Component Selection in the top-performance, world-class OLIS CPL Solo
The CPL Solo uses the most stable light source available. Today’s LEDs -- operated with OLIS circuity – are effectively perfect, showing no change over days of continuous use, thus being absolutely ideal for lengthy acquisition periods as are common for CPL. Our agile company moved to LED excitation sources about three years ago for CPL and our single wavelength CLARiTY UV/Vis photometers. We have been designing our own circuitry for nearly 50 years.
The use of the LED as the excitation source allows positioning of the LED within millimeters of the sample. There is no means for a higher intensity light for exciting the sample than this, other than a laser. For laboratories choosing to do so, second and even third LEDs can be used to illuminate the sample, doubling or tripling the exciting light intensity at very modest additional cost. Each LED is optionally filtered to provide the desired bandwidth and removal of any spurious wavelength(s) emitted by some LEDs (‘stray light’ or ‘ghost light’).
The CPL Solo uses the highest throughput emission monochromator available: the OLIS RFL single grating monochromator. This small monochromator (20 x 22 x 12.5 cm) monochromator has one 40x45 mm grating and one 35 x 35 mm mirror. Recall, every optical surface that the light beam encounters absorbs roughly 5-10% of the photons. Clearly, the fewer the surfaces (mirrors, gratings, prisms) the light encounters, the higher the light remaining as output.Both additive and subtractive double monochromators are available and were considered for this use.
The additive Cary prism-grating monochromator has outstanding throughput in the 185-2600 nm and is used in highest precision absorbance and circular dichroism spectrophotometers. When this monochromator is coupled with a 150-watt xenon arc lamp, it becomes an ideal excitation source for the OLIS DSM 172 CD & CPL, where the prism-grating Cary monochromator supports absorbance and CD in the 185-2600 nm region and CPL excitation across the same broad range.Coupled to a dimly emitting sample, however, the many mirrors, lenses, prism, and grating in the monochromator render sensitivity far lower than through a monochromator with only a single mirror and single grating. The Cary has 5 mirrors in the monochromator (and another 6 within the chopperdetector chambers, which are used in absorbance applications) plus the prism and grating. See Addendum A, Optical Train of the Cary 17.
The subtractive double grating monochromator[1] by OLIS, known as the Hummingbird (Hummer) is used in the very popular OLIS DSM 20 CD, as well as OLIS DM 245 with a 150-watt xenon arc lamp. This monochromator is far smaller and more simple than the Cary prism-grating, having only two mirrors and two gratings (see Addendum B, Optical Train of the Hummingbird). However, to achieve its small size and modest price point, it remains a slightly lesser optical system than the outstanding Cary monochromator, and is less attractive as an emission monochromator in a CPL than a smaller, less expensive, higher-throughput single monochromator.
The subtractive double grating monochromator with moving intermediate slit, invented and patented by OLIS founder Dr. Richard J. DeSa, has such high light throughput that a 75-watt xenon arc lamp is common for most applications, including absorbance scanning to 1,000 scans/second. The most outstanding characteristic of this instrument is its millisecond spectral scanning, rendering it the most attractive monochromator for kinetic studies.This strength is not beneficial in very low light cases such as CPL, so this mode of operation will not be used, taking away the strongest argument for this premier monochromator. The six mirrors and two gratings in the monochromator absorb well over 50% of the incoming light, rendering it unattractive for low light level emission studies. When bright emission is available, however, the performance achievable is unique and astonishing. See, for instance, Addendum D, the result of a one second emission acquisition period from a single firefly. A firefly has an emission signal many orders of magnitude brighter than most CPL samples. Also shown, Addendum E, is a one millisecond absorbance scan of holmium oxide.
The CPL Solo uses the most sensitive detector available. Hamamatsu came out with this photon counter years ago, and it changed our opinion about producing fluorescence instruments. Its exquisite sensitivity comes in a tiny square box. https://www.hamamatsu.com/us/en/product/optical-sensors/pmt/pmt-module/photon-counting-head/index.html
For CPL and anisotropy – where we must capture not only the intensity but also the polarization status – we operate this detector as a gated photon counter. In this mode, we send data points to our FPGA chip, sorting each datum by time and thus polarization state, into two counters, one for left and the other for the right. When using the PEM, this left-right switching is occurring at 50 KHz. Keep in mind: a modulator does not flip perfectly from left to right, but produces a sinusoidal wave as it transitions from left to right, the peak and valley of which are maximum L and minimal R or the reverse. Our extraordinary ambitious data collection code – written decades ago for our digital CD data collection by Denghui Cheng[2] – reads and uses the entire waveform, calculating the answer using ten readings along each wave for every em(L) or em(R) data retained.
The CPL Solo UV/Vis employs the new Hinds PEM-200 modulator, the best commercially available modulator. The CPL Solo NIR employs a quarter-wave plate, the best modulator for 600-1700 nm.
Polarizers are the best we can purchase for the spectral range where they will be used. A 15 mm, grade-A Glen-Taylor polarizer (the best one available) is used for the UV/Vis model. A polarizer is optional at the excitation position – easily add or removed to study this phenomenon -- and is mounted at the entrance slit of the monochromator for emission. For the NIR model, a quarter-wave plate and grid polarizer are used.
