This newsletter looks at the nature of Aloe carbohydrates and considers the roles they play in Aloe’s biomedical activities.Aloe Vera contains special carbohydrates, some of which are a very important part of the “active principies’’ which account for some of the medical effects of this famous plant.

The Carbohydrate Fraction In Relation to Other Aloe Solids

The solution which results from extracting Aloe leaves is dilute because the plant is a succulent andstores water in its leaves. These levels of total solids have been reviewed in Newsletter No 11. AnAloe vera Gel product typically contains about 0.46% to 0.6% solids, occasionally up to 0.8% and averaging just about 0.55%. We expect an Aloe Whole Leaf Extract to average about 1.25% solids with a likely range from 0.8% to 2.0%. The question now arises as to what part of these rather low solids concentrations represent the active principles of Aloe and what part represents physiologically inactive material which merely accompanies the active principles.

In all Aloe extracts, quite ordinary and common substances with small molecules make up a very large proportion of the total solids. These mainly comprise organic acids (such as oxalic, malic, lactic and citric acids), chloride, free sugars (glucose and fructose) and metal ions, such as potassium, sodium, calcium and magnesium. These are all of them such common and well known components of biological materials generally, including foods, that we can be sure that no special physiological activities attach to them. Low levels of most of the common amino acids have also been found along with other substances with a more fatty character known as fatty acids, sterols, hormones called eicosanoids and terpenes. Together these small molecule substances make up from 75% to 90% of the total solids of the Aloe. That leaves only 25% down to 10% of the solids as larger molecule substances.

This does not mean that the small molecule fraction is entirely inactive, far from it. But it is true that the substances which make up the largest part of this fraction are inactive. The minor small molecule components, like the sterols and eicosanoids mentioned above are active in connection with inflammation. Other very minor components with biological activity are the plant hormones called gibberellins and auxins and the group of substances related to aspirin, known as salicylates. Then there are also small molecule components of the phenolic fraction (meaning the same as “Exudate” or “Aloin” fraction) present. Some of these, which include substances called anthroquinones, anthrones and chromones, are, indeed, active principles, and there is reason to think that one of them – aloesin – may be quite important as an active principle. These are due to be discussed in Newsletter 14. There is no doubt that the small molecule fraction taken as a whole possesses active anti-inflammatory and healing effects. The point is that the substances responsible for these effects are present in minute quantities. The bulk of the small molecule substances are, indeed, inactive. Obviously the tiny quantities of active principles present in this fraction of Aloe are very potent at extremely low concentrations. The components of the anti-inflammatory principle appear to be particularly prevalent in the small molecule fraction.

The number of different substances acting in concert, and the way in which they work alongside the large molecule fraction, serve to confirm the relative complexity of Aloe’s actions. This certainly gives scope to those who say that with Aloe, as with other herbs, “the whole is more than the sum of its parts”. However, the fraction of Aloe which has attracted the most systematic study for its biological activities by researchers in many different parts of the world is the large molecule fractions. As mentioned above, this fraction is much smaller in amount than the small molecule fraction, though it does clearly entrain a lot of Aloe’s biologically active principles. This fraction contains small amounts of protein and nucleic acids. The tiny protein component includes some enzymes, amongst which there is bradykininase, which is one of the anti-inflammatory substances in Aloe. But this fraction is primarily composed of complex (that is large molecule) carbohydrates. These are known as polysaccharides or complex carbohydrates. Research has consistently shown that they are keybearers of both healing and immunostimulant activity. They are large molecules built up in chains from small molecule sugars, as will be described below.

