solenoid valves control the mixer inputs; a third solenoid valve stops the flow in 2 msec. The thermal system and calorimeter were analyzed mathematically and two computer programs developed which make corrections for the response time of the system, correct the data for heat diffusion from the observation tube during the reaction, and calculate the first‐order rate constant. The dehydration of carbonic acid was used as a test reaction. Experiments run at 3.8, 18, 24.1, and 36.9°C, using 0.04 molar NaHCO3 and 0.02 M HCl gave results thermally of 3.12, 15.8, 22.7, 49 sec−1 and optically of 3.15, 15.1, 20.6, for the rate constant and 2100, 1413, 970, and 460 cal/mole for ΔH.

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