HOW  MANY  ONION  SKINS  ARE THERE?

Walter Dioni    -    Cancún, México

When we speak of "onion skin" it seems that everyone knows what we mean. But in reality the onion, which is the bulb of Allium cepa (and a species with several varieties), has many "skins", as this simple check will show

It's evident that there are many skins: every scale has two.

FIG. 1, 2 (Fujifilm FinePix A900 Camera)

Fig. 3 – the mesophyll is composed of large polyhedral cells. As this is an underground leaf, the mesophyll lacks the chlorophyll of the aerial leaves. In comparison, the epidermic cells, here seen in transversal section, look tiny. Mesotome section; mounted in glycerin. 1:1 cut from the original 3 Mpx picture. Canon Powershot A75

As you can see (fig 2) each onion bulb scale has two epidermal layers, one internal and one external. Almost all protocols of practical exercises that use "onion skin", makes it clear "to use the skin from the inner side of the bulb scale."  Would both be different?

                 Fig 4 – Inner epidermis – without stomata                                                                 Fig. 5 – Outer epidermis– with stomata

No stain applied, mounted in glycerinated water – 10x objective – 1:1 crops from the 2 Mpxs originals, additionally reduced to be included in the article.

Fig 6 – 40x obj. – Inner epidermis - Water mount, no fixatives. Rheinberg disc (black center, orange border). The cytosol is seen as a gel with clouds of granulations (probably mitochondria).
Canon Powershot A75

The following pictures (6 and 7) shows a bigger image of one stomata (x40, reduced to be included here) and a topographic view of the outer skin (x 10, also reduced; the “stained glass” effect is due to an irregular staining with 1% aqueous methylene blue). It does not have scientific value but shows what I want.

Fig 7 and 8 – Outer epidermis – two images by the DC3 Motic camera. The methylene blue solution has not permeated the cell walls, and the nuclei are not stained.

What are these differences?  I think they are easy to see. The outer epidermis has cells of many sizes, with an irregular design, and has stomata (not excessively frequent in this case, but undoubtedly present). Note also that at least four of the cells in this tiny fragment of the surface of the scale (fig 5), has 2 nuclei.

The inner one (fig. 4), on the contrary, has the architecture we are used to assign to the “onion skin”. The skin that is shown to us over and over ... over decades. Narrow elongated polygonal cells in a well builtstructure, each cell with one nucleus, and its nucleoli (1,2 or 3).

Fig.9, left, onion bulb drawing, in Bastin, 1895 – Fig.10, right, onion, with its foliar “stem”, and a fully developed leaf that normally we don’t see , Bentley, 1887.

The scales (food reserve leaves) form the bulb of the onion (the underground part of the plant). But if you compare it with leeks (Allium ampeloprasun) although it has no bulb, you see the same concentric structure on the stem, and you could see that the "white part" is continuous with the "green leafy part" generally discarded by chefs, and which in onions is cut flush to the bulb by the vendor. (Our fig. 1, and also fig. 8, from Bastin 1895). The green part, if it could grow freely in the plant, could become long narrow leaves that by its own weight would bend to the outside, (fig 9, from Bentley, 1887).

In such a position, which so far we call "the outside", would be "the underside" of the leaf. A lightweight review in your Botany book providing elementary notions of the structure of the leaf, will tell you that it is at the “inferior face” where there are the stomata, exclusively, or more numerous, than at the "top side” of the leaf.

Leaf sheaths (scales) have shown with their stomata that they actually are modified onion leaves, and that its external face, which have the stomata, is equivalent to the underside of an aerial leaf.

By using the "inner skin" the biology teacher focuses the student to the study of the plant cell, and one plant tissue, the epidermis, without having to divert their attention to explanations about the stomata, its structure and its function, nor on the homology between a bulb scale and an aerial leaf. It is a useful trick.

And... the third onion skin

Normally the other "onion skins" usable for scholarly lectures (the outer papery layers or tunics), are reserved for more advanced courses, in which it is studied the secretion by plants of different chemicals, especially calcium oxalate, which form clearly visible and very interesting crystal structures.

To see them, cut with scissors a 1.5 x 1.5 cm of the tunic (the upper portions are richer in crystals) and hydrate it in plain water for many hours. If colored, the tunic would diffuse some color (it's a useful pigment that people use to tinge textiles or Easter eggs) and become pliable. The more pigment lost, the better image you have. It only remains to make a wet mount.

Really the tunics are not an epidermis. They are the compressed dry remnants of one, two or more exterior scales of the bulb. Some focusing would convince you that the crystals are embedded in the dry mesophyll. This is why also you can see, as in any leaf, the veins with the vessels that conduct water through the scale. The “skin” for this picture (fig. 14) was first embedded in nail polish solvent, and mounted in nail polish, to make it more “transparent”. (3 pictures stacked with CombineZ)

Fig 11 to 13 shows crystals of oxalate found in the tunic of a white onion. Fig 14 – conductive vessels, white onion (all pictures with 40x obj.) Logitech 9000

Shallot’s tunic, fig 15 – 10x; 16, 17, 18 – 40x – Logitech 9000 - 800x 600 cuts, reduced to publish it here.

Fig 19: Some additional pictures of individual crystals in the tunic of a white onion and a shallot. Clips from 2 Mpx pictures taken with the Canon Powershot A75. (obj x 40) but the central row ones, which were taken with the 100x, these 3 pictures were cropped and reduced.