The Nature of Simple Sugars

To conceive the nature of polysaccharides it is first necessary to understand about sugars and the wayin which they become linked together. A very good attempt to explain this has been published by R.P.Pelley “Aloe Quality Control: Elementary Sugar Chemistry”, in Aloe Verities 1994. However, this is stillfairly technical and includes the use of chemical formulae, it requires one to understand electrical charges on molecules and to follow a lot of tuition and arguments about “stereochemistry”. In short, it requires considerable exercise of the grey matter on topics entirely unfamiliar to non-scientists. Here I shall try to avoid making this quite so necessary. Starting with a sugar like glucose, one needs to envision that its molecule (the smallest part of it that can exist and keep its identity) is based upon a chain of six carbon atoms liked together. Whilst this type of molecule can exist as a straight chain of carbon atoms, it has a marked propensity to curl itself round and form a ring. Most of the sugar molecules will be in the ring form. Attached to each carbon atom in the chain are hydrogen atoms and oxygen atoms arranged in a particular pattern. Such a molecule is quite small (it definitely falls into the small molecule fraction of Aloe). It is an example of a “simple sugar”. That means that each of its molecules comprises a single six- carbon unit by itself. The way in which a simple sugar like this is converted into more complex molecules is by linking the rings together in chains. Simple sugars most commonly have either six carbons in the molecule as described above, (a class of sugars called hexoses) or five carbons (a group of sugars called pentoses). In some situations you get 4-carbon sugars or 7-carbon sugars. However, in looking at the active principles of Aloe we need only consider 6- carbon sugars. These belong to the group called “monosaccharides”, which refers to the fact that they comprise only one sugar unit per molecule.

The 6-carbon sugars which commonly occur in foods are glucose, fructose, galactose and mannose. Since they all have the same length of carbon chain and also all join up head-to- tail to form rings, the only differences between them are in respect of the ways in which the oxygen and hydrogen atoms are arranged around the carbon chain. Fucose and rhamnose are also 6-carbon sugars but they differ in having slightly less oxygen in the molecule. They are also less common and less abundant. These sugars are all of them small molecules, indeed, their molecules are all the same size. Their combinations of oxygen and hydrogen atoms make them very readily soluble in water and small molecular size makes them very diffusible. When their solution is enclosed in what is called a “semipermeable membrane” (a membrane which allows small molecules to pass through but acts as a barrier to large molecules), these simple sugars pass through but more complex carbohydrates are retained. This is the basis of the separation technique known as dialysis, which may be used in practice to separate these fractions.

These simple sugars, or monosaccharides, are plentiful in Aloe extracts. Pelley and Wang of the university of Texas Medical Branch reported that 0.28% by weight was present in Aloe gel extracts having 0.6% solids. In other words just under half of the solids were accounted for as simple sugars. Of these, 95% was glucose and 5% fructose, other types contributing, apparently, only trace amounts. Whilst these will, of course, be used as an energy source when ingested by humans, they have no other physiological action.

The Nature of Disaccharides and Oligosaccharides

As indicated above, the molecules of simple sugars (in their ring forms) can become linked together inchains. The link between one sugar unit and the next is effected by a single chemical bond. The simplesttype of molecule that results from this process contains only two sugar units linked in this way. Such a substance is no longer just a simple sugar. It is called a disaccharide. Sucrose, or common sugar, derived from sugar beet or sugar cane, is a disaccharide in which the two sugars present are glucose and fructose. Others are milk sugar, or lactose, containing glucose and galactose, and maltose, which comes from breakdown of starch, containing two molecules of glucose. If three sugars link together this forms a trisaccharide. Larger chains in which the number of sugar units are still small (one might say “a few”, or “several” are called oligosaccharides. All of these are still reckoned as small molecules and will pass through a semipermeable membrane. Oligosaccharides can have physiological effects, but no such effects are known in Aloe.

The Nature of Polysaccharides

As the chains of sugar units which make up oligosaccharide are lengthened much further, they eventually build up to molecular sizes which have to be classified as large and these will now be retained behind a semipermeable membrane. The best known example of this is starch, which is composed of nothing but glucose units linked together. An important component of starch, amylose, usually contains up to 300 glucose units per molecule. The molecular weight (the usual chemist’s measure of the size of an organic molecule) is up to about 50,000 and this substance will be easily retained behind a semipermeable membrane. Starch is a near ubiquitous carbohydrate energy store in plants. The chains of sugar units
may be either a continuous single chain or can be branched. Wherever a polysaccharide is composed wholly of glucose units it is referred to as a glucan. Those containing only galactose units are galactans, those containing only fructose units are fructans and those containing only mannose units are mannans. Those containing two kinds of sugar units in combination are named according to the particular sugars they contain. Hence, there are, for example, glucomannans and galactomannans.

