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rapid Synchrotron Radiation Circular Dichroism (rSRCD)

A plane mirror after the M3 horizontally deflecting mirror in the AU-AMO beamline can be moved to direct the monochromatised light into the rSRCD branchline.

The figure above shows a schematic overview of the beam line. The source of this beam line has a 2.4 m long undulator with a magnetic period of 53 mm, which, in combination with the relatively low electron energy of the ASTRID2 accelerator of 0.58 GeV, allows the generation of intense UV light up to a wavelength of 290 nm. The small divergence and narrow width of this UV light source makes it well suited for a stopped flow instrument, where the internal light beam width must be kept below 1 mm. This can be achieved without any further optics than those in the AU-AMO monochromator which selects the UV wavelength of interest. Note that this wavelength can be freely chosen below 300 nm, and is not restricted as more conventional light sources are, where intense single lines of a lamp limits the wavelength to certain discrete values. The UV light from the AU-AMO monochromator is directed into the rSRCD branch line via a flat MgF2 protected aluminium coated mirror. This mirror has a sideways, as well as a reflection angle adjustment, which makes easy alignment of the light into the stopped flow instrument.

After the exit slit in the rSRCD branch line, which selects the wavelength from the grating in the monochromator, the light passes through a high quality excimer laser graded CaF2 window (Vacom, Germany). The window is cut perpendicular to the <111> crystal axis of CaF2 to minimize birefringence, which would otherwise change the polarization of the light. This window separates the ultra-high vacuum of the AU-AMO beam line from the nitrogen-purged environment of the stopped flow system.

The polarization of the light from the monochromator is nearly fully horizontally polarized, but to ensure 100% linear polarization, the light passes through a MgF2 Rochon polarizer (B.Halle GmbH, Berlin, Germany). The following optical element, a Photo Elastic Modulator (PEM - Hinds, Oregon, USA), converts the horizontally polarized light into alternating left and right handed circularly polarized light with a frequency close to 50 kHz. Thus, the CD signal of a chiral sample will oscillate in phase with this frequency, and can be detected using a lock-in amplifier.

Stopped flow instrument

The stopped flow instrument chosen for the rSRCD facility is a model SX20 from Applied Photophysics Limited (APL - Surry, UK), which can perform multiple fully automated mixing shots. The photomultiplier tube (PMT) and data acquisition electronics from APL records CD signal, with time resolutions down to 0.1 ms. The APL electronics controls both the PMT high voltage and the PEM, using the DataPro SX20 software. The wavelength of the beam line is set via the ASTRID2 control system software (ConSys), which can set both the monochromator and the undulator to the desired wavelength, as well as directly access the APL software for setting the PEM to the correct setting for CD measurements.

The stopped flow cell in the SX20 stopped flow system is a clear quartz cell; mixing of dark and clear quartz, often used in fluorescence and UV-VIS absorption stopped flow instruments, is avoided to minimize stress-induced birefringence in the quartz. This would otherwise interfere with the circular polarization state of the light, and thus might lead to incorrect CD measurement. Careful masking of the clear quartz avoids light passing through the quartz, and thus not through the sample, to reach the PMT detector, which otherwise can led to a falsely low apparent CD signal of the sample. The quartz cell can be oriented in two directions, with either 2 mm or 10 mm light pathlength. The former is used for most experiments. The two sample syringes and the quartz cell can be submerged in a water bath to set the sample temperature using a water cooler/heater. Typically the temperature can be set between 5°C and 50°C, with temperature oscillations of about 0.1°C. The two mixing syringes of the SX20 system can readily be exchanged for either two 1 mL syringes or one 2.5 mL and one 0.25 mL syringes, thus providing either 1:1 or 1:10 mixing.

Flux

The flux of light entering the SX20 stopped flow system has been measured with a Si diode (AXUV100G, Diode Corp). At a wavelength of 195 nm the flux from the rSRCD branch of the AU-AMO line is 5x10^13 photons per sec. into a beam size of 1x1 mm2. As this beam size is smaller than the stopped flow full aperture of 6x1.5 mm2, all of the flux enters the cell. In comparison, flux measurement carried out on a new high flux Xe-arc lamp based conventional CD instrument, gives a flux density at 195 nm of 2.5x10^10 photons per sec per mm2 (full beam size 8x11 mm2). This corresponds to roughly 2.5x10^11 photon per sec. entering the stopped flow aperture at a (comparable to the rSRCD) bandwidth of 1 nm. Other light sources than Xe-arc lamps are frequently used on conventional CD instruments for stopped flow applications. These include Xe-Hg lamps and even light emitting diodes (LEDs). Xe-Hg lamps have intense, but discrete, lines, and LEDs are single line light sources, which may increase the flux by up to a factor of 10. However, none of these higher intensity light sources are available below 200 nm. Therefore, the light flux which enters the rSRCD stopped flow cell is at least 15 times, and below 200 nm at least 100 times, higher than available from the best commercially available CD instrument/light source combinations, corresponding to more than 4 to 10 times better signal to noise ratios.

Last Modified 05 August 2019