In all the pictures crystals are embedded in the mesophyll, and are seen through the superficial layer of the epidermis. As my microscope is not a DIC with the power to make net optical sections, the images are not very sharp.

Fig. 20: 10x  obj.– distribution of crystals (mostly one in a cell) in the mesophyll. Shadows of the limits of the long cells of the epidermis are seen, out of focus. Canon Powershot A75

In other plants are also common calcium carbonate crystals. Why do I know that the onion ones are calcium oxalate? Because it's said (by microscopists from the 18 century to now) that is not difficult to establish a diagnosis, using two chemical reactions, which can be controlled with the microscope:

In principle it is interesting that, fortunately for us microscopists, these crystals are insoluble in water, alcohol, acetic acid, or even in the hypochlorite, which is commonly used to destroy the cytoplasm and clean plant sections to be colored and mounted. This is why we can see them easily.

But...

Oxalate crystals are insoluble in acetic acid, but soluble without effervescence in hydrochloric acid

The crystals of carbonate by contrast are soluble in acetic acid and hydrochloric acid, with effervescence

As in my samples I submit to, and they resist the acetic acid, I can say they are oxalate, but ... I don’t want to risk my microscope objectives, exposing them to vapours of hydrochloric acid, so, please, believe me, as I believe all the botanists that spoke about these crystals from the 18th century to today, that they would dissolve readily. Amen.

What are the calcium oxalate crystals for? Included as they are in the external wrapping (the “tunic”) of the bulb, it is generally considered that they may be, by its taste, and its sharp edged structure, an element of deterrence for those underground invertebrates that could be tempted to attack the onion. And possibly they are also a form to sequester, and to turn inoffensive, the oxalic acid produced by the plant metabolism itself.

 The preparation I made to see the crystals of the white onion, give me an unexpected gift. It's not uncommon when seeing the epidermis with the higher powers, that you suspect to be viewing the plasmodesmata. These are cytosol connections between adjacent cells made through little holes in the cell walls. These are pictures (with the 40x obj.) of the plasmodesmata piercing the thick wall between cells of the tunic’s mesophyll.

Fig. 21 and 22: plasmodesmata in mesophyll cells

Another interesting trait was the very long cells, that build the upper (internal) tunic’s epidermis .

The above picture (fig. 23) is a montage of 3 individual pictures showing the special epidermis of the tunica: cells are very long and narrow. See the position and size of the nucleus that have to manage the long cell in the second row from the bottom. It is out of focus because my interest was to show the cellular walls. Original pictures taken with 40x obj. with the Logitech 9000

  THE  END

 OH! NO, NO, NO, OH! NO – it can’t be the end !

 Uh! Yes! Yes! There must be some additional words. I must not forget a fourth kind of onion epidermis: the information-rich epidermis of the red onion!. You know: beside being an onion epidermis with all the details we have discussed before, it has the sap in the central vacuole of the cell, tinged with an anthocyanin red pigment (write anthocyanin in your browser). It is the favorite to those teachers that must teach “OSMOSIS” and “PLASMOLYSIS”. I give you the “home work” of digging out of your books or from your browsers the knowledge of what osmosis is... if you don’t have learned it before.

 This has been treated before in MICSCAPE by Mike Samworth, (a long time ago in terms of this electronic era)

 ( http://www.microscopy-uk.org.uk/mag/art97/maysnp2.html)

 but I like to add my own two sand grains. There are more in Internet, I see at least 14 related images. And at least one valuable video on YouTube.

 My first surprise, working with the external epidermis (the only colored one, the inner one has no color) of my beautiful and big red onions, was that it was very difficult to obtain the normal peel which is so easy to get from the white onion, even from the outer epidermis. I noticed also the faint color of the peel, and that there are only a few nucleated cells.

 After an extra couple of attempts I realized that the cells were broken open, and I have in my slides only the upper wall of the cells, with some adhered cytosol, and the nuclei that by chance were near the upper wall. Most of the cytosol, and the colored sap in the vacuoles were left clinging to the mesophyll.

Fig 24: 40x obj. - a water mount of the outer epidermis of a scale of red onion. The nucleus is perfectly round and with a clear limiting border. Very faint color. Probably there is only half a cell here. Compare with fig. 26 – Canon Powershot A75.

Fig 25: a peel with only the upper wall of the cells with a very thin layer of citosol, no nucleus.

People don’t tell about this when explaining the protocol of the laboratory practices. The following two pictures (26 and 27) were taken from a thin tangential section of no more than 0.5 square centimeters that interest not only the epidermis, here shown, but a thin section of the mesophyll, which, being out of focus is not showing here. The short focus deep of the 40x allows this. Pictures were taken with the Canon Powershot

To see the famous plasmolysis you must prepare a concentrated sodium chloride solution (kitchen salt you know). It is enough to make a solution with 1 teaspoon of salt (more or less 6 g) in 100 ml of clean freshwater, and replace the water under your coverslip with the salt solution using the usual trick of putting a drop of salt solution at one border of the coverslip and absorbing water with an absorbent paper from the other side.  In no more than 10 minutes the effect is totally displayed. In YouTube there are some good videos, showing not only the plasmolysis but also the deplasmolysis when returning the epidermis to fresh or distilled water.

Can the onion skin give us more fun, for so little money? ...yes! it can ... see the next article.