Pentose sugars also form polysaccharides called, for example, arabans or xylans, but these need not concern us further here. Still other types of polysaccharide involve, not simple sugar units, but derivatives of simple sugars (i.e. the sugar molecule has been modified). The oxidation of a simple sugar to an acid result in a hexuronic acid, the one derived from glucose being glucuronic acid and that derived from galactose being called galacturonic acid. Or the sugar may be altered by the addition of a nitrogen-containing group called and amino group. This leads to a hexosamine, the one from glucose being called glucosamine. Polymers (polysaccharide-like materials) made up from these amino sugars are called glycosaminoglycans. Polysaccharides of this type are not common in plant products.
Some plant polysaccharide exist in a form which chemists call “acetylated”. This means that some of the oxygen molecules which form part of the sugar units are combined with a chemical grouping derived from acetylation may also help to identify the species from which the polysaccharide is derived.

The Nature of Aloe Polysaccharides I

Polysaccharides other than Glucomannan

The polysaccharide of Aloe which primarily bears physiological activities has for a good many years been recognised as the glucomannan. We shall look more closely at this material. It is interesting to consider first what other polysaccharides occur in Aloe. Types of polysaccharide other than glucomannan that have been found in Aloe vera include some which contain “uronic acids” (a term which is most likely to include glucuronic acid and/or galacturonic acid (see above). Specifically, Farkas (1963) found 2.4% of “uronic acids” and work in West Bengal by Mendal and Das (1980) found much galactose polymer (galactan) and also galacturonic acid.

This work stands out for not according pride of place to glucomannan in Aloe vera. However, results are different according to whether Gel or Whole Leaf Aloe material is studied. Work by Waller et al (1978) found “trace amounts of rhamnose, xylose, galactose and either arabinose or fucose”. Work by Segal et al (1968) also reported the presence of small amounts of arabinose, galactose and xylose. It seems clear then, that at least minor amounts of these other sugars are to be found in Aloe vera.

The question arises about what the differences might be between the polysaccharide components of the Gel and the Rind of Aloe vera. Any such differences will naturally tend to be reflected in the composition of Aloe vera Gel extracts versus Whole Leaf Aloe vera extracts and may well be reflected in differences of biological activity. Overall the literature and personal communication with researchers in the field suggest that glucomannan is, indeed, the principle polysaccharide of the Gel and that whilst the Gel probably also contains some polysaccharide material with arabinose, galactose, xylose, fucose and rhamnose components, the quantities of these are rather insignificant. It also appears clear that galactans and galacturonic acid polymers do occur in Aloe vera and that they are principally located in the outer rind of the leaf. These can therefore be expected to be significant in Whole Leaf Aloe vera but not in the Gel. In work published by Schmidt & Cutler (1994) the polysaccharides of Whole Leaf Aloe were found to be 27- 30% galactan and 70% glucomannan. As to whether the galactose-containing polysaccharides in any way contribute to the biological activity of Whole Leaf Aloe seems to be unknown at the present time.

The Nature of Aloe Polysaccharides II

Glucomannan

Ever since the work of Roboz & Haagen-Smit (1948) it has been recognised that the polysaccharide of Aloe Gel (the main Aloe product used in the past) was primarily composed of Glucomannan. However, there has not always been perfect agreement about the ratio of glucose to mannose in this material. Roboz & Haagen-Smit found that ratio to be about 1:1. Waller et al found this ratio to be about 4:5. However, later investigations have made it very clear that the overall balance of the composition of glucomannan is very much more in favour of mannose. In a very well- respected study in 1979, Gowda et al. showed that the ratio of glucose to mannose overall was 1:6, but actually they found four different fractions of polysaccharides, all glucomannans. Individually their ratio of glucose to mannose varied from 1.5:1 to 1:19. Some of the earlier analytical difficulties may be accounted for by analysing only part of the full complement of glucomannan in a sample. Gowda and colleagues showed that the glucomannans were linear, that is, their sugar units are arranged in unbranched chains. They also showed that the links between the sugar units were of a special kind which
chemists call 1:4β. Also, most authorities agree that Aloe glucomannan is, to some extent, acetylated These, then, are the main characteristics of Aloe glucomannan.