Software is the mature, powerful, inclusive, and yet easily mastered OLIS SpectralWorks data acquisition/ instrument control Win 10 program. If you can conceive of a feature, you will find it in this excellent software package. This user-friendly, fast, and attractive program belies the muscle behind the screen. The code written for anisotropy in the 1970s was expanded to become the code for circular dichroism in the 1990s and for CPL in the 2010s. See Addendum F: The Math behind OLIS spectrophotometers’’ freedom from an analog lock-in amplifier (and G-factors)
An instrument is only as good – and no better – than its components. The OLIS hardware and software are exactly what you would choose, if you were to produce your own world-class CPL. The instrument is built a particular way because it is the best way.
Many alternative designs are available to a company with the expertise of OLIS.
However, for the most electronically simple, functionally specific, and most sensitive, we have the CPL Solo as described above.
A running joke amongst the engineers is “We’re always looking for light.” Maximum light in, maximum light detected: this is what makes a great spectrophotometer.
[1] For more on the “Subtractive double monochromator,” See Addendum G
[2] Horrifically, Denghui and his bride, Caixia, were killed in an automobile accident on their way to work in June 2011. One has to wonder what Denghui could have achieved for mankind next. Denghui Cheng Obituary - Athens, GA (dignitymemorial.com)
ADDENDUMS
Addendum A: Optical Train of the Cary 17 – An additive with many optical surfaces, albeit nearly peerless world-class performance, 185-2600 nm
Addendum B: Optical Train of the Hummingbird, our own small monochromator for UV/Vis use, 170-700 nm for CD or 200-850 nm for absorbance
Addendum C: Optical Train of the OLIS RSM 1000, our own UV/Vis rapid-scanning monochromator with gratings that can be exchanged between experiments for ease in optimizing for any spectral region 190-1100 nm (i.e., 300-530 nm, 500-960 nm, etc.)
Addendum D: One Second Emission spectrum from one Firefly using the subtractive double grating monochromator commercialized as the OLIS RSM 1000
Addendum E: One Millisecond absorbance spectrum of holmium oxide using the subtractive double grating monochromator commercialized as the OLIS RSM 1000
Addendum F: The Math
The math behind OLIS spectrophotometers’ freedom from an analog lock-in amplifier, valid for CD (and CPL[1]) spectrometers:
The measurement begins with the digitization of the voltage signals generated by the photomultiplier tubes (PMTs). These signals represent the time-dependent intensity of the light detected by each phototube. Once digitized, the CD information is extracted from these signals in an entirely digital manner using mathematical manipulations derived by rigorous analysis of the optical system.
The digitized signal consists of a DC level, a 50 kHz signal, and noise. The digital processing derives the CD amplitude from the DC and 50 kHz signals while repressing the noise.
Step one is to measure the DC component of each signal by computing the average of each signal. This DC level is derived from the same data as the 50 kHz signal, so there is no need to deal with independent amplification of each component and thus no need for a photometric calibration using a chemical standard.
With the DC amplitudes known, second step is to normalize each signal by dividing each by their respective DC level. The resulting signals have DC amplitudes of unity with normalized 50 kHz components. The CD information is derived from these normalized signals; thus, the absolute DC amplitude of either signal before normalization is of no consequence to obtaining the right answer:
CD by first principle and without lock-in amplifier and associated calibration constant, k.
[1] CPL = em(L) – em(R), whereas CD = abs(L) – abs(R). In the case of CPL, this entirely digital method also obviates the need for G-factors.
We close with a quotation from W. Curtis Johnson's chapter in Fasman's 1996 book, CD & Conf Ana of B (Plenum Press):
“There are three methods for measuring the CD of a sample. The most straight-forward is to measure the absorption for each rotation of light and subtract directly the measurement for right circularly polarized light from the measurement for left circularly polarized light. However, this requires making each measurement to great accuracy, which was not practical until the recent availability of powerful and inexpensive computers...Inexpensive, high speed digital computers have made direct subtraction of left and right circularly polarized beams a practical method of measuring CD. This method is pioneered in a commercial instrument for electronic CD by On-Line Instrument Systems (Olis, Bogart, Georgia)..This dual beam collection and direct subtraction method has two advantages in addition to the flat baseline. First...CD signals are measured correctly. Second, the instrument is measuring absorbance directly, so there is no constant of proportionality to calibrate.”[Quoted from pages 640 and 644-5, underlining added]
If you do not have this book, Johnson notes the two other means of measuring CD are: “to measure the ellipticity imparted on linearly polarized light that passes through the sample. This method is limited by the quality of the linearly polarized light and the mechanics of measuring the small perpendicular component of the elliptically polarized light. The third method is to modulate the light between the two rotations, and measure the difference at each wavelength. This method is well suited to analog electronics, and is the method used by most commercial instrumentation.”
Addendum G: Attributes of a Subtractive Double Grating Monochromator
Two key attributes of the subtractive double monochromator are its production of a homogenous output beam (useful to us and thus addressed here) and zero temporal dispersion[1]. The subtractive double is used in Raman spectrophotometers for these reasons. Both the OLIS RSM 1000 and OLIS Hummingbird monochromators are subtractive double monochromators and thus produce a homogenous output beam, regardless of bandwidth.
Output beam of an additive with 6 nm bandpass
Sample is illuminated with spatially separated wavelengths. So, if the sample is a solid or a suspension, literally different experiments are happening along the length or breadth of the sample, unintentionally. One portion sees 278 nm light, another 282 nm light, etc.
Output beam of a subtractive with 6 nm bandpass
Sample is illuminated by a homogenous blend of all the wavelengths. What one portion of the sample sees is identical in every way to what other portions of the sample sees. You are conducting one experiment.
[1] The length of time for the incoming light to exit the monochromator is independent of the wavelength.