A Gallery of l - Arginine Photomicrographs

(using a variety of illumination techniques)

by Brian Johnston   (Canada)

L-arginine is an amino acid present not only in humans, but in all life forms.  It plays a part in the synthesis of proteins on which the function of all living cells depend.

This fairly simple organic molecule is composed of carbon, hydrogen, nitrogen and oxygen atoms as can be seen in the structural formula.

The space-filling model gives a better idea of what the molecule would “look like” if we could see it.  (Both images were produced using Hyperchem software.)

Although the melting temperature of this white crystalline solid is low enough ( 221 degrees Celsius ) to enable melt specimens to be prepared, I decided instead to use its high solubility in water to produce crystals by evaporation.  A quantity of the solid was dissolved in several millilitres of cool distilled water to produce a fairly concentrated solution.  A couple of drops of this solution were placed on a clean slide and the slide was placed in a cool dust-free location.  After all of the water had evaporated, the slide was checked under the polarizing microscope to see whether any photogenic structures were present.  If the slide was a good one, it was allowed to dry completely for a further week and then a drop of the mountant Permount was placed on the crystal layer and a cover-slip carefully added.  (This final step allows the slide to be kept in pristine condition for a long period, but it has the side-effect of turning the crystal layer a pale yellow-orange colour when viewed by normal transmitted light.) 

Note that l-arginine may act as an eye or respiratory irritant, and should therefore be handled with care.

I must admit that I prefer to produce melt specimens of compounds because there are fewer variables that can act to the detriment of the finished product.  In specimens produced by evaporation of the solvent,  the concentration of the solution is of critical importance.  If the solution is too concentrated, unsightly amorphous lumps of crystal tend to form.  If the solution on the other hand, is too dilute, the thin crystal layer that forms tends to produce shades-of-gray images under the polarizing microscope, instead of the colourful forms desired.  Only experimentation will find the ideal parameters.  In the case of l-arginine, a relatively concentrated solution seemed to work best.

The images in the article were photographed using a Nikon Coolpix 4500 camera attached to a Leitz SM-Pol polarizing microscope.  Images were produced using several illumination techniques:  transmitted light, dark-ground illumination, phase contrast and polarized light.  Crossed polars were used in all polarized light images.  Compensators, ( lambda and lambda/4 plates ), were utilized to alter the appearance in some cases.  A 2.5x, 6.3x, 16x or 25x flat-field objective formed the original image and a 10x Periplan eyepiece projected the image to the camera lens.

The image to the left below shows a field near the edge of the cover-slip illuminated with transmitted light. On the right is the same field using polarized light.  It is interesting that much of the material visible in the left image does not appear in the right image.  This material is referred to as isotropic since light travels through it at the same velocity in all directions.  Anisotropic material, (which forms each circle in the right hand image) is referred to as birefringent  because it splits the light rays into two sub-rays which travel through the material at different velocities.  This phenomenon (which depends upon the packing of the molecules in the crystal) results in the material being visible between crossed polars.

The image to the left below shows an area near the centre of the cover-slip, where crystal growth has begun at a large number of sites which are close to one another.  The circular growth fronts have run into one another producing the strange black intersection lines that can be seen in the higher magnification image on the right.

Under still higher magnification, the detail in the circular structures can be seen more clearly.

Compensators can be used to alter the appearance of anisotropic crystals by slowing the velocity of one of the sub-rays travelling through the material.  In the example on the right below, a lambda/4 compensator was used beneath the crystal field and a lambda compensator was positioned above.  This results in the brilliantly coloured circles and background.

At other locations, (usually near the edge of the cover-slip), different crystal formations sometimes occur.  The feather-like structures below are illuminated by polarized light with no compensators (on the left), and with the addition of a single lambda compensator (on the right).

A low power image of a field between crossed polars, and with a lambda/4 compensator, shows strange wave-like areas between the circular forms.

The two higher magnification images below show the ‘waves’ more clearly.

Sections of the slide near the cover-slip edge sometimes show ‘flow’ patterns.  These can be seen in the two images below.  The one on the left uses normal transmitted light illumination, while the one on the right uses polarized light with crossed polars and two compensators (lambda/4 and lambda).

Large areas of one slide preparation displayed the structures shown in the three images below.  These patterns appear almost ‘fractal’ in nature.  The black and white image uses dark-ground illumination while the other two use normal transmitted light.

The image on the left below uses dark-ground illumination.  The higher magnification image on the right, of another location, uses phase-contrast illumination to distinguish finer structural details.

This final image shows some rather amorphous circular structures illuminated with a dark-field condenser.

Evaporation specimens, with their many controlling variables, tend to produce very different results when several slides are prepared under different conditions.  This was certainly true of l-arginine.  My favourite field is shown as the first image of this article.  It reminds me of the false-colour image of a martian landscape.  Images like this one are what make this hobby so rewarding!

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