Subsequent work has made this component of Aloe vera a very familiar substance and has clearly indicated that the immunostimulant and cell-proliferative activities of Aloe reside, at least in part, in this component. Glucomannan has, indeed, been isolated and marketed as a product in its own right, separately from the other components of Aloe. Further on in this Newsletter we shall be looking more closely at its biological activities.

The Nature of Aloe Polysaccharides III Other Aloe Species

Aloe saponaria

Early work of Yagi, (1984) on the structure of the polysaccharides in Aloe saponaria (Hill) Haw. found two principal fractions, one of which was purely mannan (Mannan 1) and the other a glucomannan (Mannan 2) with a heavy preponderance of mannose. Mannan 1 inhibited induced oedema and was therefore anti-inflammatory. This species has also been shown to contain a glycoprotein (a protein with carbohydrate attached to it) with immune-active properties.

Aloe arborescens

Ovodova et al (1975) isolated a polysaccharide from Aloe arborescens comprising galacturonio acid. Yagi, (1977) studied Aloe Mannan Polysccharide, from Aloe arborescens Mill, var natalensis Berger. They found a main polysaccharide (aloe mannan) of Aloe arborescens var natalensis, obtained it pure and proved it to be a partially acetylated p mannan, apparently free from glucose. The molecular weight of aloe mannan was calculated to be approximately 15,000 (about 80 sugar units). An inhibiting effect of aloe mannan was found against an implanted tumour called sarcoma-180.

Aloe vahombe

Radjabi et al (1983) studied the Glucomannan from this species and found it to be a single type of unbranched acetylated glucomannan with a glucose / mannose ratio of 1:3 and that the linkage between the sugar units was of the 1:4 type as in Aloe vera.

Aloe plicatilis

Structural Studies were done on Polysaccharide from Aloe plicatilis Miller by Paulson et al. (1978). It contained a single type of unbranched acetylated glucomannan.

The Biomedical Actions of Separated Aloe Polysaccharides

The biomedical actions of Aloe that are attributable to its carbohydrate moiety have been described already under the appropriate Newsletter headings such as Immunostimulant (Issue 1), Healing (Issue 4) and Infections (Issue 9). However, with the exception of a few of the authors quoted, reference was usually being made there to the carbohydrate of Aloe acting within a milieu of Aloe itself in which all the different active principles could act together. Here, at this point in the series of Newsletters, the aim will be to make it clear that Aloe glucomannan, after separation and purification, free from other active principles, retains these immunostimulant and cell-proliferative activities and, apparently, also retains some of the anti-inflammatory activity. In this form it is obviously able to be rigorously dose-controlled and standardized and is suitable for injection therapy and for intensive topical application. For people who want to use Aloe for internal conditions, there is, of course, absolutely no need to consume the glucomannan in purified form. Aloe is best known and best liked as a drink of Whole Aloe, either Gel or Whole Leaf Extract. But it is nonetheless interesting to know that the purified glucomannan possesses impressive activity.

In all, the Aloe vera Information Service has listings of no less than 154 literature references relating to the use of purified Aloe glucomannan. Some of these have been mentioned already: for example, the paper by Sheets et al. (1991) using A. vera, reporting major benefit to cats infected with feline leukaemia virus, discussed in Issue No 9, and the paper by Imanishi et al. (1981), using A. arborescens, reporting inhibition of growth of a mouse fibrosarcoma and discussed in Issue 6.Additions to these examples are needed here only to emphasise that the work has been carried quite a long way and has established the bioactive role of glucomannan from Aloe vera well beyond any reasonable doubt.

