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Although a major part of histotechnology is based on the staining of proteins in one way or another, proteins are not the only biological material with which we are concerned. Both carbohydrates and lipids are significant in both healthy and disease states, and it is important that their demonstration be reliable. Before that is possible, an understanding of the nature of the materials is necessary. This page discusses the structure of carbohydrates as they relate to human histology. It is not an exhaustive explanation, but will provide the background for demonstrating these materials.

Carbohydrates are carbon compounds with hydrogen and oxygen in a 2:1 ratio, from which the name is derived, i.e. watery carbon or hydrated carbon. The 2:1 ratio of hydrogen to oxygen is, of course, H2O, the formula for water. The term carbohydrate is also used to refer to compounds derived from these by oxidation or reduction, by the addition of amino, carboxyl and phosphate groups, or by their being linked with proteins and lipids.


The basic unit for carbohydrates is a sugar. There are numerous examples of sugars, which are more properly called monosaccharides and have the general formula of Cn(H2O)n. The prefix mono means “one” or “single”, while the word saccharide means it is a sugar, i.e. it is a compound composed of a single sugar. The ending -ide in this context means “in the class of”, so a monosaccharide is a compound composed of a single molecule of sugar and classed as a carbohydrate.

In addition to “mono”, other prefixes which denote numbers may be encountered. The prefix di- means two, so a disaccharide is a carbohydrate composed of two sugar molecules, and tri- denotes carbohydrates with three sugar molecules i.e. a trisaccharide. The word Oligosaccharide means “a few sugars” and refers to carbohydrates with several sugar molecules. Polysaccharide means “many sugars” and refers to carbohydrates composed of large numbers of sugar molecules, such as glycogen, cellulose or starch.

The names of carbohydrates usually end with -ose, e.g. sucrose, which is more commonly known as table sugar or cane sugar, and is a disaccharide composed of one molecule of glucose and one molecule of fructose. Due to the fact that they contain hydroxyl groups, sugars are sometimes referred to as alcohols or ols.

If sugars contain an aldehyde (-CHO), they may be referred to as aldoses or aldehyde sugars. If they contain a ketone (-C=O), they may be referred to as ketoses, keto or ketone sugars. Of the two formulas shown, the leftmost is a general formula for an aldose and the second is for a ketose.

Aldose chemical structure


Ketose chemical structure


Monosaccharides may also be grouped according to the number of carbon atoms linked together in a chain making up their molecule. The pentoses are an important group, as ribose and deoxyribose, both pentoses, are incorporated into RNA and DNA, respectively. Similarly, hexoses are important as the group that contains glucosegalactose and fructose, all sugars important to human metabolism. Glucose and galactose are aldoses, while fructose is a ketose.

3 carbon chaintriose
4 carbon chaintetrose
5 carbon chainpentose
6 carbon chainhexose
7 carbon chainseptulose

The carbon atoms in sugars are often numbered so that it may be specified where in the molecule a reaction has occured.

Numbers 1 through 6 arranged vertically
Glucose chemical structure


Galactose chemical structure


Fructose chemical structure


Carbon number 1 is the topmost carbon in each case in the formulas above, i.e. the CHO for glucose and galactose, and the CH2OH for fructose. The other carbon atoms are then sequentially numbered downwards. The way the formulas have been illustrated above is called the Fischer projection.

A comparison of the formulas for glucose and galactose shows the difference between them to be that one of the carbons (number 4) has “HCOH” in glucose and “HOCH” in galactose. However, this difference could be at each of the four carbons (numbers 2 to 5) giving 16 possible monosaccharides, known as stereoisomers.

Allose chemical structure


Altrose Chemical Structure


Glucose chemical structure


Mannose chemical structure


Gulose chemical structure


Idose chemical structure


Galactose chemical structure


Talose chemical structure


Of these, eight will be mirror images of the other eight and are known as enantiomers of each other. They are grouped according to carbon 5. Eight of them will have “HCOH” there, and the remaining eight will have “HOCH”. Those with “HCOH” are the D form of each monosaccharide, and those with “HOCH” are the L form. Note that the “D” and “L” are in capital or uppercase letters. The eight formulas immediately above are all of the D-enantiomers, and “D-” could be put as a prefix to the names given, i.e. D-glucoseD-galactoseetc. The two formulas below show the difference between the “D” and “L” enantiomers, using glucose as an example. D-glucose is probably the most widely distributed hexose in biological systems, certainly it is the one we are most concerned with in anatomic pathology.

D-glucose chemical structure


L-glucose chemical structure


Designations as the “D” or “L” enantiomer is based on a similarity to the positions of the groups relative to each other in the structure of glyceraldehyde. In that compound, the “D” and “L” refer to the optical rotation of light in a solution, but this is not carried over to the monosaccharides and the lowercase d and l have been used to designate the optical refraction of the specific molecules. These optically active compounds may exist as mixtures of both the “D” and “L” forms, in which case they are designated “DL”. If present in equal amounts, they become optically inactive as the right and left rotations of light cancel each other out, in which case they are called racemic mixtures.

