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

In the previous issue of our magazine we were able to demonstrate the detection of tardigrade carotenoids by means of a DIY Raman microscope spectrometer. In order to check whether the carotenoids might simply stem from food intake we can add a Raman spectrometer look at moss plants:

Fig. 1: Results from the DIY Raman microscope spectrometer (cf. previous issue #200).
- Red curve: Raman spectrum of a 50 µm sized Echiniscus sp. "tun" (dry state tardigrade)
- Orange curve: Raman spectrum of a slice of carrot
- Green curve: Raman spectrum of a moss plant
As you can see the carotionoids are present in the moss plant as well (the green exclamation marks indicate the two main carotenoid bands which are weak but do occur at the correct Raman Shift values.

This is a fine result! Additionally the green curve is indicating the presence of other chemical compounds. We think that the huge band on the right hand side of the spectrum could be attributed to the chlorophyll variants within the moss. Possibly the carotenoids are protecting the chlorophyll against UV attack and oxidation as their double bonds are able to absorb potentially destructive photons:

Fig. 2: Structure formula of the most common carotenoid (ß-carotene). This molecule has many neighbouring (conjugated) double bonds which are able to bond oxygen. Though the carotene might help to preserve the dry state tardigrade humans should avoid excessive carotenoid consumption. Some very prominent people's faces are said to be colour shifted to orange because of carotenoid additives.

In summary and in comparison with the microscopic evidence we might conclude that the Echiniscus tardigrades are treating the chlorphyll and the carotene in a very different manner: the chlorophyll appears to travel primarily through the stomach-intestine-pathway, whereas the carotene will be found in the body cavity outside the stomach-intestine-region, too. Some part of the chlorophyll might even remain undigested as we can see from the still green "garbage bags" which are left back during the moulting process.

With respect to the DIY Raman spectrometer we will proceed slowly. You will find excellent internet reports by people who have built their own Raman spectrometers everywhere in the internet. But nevertheless the DIY Raman remains a challenge. The individual situation might be very different because of:

-- Different demands (measurement range, sensitivity, precision, resolution)
-- Different microscopes to be used as a basis for the DIY construction
-- Different budgets
-- Locally different access to filters, laser modules and spectrometer modules
-- Last but not least: different safety requirements (laser safety, e.g. in schools!)

As a consequence we are going to present a rather simple, almost generic construction plan. It is essential to understand the basic principle first and to become creative second. In its simplest form a Raman microscope spectrometer can be described as follows:

>> Strong laser light, precisely focused on a tiny object area
>> 99,9 % of the light is not absorbed by the sample. Only a very small fraction of the photons is coming back loaded with Raman information
>> After filtering out the unchanged (useless) laser light by means of a so-called long-pass filter the Raman signals are coming out
>> These have to be wavelength-splitted by means of a classical light spectrometer
>> and in the end there might be a spectrum, hopefully!

Compared to e.g. classical infrared spectroscopy the "Bauplan" is rather simple and is consisting of relatively few components which are:

Solid-state laser (strong laser pointer) - focusing optics - sample holder - laser rejection filter - focusing optics - VIS spectrometer module.

More details will follow.

© 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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