A PhD dissertation presented by Chinnah (1990) is entitled “Evaluation of the Antiviral, Adjuvant and Immunomodulatory Effects of a p-(1,4)-linked polymannose (Acemannan).”The study was concerned with Newcastle disease virus in chicks and concluded that the Acemannan was an effective anti-viral substance and was also an immunostimulant.
A paper by Egger et al. (1995) showed that Aloe vera (3-(1,4)-linked acetylated mannan acted as a stimulant of cell replication in animals whose bone marrow had been suppressed by irradiation. Harris et al. (1991) contributed a paper on “The Efficacy of Acemannan in treatment of canine and feline spontaneous neoplasms” showing significant anti-tumour action, believed to be mediated through stimulation of macrophage action and the release of tumour necrosis factor. King et al. (1995) reported similar effects with dogs and cats with spontaneous fibrosarcomas. McDaniel and McAnalley (1992) reviewed ‘The Antiviral and Anti-Tumour Immunostimulant Isolated from Aloe Barbadensis Miller” and concluded that “Despite initial scepticism, evidence that the aloe plant contains an active medicinal molecule and that moiety is the complex polysaccharide acemannan is irrefutable” and go on the refer to some 100 scientific papers which support that assertion.

Separating Aloe Glucomannan into Fractions and Ascribing them Different Functions

So far the Aloe glucomannan has been discussed mainly as though it were a single homogeneous material. Nonetheless, the work of Gowda et al., quoted above, showed that the ratio of glucose to mannose was overall 1:6, but four different fractions of gluco- mannans were present with individual ratios of glucose to mannose varying from 1.5:1 to 1:19. This showed that the glucomannan was what chemists call “polydisperse”, meaning that whilst the whole of it was clearly glucomannan, there were differing varieties within it.

The clear demonstration that several important biomedical functions of Aloe reside within the
glucomannan has motivated American researchers to isolate the glucomannan, separately from other Aloe components, and to market it as a cold-processed freeze-dried powder, which can be either administered orally or by injection. It has also motivated several American researchers to actually fractionate the glucomannan fraction. That is, to separate out the various sub-fractions of the glucomannan which the work of Gowda et al had shown to be
possible. This is generally being done on the basis of molecular weight. The polydispersity of the glucomannan is reflected in its molecular weight. Dr. Ivan Danhof of the North Texas Research Laboratory, in public statements, as well as in personal communication to this author, has indicated that the molecular weights of components of this material range from just a few hundreds up to a maximum of about 4,000,000. The further detailed fractionation of glucomannan on a molecular weight / molecular size basis continues to be a matter for research. Dr Danhof has stated that fractions with molecular weights of 12,000 and 57,000 are responsible for the anti-diabetic effects, i.e. these fractions exert a hypoglycaemic effect which is independent of insulin. He states, therefore, that they exert a beneficial effect upon both Type I and Type II diabetes. Further, he connects the immunostimulant effects primarily with the very high molecular weight fractions between 1,000,000 and 4,000,000. This work leads one to envision a future for Aloe in which very sophisticated medical products are generated, targeting specific biomedical activities, and therefore, specific ailments, with carefully graded glucomannan subfractions with selected molecular weight distribution. Dr Danhof does not appear to have stated what particular biomedical activities might be targeted using glucomannan fractions with molecular weights between 57,000 and 1,000,000. It is not clear either whether any of the subfractions being isolated in the USA have markedly different glucose / mannose ratios or whether any of them actually correspond with the subfractions isolated and studied by Gowda. The larger US Aloe companies tend to be involved in this work and the results are closely guarded at the present time by secrecy or by patents.
The work of Dr Danhof is specifically orientated towards the complex carbohydrates of Aloe and to isolating and using them separately from the small molecule fraction. He has stated that in his work as a gastroenterologist he has had success in relieving Crohn’s disease and ulcerative colitis through using purified Aloe polysaccharides by oral administration, although drinking the juice, for these serious conditions, was ineffective. Presumably, this is a question of the amount of the Aloe polysaccharides that can be consumed by the two methods. It is an interesting slant upon the treatment of those particular diseases and extends the statements made on the gastrointestinal system in Issue 3. Reich et al. (1994) in preliminary studies with oral Aloe glucomannan, administered to patients with ulcerative colitis, similarly concluded that the Disease Activity Index and the Signs and Symptoms Index were improved by the
treatment. It is stressed once again that most Aloe using members of the public and most Alternative Medicine Practitioners will probably always wish to use whole Aloe. What the above discussion illustrates for them is that the Aloe glucomannan is rather sensitive material. Any degradation of the glucomannan in a way that reduces its average molecular weight, or which alters in any way the actual molecular weight of specific fractions within the glucomannan, will tend to have negative results as far as biomedical activity is concerned. Such degradation can occur in processing, as stated in Newsletter No 8, through the use of heat, exposure to high salt concentrations, or by enzyme action with either natural Aloe cellulases or added cellulases. The loss of activity will not be “across-the-board and it may be confined to just those activities which are associated with the degraded glucomannan fractions. In this situation, the immunostimulant activity needs to be especially monitored.

