**2.3. Transformations of polysaccharides in solid state and in solutions**

Macromolecules characteristically have broadly varying molecular weights, from ~103 to ~107 Da. Macromolecules in polymers involved in technological production of various materials may experience mechanical action and partial destruction (breakdown of chains) whereby their molecular weight becomes ever more heterogeneous and diminishes on average [38]. The destructive change may be especially prominent in "dry" technological processes, such as pulverization, pelleting, or mixing, e.g., in mechanochemical solid-state complexing of drugs with water-soluble polymers (polysaccharides). Partial destruction of polymers may change their toxicological properties which have to be cautiously monitored when making new drugs and food products.

Molecular weight patterns were studied [39] in polysaccharides (dextrans 10, 40, and 70, HES 200/0.5, and larch AG and acacia FG gum) by gel permeation chromatography (GPC) [40] of samples treated in rotary and planetary mills; the obtained materials were tested for their toxicity.

580 The Complex World of Polysaccharides

complexing on mechanical activation of solids.

be ~> 100 ms, judging from the conditions of slow exchange.

to slower exchange of molecules and stronger binding.

when making new drugs and food products.

hydrogen nuclei in the sample. The mobile phase may correspond to fragments of AG macromolecules, possibly, side chains, as one may reasonably hypothesize given that water content in AG never exceeds 2 wt.%. This very fact appears to facilitate AG-Drug molecular

*AG-Drug systems* most often exhibit distinct biexponential kinetics as evidence that the drug molecules are either free or bound in complexes with AG. The bound molecules are more abundant and less mobile in milled samples, while the free ones keep almost invariable NMR relaxation times. The characteristic 1H NMR bands of clozapine and mezapam move to low field on complexing, possibly because the molecules become protonated at the account of minor remnant uronic acid present in AG, the shift being likewise greater in the milled samples. However, no complexing-related shifting appears in the cases of indomethacin and diazepam. The life time of drug molecules in complexes with AG must to

The system AG-diazepam offers an illustrative example. Solutions of these mixtures not subjected to mechanical treatment show mono-exponential relaxation behavior, but with shorter times than in free diazepam, likely as a result of rapid solution-complex molecular exchange. The milled mixtures, on the contrary, have biexponential kinetics corresponding

Thus, dynamic NMR spectroscopy of all Drug-AG solutions indicates formation of supramolecular drug-polysaccharide complexes, like the data on solubility increase. Most likely, the complexing sites are at side chain spaces in the branching macromolecules. Unlike cyclodextrins, ensembles of polysaccharide molecules (including arabainogalactan) are micro-heterogeneous in mass and structure. As a result, molecular modeling of the complex is very difficult. The binding mechanism appears to lie mainly with hydrophobic interactions [33, 34] which are typical of guest-host cyclodextrin complexes. A certain support to this hypothesis comes from stronger binding of highly lipophilic drugs which are almost insoluble in water. In this case, the branched structure of AG macromolecules [35, 36] is especially favorable for complexing. However, Coulomb interactions may contribute as

well in the presence of acid-base groups in polysaccharides and drugs [37].

**2.3. Transformations of polysaccharides in solid state and in solutions** 

Macromolecules characteristically have broadly varying molecular weights, from ~103 to ~107 Da. Macromolecules in polymers involved in technological production of various materials may experience mechanical action and partial destruction (breakdown of chains) whereby their molecular weight becomes ever more heterogeneous and diminishes on average [38]. The destructive change may be especially prominent in "dry" technological processes, such as pulverization, pelleting, or mixing, e.g., in mechanochemical solid-state complexing of drugs with water-soluble polymers (polysaccharides). Partial destruction of polymers may change their toxicological properties which have to be cautiously monitored See Fig. 2 for example chromatograms of AG and Table 3 for calculated molecular weights of the analyzed polysaccharides before and after mechanical treatment.

**Figure 2.** GPC chromatograms of 0.02 wt.% arabinogalactan water solution. 1 = native; 2 – 5 = subjected to mechanical treatment: ball mill, 2 hours (2), ball mill, 24 hours (3), planetary mill, 10 min (4), extremely intense treatment, mixed ball loading (5); Eluent: H2O/0.1 N LiNO3

Polysaccharides ground in a high-rate planetary mill diminish markedly in molecular weight and change slightly their polydispersity index Mw/Mn. The mechanical destruction is stronger in polymers with larger molecular weights, which agrees with published evidence [39]. Note that highly branching macromolecules (HES and AG) break down into roughly equal fragments. The Mw/Mn ratios in polysaccharides do not grow much, possibly, because destruction mostly affects their high-molecular fractions. Destruction is apparently controlled by the structure of polysaccharide molecules and physicochemical chain breakdown mechanisms. According to a model for linear synthetic polymers [41], the chains that occur in the middle of macromolecules are especially prone to failure. Destruction of HES and AG is qualitatively similar to that model, though dextrans and fibregum may deform by a different mechanism.

The results for larch arabinogalactan are worth of special consideration. High-rate treatment in a planetary mill, especially with mixed ball loading reduces strongly the AG molecular weight. According to chromatograms (Fig. 2), its Mw 17.3 kDa macromolecules split quantitatively into two almost equal parts of Mw = 8.3 kDa, while their Mw/Mn ratio decreases to 1.08 [23]. Therefore, the native AG molecules may consist of two relatively weakly bonded fragments of equal molecular weights and easily break down on milling [23,39]. Note that AG macromolecules with MM (Molecular Mass) ~9 kDa are likewise the main product of chemical destruction of Canadian larch AG [42].

Furthermore, the analyzed polysaccharides experience almost no mechanical failure on lowrate grinding in a rotary mill (Table 3). Thus, ball rotary milling appears to be most often preferable, as molecular mass changes in technologically produced polymers are commonly unwanted in view of their further use in dietary supplements and drugs, otherwise additional tests and standardization may be required.


**Table 3.** Molecular mass distribution of polysaccharides

Toxicological tests of the milled polysaccharides show that a single intragastric injection administered at doses from 500 to 6000 mg/kg body weight caused no death in experimental animals. Their appearance, behavior, and state were within the background over the whole dose range; no statistically significant changes in body temperature relative to the control was observed, and body weight growth was uniform in all groups. Injections of the tested polysaccharides neither induced any considerable effect on the central nervous system of the mice. Patomorphological postmortem examination of mice in 14 days after polysaccharide administration revealed no pathology in thoracic and abdominal cavities. The median lethal dose LD50 for all polysaccharides was over 6000 mg/kg body weight on single intragastric injection.
