Tardigrades, carrot carotenoids and the DIY Raman spectrometer (VI)

It goes without saying that the ordinary microscope will remain our primary instrument for tardigrade studies. Nevertheless, a spectrometer might contribute some interesting information: see February and March issues of our magazine.

Spectrometers are included in some of our school education plans - in theory. But when asking around among your friends you will notice that the individual amount of spectroscopic knowledge is actually very low, mostly non-existent. We think that a basic understanding of spectroscopy should be considered as an essential part of general education: spectroscopy provided virtually all information about the chemistry of distant stars, about their genesis and final disappearance and about the expansion of the universe. Spectroscopy helped to understand the properties of infra-red and ultra-violet light. Nevertheless many teachers think that it might be much more important to learn poems by heart or to memorize the biographical data of some classical authors ... a pity.

All this makes up our justification and motivation for the tiny lecture in practical spectroscopy which is going to follow here. Besides, spectroscopy is essential for a basic understanding of our continued Raman experiment:

Fig. 1: The context - a further sequel to our Raman DIY spectrometer experiment. As already explained previously, Raman radiation is generated by an interaction between laser light and the material under investigation. Those Raman signals are tremendously weak but they bear information which can be used for chemical analysis of many materials. In our experiment we are sending strong laser light through the microscope objective upon the surface of the sample. Most of the laser light will be simply reflected from the sample's surface, thus remaining useless. But a small percentage will come back with a so-called Raman frequency shift. The strong, simply reflected laser signal and the weak Raman signals will pass the microscope objective once more on their way back and will finally arrive at the top of the microscope photo tube. At this point we are catching the sharply focused laser beam by means of a hole within an x-y-positioning plate (1) which serves as an entrance towards the spectrometer light guide (3) which again is passing the signals on to the spectrometer (4).
Please make sure that the sharply focused laser cannot reach anbody's eye! So please, use eye-piece protective caps or direct the eye-pieces' output to a close wall in order that nobody can be harmed by the laser radiation!

Basically that's already the complete Raman experiment. But the history of scientific spectrometry is interesting and even fascinating as well. It appears quite rewarding to look back a little bit in history and to enjoy a glance at the equipment used at those times when lasers, interference filters and CCD electronics were yet non-existent. Besides, when asking around which kind of scientific instrument might have been most important in the history of mankind, you will inevitably end up with two answers: the telescope and the microscope. The third extremely most important invention, namely the spectroscope, will never be mentionened though the microscope and the telescope both have taken profit enormously from spectroscopy (both the microscope and the telescope can be used for spectroscopic analysis!). The reason behind the non-understanding of spectroscopy is probably that telescopes and microscopes are delivering images which can be easily understood, sometimes just by looking at them. No physics and no math are necessary. In contrast, spectroscopic data are rather abstract and have to be translated in order to understand and make use of them.

Fig. 2: The pocket spectroscope is a precursor of the modern electronical spectroscope. There is an entrance slit which can be adjusted in order to let pass various amounts of light: when made slim the spectral lines will appear sharp but dim whereas a wider slit will admit more light at the expense of sharpness (bright lines will appear broad, not crisp). The center of the tube contains a chain of three (sometimes five) glass prisms which are forming a prism chain, splitting up the entering light in a rainbow manner, as can be seen on the next image:

Fig. 3: View through a pocket spectroscope, in this case an instrument with am in-built wavelength scale. You will notice the typical sequence of the rainbow colors - together with their respective wavelengths between ca. 400 and 700 nm. On the left side you would enter the (invisible) world of infrared radiation, on the right side the world of ultraviolet and gamma-radiation - both of which are invisible for us. As can be seen from the image it is difficult to tell the exact wavelengths' numbers and there appears to be no way to measure a specific intensity at a given wavelength.
The spectrum shown is the one which appears when directing the spectrometer towards an Almandine gem lit by strong white light. The Almandine is absorbing strongly at three positions in the spectrum. Therefore the respective regions will appear less intense than the rest of the spectrum which didn't interact with the mineral (the dark absorptions occur at ca. 505 nm, 525 nm and 575 nm). So you might use this instrument e.g. to tell apart Almandine from red glass at a flea market.
There were some other famous applications used in the past: people could direct the instrument towards an approaching cloud. The intensity of the water lines within the spectrum of the cloud helped to make predictions about the water content of the cloud. As a consequence some rudimentary local short-term weather prognosis became possible.
Furthermore, when directing the instrument towards the blue sky you will notice some faint black lines, the famous Fraunhofer lines (which helped to investigate the chemical composition of the sun). Furthermore this type of simple spectroscope was used in forensics, in order to tell apart red ink stains from blood stains.

Fig. 4: Special spectroscope for chemists. A tiny test tube holder and a mirror are integrated within the instrument's design thus allowing to superimpose a sample's spectrum with some kind of external comparison spectrum, e.g. from a flame. The image is showing a test tube with an orange carotene solution. But, frankly speaking, the knowledge gain is not very impressive in this case, as the carotene will merely absorb broadly in the blue and blue-green part of the spectrum ...
This is one of the reasons why we are building the DIY Raman spectrometer which will render sharp signals when encountering carotene in a given sample.

The bad news for the friends of the historic instruments is: once you will have worked with an electronic (quick, precise, solid-state) spectrometer chances are low that you will continue to use your old-fashioned, purely optical spectrometer. It is a pity as many of those old spectrometers are so much beautiful!

Fig. 5: The instrument used for our Raman experiment was sold by a company called Science-Surplus which is based in the U.S.A.The price was ca. 250 US$ including overseas freight. It came as "unadjusted " but worked perfectly out of the box. The instrument has to be connected to an old-fashioned "RS232" (serial) port. But this is not a serious drawback (at least as long as you have this type of connector on your computer). The instrument was delivered with a standard 1.800 lines per millimeter grid, a value which is quite ideal for our Raman experiment.

Fig. 6: View into the Science-Surplus spectrometer. There is also a cooling fan inside (the black item in the center of the image, the one with the parallel grooves). It can be activated but we do not recommend it: you will achieve an improved detector performance (less signal noise) but at a price: the ventilator is rather loud and you will run into serious condensation risks.

Fig. 7: View into the "heart" of a spectrometer, the so-called spectrometer cell. The light is entering through a small slit into the optical cell and is split up into its range of wavelengths by means of two mirrors and a diffraction grid, thus ending up as a rainbow, similar to the one shown in fig. 3. This rainbow pattern is being projected onto the CCD chip (the chip with the silvery legs on the right) which is made up by a linear chain of ca. 2,000 sensor cells. Each of those CCD pixels is able to measure the light intensity at its specific position (which is equivalent to a specific wavelength as well). The rest of the job is being performed by a computer.

© Text, images and video clips by  Martin Mach  (webmaster@baertierchen.de).
The Water Bear web base is a licensed and revised version of the German language monthly magazine  Bärtierchen-Journal . Style and grammar amendments by native speakers are warmly welcomed.

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