Testing for the Presence of “Proper” Aloe Carbohydrates

It is a natural corollary to the above paragraphs to ask “how can we be sure that whole Aloe products, either whole Aloe Gel or Aloe Whole Leaf Extract, contain the correct amounts and types of biomedically active Aloe carbohydrates? It is a crucial question since, referring again to Issues Nos 8 and 11, this Is a matter which impinges directly upon the consumers ability to get value for money. The first question – that of getting enough Aloe polysaccharides – relates once again to performing the “Alcohol Precipitable Hexose Test” as described in Issue 11. The reader is reminded that the Alcohol Precipitable Solids Test is not meaningful or acceptable in this context. Having sufficient levels of Alcohol Precipitable Hexose, however, measured against a standard, is an indication that a product contains a good level of Aloe polysaccharide. Aloe Gel and Whole Leaf Extract will have to be measured against rather different standards. Standard values for Alcohol Precipitable Hexose (poly-saccharides) quoted by Schmidt and Cutler were 6.22% of total solids for Aloe Gel and
0.99% for Whole Leaf Extract. This major difference arises partly because the total solids in
Aloe Gel are very low, so the Aloe polysaccharides make up a large proportion of them.
The second question – that of checking whether the Aloe polysaccharides present show an
optimum pattern of distribution between the various subfractions of glucomannan – can only be answered by a sophisticated analytical procedure which subjects the separated glucomannan to molecular sizing. The chances are that few commercial products will perform well on that test at the present time. Few commercial products are genuinely cold- processed from beginning to end of the process. Few products, apart from 1:1 products (i.e. not concentrated above natural strength) avoid exposure to elevated concentrations of salts at the elevated temperatures. Few, regardless of whether they are Gel or Whole Leaf, avoid the use of enzymic treatment.Moreover, the complete omission of enzyme treatment is usually incompatible with optimum organoleptic properties. Consumers begin to complain about the taste and the mouth-feel of the products. There is a situation here where, perhaps, a special product for minority users is called for which would retain the maximum of biomedical power regardless of the unavoidable impact upon taste and acceptance.

Using Carbohydrate Patterns as a Check for Validation of Aloe

This last point really bears most directly upon the topic of Issue 11 on Quality Control. The
particular patterns of distributions of Aloe glucomannan subfractions is very specifically
characteristic of Aloe.The appearance of peaks of material, during molecular sizing
chromatography, having particular peak sizes at particular molecular weights gives a pattern
that could never be mistaken for anything other than Aloe. These special subfractions of
glucomannan can therefore act as marker substances for Aloe. Therefore, detection of their
presence can be regarded as, perhaps, the ultimate test to determine whether a substance
offered on the market is really Aloe or not. The test measurements, being sophisticated,
would be relatively expensive. Producing fake products that would pass these tests would not
be totally impossible either, but such fakes would themselves be sophisticated and expensive,
so the cost-effectiveness of overcoming regulation in that way would be diminishing.