For kinetics applications, the diode-array technology of the HP 8453 UV-visible spectroscopy system offers substantial advantages over conventional scanning spectrophotometers. It can measure full spectra (190-1100 nm) at up to 10 spectra per second.

UV-visible biochemical analysis software for the HP ChemStation includes time-based data acquisition and basic kinetic evaluation (zero- or first-order). An open-sample-area design makes it very easy and convenient to use third-party accessories.

The chemists at Applied Photophysics Ltd., a comarketing associate of Hewlett-Packard, are specialists in stopped-flow kinetics measurements. Their RX.2000 rapid-kinetics accessory is used to obtain rapid-mixing, stopped-flow kinetics data. It is robustly built and simple to use. It offers sample economy, a fully thermostatted flow circuit with a flexible armored umbilical connection to the triple-annealed observation cell (2- and 10-mm path lengths), and an 8-millisecond dead time. Applied Photophsysics also offers the Pro/K software, a powerful global analysis package for advanced kinetics data evaluation.

Configuring the RX.2000 accessory with the HP 8453 spectroscopy system is simple:

• Fit the observation cell into the cuvette holder of the HP 8453.

• Connect the HP 8453 GPIO port to the RX. 2000 DIN socket.

• Load the data conversion macro to transfer data easily from the HP ChemStation to the Applied Photophysics Pro/K software.

No other changes to hardware or software are required to operate the HP 8453 and RX.2000 together to obtain rapid, high-quality stopped-flow kinetics data.

In the example shown here, the RX.2000 was used with an HP 8453 and an HP ChemStation with UV-visible biochemical analysis software for time-based data acquisition. The 2-D display shows spectral changes in the range 500-580 nm during the reduction of cytochrome C (6.5 mM) from the ferric to the ferrous iron state by ascorbate (1 mM) in tris buffer pH 7.0. The rise in absorption at 550 nm is indicative of the reaction which occurs over about 30 seconds. The data file obtained with the HP 8453 was converted to Pro/K format.