I

Intermediate

Processing Untreated Decolorize Untreated Decolorize ^rea^ Decolorize Untreated Decolorize Untreated Decolorize Untreated Decolorize

Final

Lyophilize Lyophilize Lyophilize Lyophilize Lyophilize Lyophilize Lyophilize Lyophilize Lyophilize Lyophilize Lyophilize Lyophilize

Processing CHS Protection Mean% +SEM Anti-Inflammatory Activity

LoMW Hexose % of LoMW Solids

Untreated

Untreated

Figure 8.11 Effect of processing upon biological activity and some key chemical parameters. © 2004 by CRC Press LLC

Protection of the skin immune system varied in a complex fashion. Clean, gently handled leaves stored for 24 hours before filleting caused a borderline decrease in protective activity (42% protection versus 39% protection ±3%). Similarly, this meticulously filleted gel, with a very low anthraquinone content demonstrated only a 3% change in activity after activated charcoal treatment. On the other hand, treatment with cellulase increased protective activity from 37.5% to 43.5%, an increase of 15% where the range of variation was about 1.5% and this lead to further study. It was found that activation of native glucomannan gel by cellulase was followed by subsequent decay of the biological activity upon excessive treatment. There is a similar gross effect with WLE where cellulase action causes an increase of 50 to 57% and 33 to 38% of activity. As stated above, charcoal treatment has little effect on highest quality gel, although treatment of WLE reduces the CHS protective activity from 50—57% to 33—38%, a decrease of almost a half. These data suggest that adsorption removes from WLE one major factor responsible for protection of the skin immune system, a factor that is absent from gel with low anthraquinone content.

Normal processing steps, such as treatment with cellulase or activated charcoal also affect such biological activities as the modulation of acute inflammation. However, the A. barbadensis anti-inflammatories are affected by processing in different ways than protection of the CHS response. Thus, treatment of WLE with cellulase strongly activates an anti-inflammatory principle, while absorbing with activated charcoal has no effect. The situation is less clear cut for the anti-inflammatory activity in gel. Native gel, not treated by cellulase or charcoal has good anti-inflammatory activity (13 to 37% inhibition of croton oil-induced swelling), which is lost by adsorption on charcoal (-1.6 to 2.1% inhibition). Thus processing has a profound effect on the biological activities of aloe gel although there is no simple pattern of activity change among different biological activities. Lastly it should be realized that the skin immune system protective activity in Figure 8.11 is based on 14—15 animals per group whereas the anti-inflammatory data is based on only five animals per group.

Chemical parameters change with proper processing. Most notable are the changes that occur with carbohydrates, other than monosaccharides, during processing. Figure 8.11 shows around 11% of total solids as monosaccharides in treated and untreated gels. On the other hand, polysaccharides, which made up on the average 18% of solids, varied between samples. The highest values (27.2% of solids) were found with WLE, which was exposed to neither cellulase nor activated charcoal. Treatment with activated charcoal reduced this to 23.9%. Depulping treatment of WLE with cellulase, even under conditions far milder than usual in the industry, destroyed all of the acetylated gluco-mannan leaving only the pectins and galactan. Similar effects on polysaccharide were observed with gel. In general, adsorption on activated charcoal caused a drop from about 22% of solids to 17%. Treatment with cellulase under extremely mild conditions reduced gel polysaccharide from about 20.6 to 19% of solids. Again, it should be noted that the time and temperature of cellulase treatment in these experiments are far shorter and lower than is usual in industry. Cellulase treatment increases low molecular weight (<1.6 kd.) saccharides (Figure 8.11, last line), of which 14% are monosaccharide as discussed above. This means that about 2% of untreated WLE and 7—10% of untreated gel can be defined as 'oligosaccharide.' These values increase considerably when WLE is treated with cellulase but not when gel is subjected to much milder cellulase treatment.

It can be seen that processing has a remarkable effect upon all aspects of A. barbadensis gel chemistry and biological activity. The activated charcoal adsorption process, used to prevent color change, caused a vast complex of compounds to be adsorbed. Large quantities of cellulase are routinely used for viscosity reduction in WLE process and before evaporative concentration of gel in thin films. Profound biological and chemical changes occur with both the charcoal and cellulase processes. Cleavage and destruction of the cytoprotective oligosaccharide from the native gel is described elsewhere (Strickland and Pelley, Chapter 12). All of these observations are made with considerably cleaner materials, under conditions considerably more mild than those generally employed in industry. Materials processed in industry are often devoid of biological activity because of bacterial proliferation, over pasteurization, excessive use of activated charcoal and addition of large amounts of cellulase followed by prolonged incubation at elevated temperatures.

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