If light is passed through a solution of a monosaccharide, it will be rotated to either the right or the left. If to the right, it is designated with the plus sign + or the lowercase d. If to the left, the minus sign  or lowercase l is used. The lowercase “d” is short for “dextro”, meaning “right” and the “l” is short for “levulo” or “laevulo”, meaning “left”. The prefix “dextro” gave rise to the term “dextrose” for the most commonly encountered form of glucose, dextro-rotatory D glucose, illustrated just above. Similarly, the prefix “levulo” gave rise to the name “levulose” for fructose, or fruit sugar. Although sometimes seen, both “dextrose” and “levulose” as identifying names for glucose and fructose are deprecated and should be avoided in scientific use.

Ring Forms

Besides the Fischer projection to illustrate the structure of carbohydrates, illustrations using rings are frequently seen. Known as the Haworth projection, they are commonly encountered. There are two types of ring formula, those based on pyranose, which has a six member ring (5 carbons and an oxygen), and those based on furanose, which has a 5 member ring (4 carbons and an oxygen).

D-glucose chemical structure


D-glucopyranose chemical structure


D-glucofuranose chemical structure


As can be seen, a pyranose ring is formed from the hydroxyl of carbon 5 reacting with the aldehyde of carbon 1 forming a hemiacetal. In the case of the furanose, the reaction is between the aldehyde of carbon 1 and the hydroxyl of carbon 4. The hydroxyl group on carbon 1 of glucopyranose, called the glycosidic hydroxyl, is reactive and can form compounds with other monosaccharides and alcohols. These are called disaccharides if with another monosaccharide, or a glycoside if with a non-carbohydrate ol, methanol for instance.

alpha-D-glucopyranose chemical structure


beta-D-glucopyranose chemical structure


Two different pyranose molecules can be formed from glucose, depending on whether the hydroxyl next to the oxygen is on the opposite side of the molecule as the CH2OH, in which case it is known as α-D-glucopyranose (the formula on the left), or on the same side, when it is known as β-D-glucopyranose (the formula on the right).

Similar compounds for keto sugars are shown below, using fructose as an example.

alpha-D-fructpyranose ring chemical structure


alpha-D-fructofuranose ring chemical structure


There is an alternate way of drawing monosaccharide rings, shown in the diagram on the left, below. This is sometimes called the “chair” structure, and it is designed to show the relationships between bond angles more clearly. It is of the same α-D-glucopyranose shown in the diagram next to it. The third diagram, on the right, gives the usual numbering sequence for monosaccharide rings.

Glucose chair chemical structure

α-D-glucopyranose chair

alpha-D-glucopyranose chemical structure

α-D-glucopyranose ring

Hexose numbers ring


Keep in mind that when the term “glucose” is used in anatomic pathology, it usually refers to α-D-glucopyranose, as that is the most common form encountered.


Monosaccharide units can combine with each other to form disaccharides, “di” meaning “two”. Probably the most familiar will be sucrose, or cane sugar, and lactose, or milk sugar. Sucrose is illustrated as a product of α-D-glucose and β-D-fructose, and lactose is illustrated as a product of β-D-galactose and β-D-glucose.

Sucrose chemical structure


Lactose chemical structure



In human tissue, the polysaccharide encountered is glycogen. In vegetable tissues, it is starch and cellulose. Some animals and fungi contain chitin. All are different kinds of polysaccharides. Glycogen, starch and cellulose are polymers of glucose, while chitin is a polymer of N-acetylglucosamine, a modified glucose.

The polysaccharide that is most important in anatomic pathology is glycogen, a polysaccharide very similar to starch. It is made up of many glucose units, perhaps a quarter to half a million or more. These are joined in long chains by 1-4 glycosidic links, i.e. they are linked together at carbon atoms 1 and 4 by the glycosidic hydroxyl. Every ten or so glucose units there is a 1-6 glycosidic link which causes the glycogen chain to branch.

Glycogen chemical structure


One of the the major polysaccharides found in plants is starch, which includes two slightly different compounds. One is amylose, a non-branching polysaccharide composed of large numbers of 1-4 linked glucose units. The other is amylopectin, a branching polysaccharide similar in structure to glycogen but with fewer 1-6 linked branches and two to three times as many glucose units between branches. Amylopectin is sometimes used as a section adhesive.

Cellulose is another glucose polysaccharide found in plants. This is a very widely distributed material as it forms a major part of the structure of plants. It is also a non-branching polymer of glucose units linked via 1-4 bonds, but in this case, they are β-glycosidic bonds rather than the α-glycosidic bonds of glycogen and starch (see the diagrams above of α-D-glucopyranose and β-D-glucopyranose). Cellulose polymers contain several hundred to several thousand glucose units with hydrogen bonds between units which stabilize the structure and make it more rigid.

Chitin is a polysaccharide found in fungi, the shells of crustaceans and some other animals. It is a polymer composed of N-acetylglucosamine rather than glucose. The N-acetylglucosamine units are linked via 1-4 β-glycosidic bonds similar to cellulose.

All of these polysaccharides can be encountered in anatomic pathology, either as a natural component of the tissue, as a consequence of disease (chitin in fungal cell walls), contamination from latex gloves (starch granules) or by chance (wood splinters, food residue). Fortunately, they all look different microscopically so even though all of them may be PAS positive, their visual appearance makes them easy to identify.

The presence of N-acetylglucosamine in chitin highlights the fact that groups other than another monosaccharide can attach to a monosaccharide. A commonly encountered material composed of a glucose molecule with an amino group attached is glucosamine, a dietary supplement often taken by those with arthritis and a common component of the mucins, glycolipids and glycoproteins. If the amino group itself has an attached carboxyl group (N-acetyl-) the compound formed is N-acetylglucosamine. As noted, the polymer of this compound is the polysaccharide chitin.

D-glucosamine chemical structure


N-acetyl-glucosamine chemical structure


A common group of compounds are the sialo-mucins, widely distributed as components of epithelial mucins. Sialomucins contain sialic acids (neuraminic acids), which are nine carbon amino sugars containing carboxyl groups. Two of these, N-acetyl- and N-glycoloyl- neuraminic acids, are illustrated below and the numbering of the nine carbons is shown on the left.

Numbering for N-acetylneuraminic acid


N-acetyl neuraminic acid chemical structure

N-acetyl-neuraminic acid

N-glycolloyl neuraminic acid

N-glycoloyl-neuraminic acid

Carboxyl groups are not the only acid groups that can form compounds with monosaccharides. Compounds with phosphate groups are quite common. Glucose-1-phosphate and glucose-6-phosphate are examples. The difference between these two compounds is which carbon has the phosphate group attached. Both are important in cellular metabolism, energy storage, etc.

It is also not a case of either one group or another group attached to a monosaccharide. It can easily be a case of one group and another. A case in point is N-acetylglucosamine-6-phosphatei.e. glucose with an amino and carboxyl group attached at carbon 2, and a phosphate group at carbon 6. Compare the formula for N-acetylglucosamine and the formula for glucose-6-phosphate below.

Glucose-1-phosphate chemical structure


Glucose-6-phosphate chemical structure


N-acetylglucosamine-6-phosphate chemical structure


In addition to adding carboxyl groups to the molecule, both the terminal aldehyde of aldose sugars and the terminal hydroxyl can be oxidized to a carboxyl group. This gives three possibilities. If the terminal aldehyde alone is changed, it forms an aldonic acid. If the terminal hydroxyl is changed, it forms a uronic acid. If both are changed, it is called an aric or saccharic acid. These are illustrated below with glucose.

Gluconic acid chemical structure

Gluconic acid

Glucuronic acid chemical structure

Glucuronic acid

Glucuronic acid ring chemical structure

Glucuronic acid ring

Glucaric acid chemical structure

Glucaric acid

Glucuronic acid is a common constituent of mucins. So is N-acetylglucosamine. Together they form the disaccharide hyaluranon, also known as hyaluronic acid with the two monosaccharides being linked by bonds alternating between the 1-3 and 1-4 positions. The polymer of this compound, in conjunction with protein, forms the basis of hyaluronic acid containing acid mucopolysaccharides. Another class of acid mucopolysaccharides are the sialomucins, already mentioned.

Hyaluranon chemical structure


As well as carboxylated acid mucopolysaccharides, there are sulphated mucopolysaccharides. These are formed from monosaccharides containing sulphate groups. Chondroitin sulphates are examples, and contain sulphated galactosamines in conjunction with glucuronic acid. Chondroitin-4-sulphate and chondroitin-6-sulphate are examples differing in which carbon has the sulphate group. Chondroitin-4-sulphate is called chondroitin sulphate type A and chondroitin-6-sulphate is type C. Chondoitin sulphate is found in cartilage, and is used as a dietary supplement for people with arthritis. Dermatan sulphate is similar but contains iduronic acid instead of glucuronic acid, and is found in the skin. Heparin is also a related compound, which contains iduronic acid and is more strongly sulphated. It is an anticoagulant found in mast cells and often used for their demonstration. Like chondroitin sulphates and dermatan sulphate, heparin is strongly metachromatic with the azures and related dyes.

Chondroitin-4-sulphate chemical structure


Chondroitin-6-sulphate chemical structure


Dermatan sulphate chemical structure

Dermatan sulphate

Heparin chemical structure


Pentose sugars

So far the discussion has focused on hexoses, particularly glucose and how it is modified for different purposes. There is another group of compounds that are very important but are based on the pentose sugars, ribose and deoxyribose. Deoxyribose has one less hydroxyl group than ribose, with hydrogen replacing it. This is shown in the diagrams below. Ribose and deoxyribose are both shown as they are usually represented and next to that the same molecules are shown with the hydrogen atoms explicitly included to emphasize the oxygen that has been removed in deoxyribose at carbon 2. The numbering scheme is shown to the left.

Ribose numbering


Ribose chemical structure


Ribose explicit chemical structure


Deoxyribose chemical structure


Deoxyribose explicit chemical structure


Both ribose and deoxyribose are important as they are the basis of RNA (ribosenucleic acid) and DNA (deoxyribosenucleic acid), respectively. However, that is not their only function. They are also important in energy transfer and cellular respiration.


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