All previous in vivo studies evaluating activity of mastic gum against H. pylori have used a crude mastic preparation which contained a high percentage (30%) of an insoluble and sticky polymer. We speculated that the presence of the polymer hindered potential in vivo activity of mastic during oral administration, and for this reason we prepared TMEWP. We have verified that the chemical consistency of TMEWP was virtually identical to that of crude mastic gum, except for the absence of the polymer, and it also presented better solubility properties and increased concentration of active constituents. Previous animal studies for the determination of the potential anti-Helicobacter activity of mastic gum were organized on a short-term administration schedule. However, we have extended our administration and hence tested the TMEWP activity over a period of 3 months.
In the present study we utilized an established H. pylori
infection model to evaluate the potential therapeutic effect of continuous TMEWP administration on H. pylori
colonization and development of associated gastritis. This model involves the mouse-adapted H. pylori
Sydney strain 1 (SS1), which colonizes the C57BL/6 mouse heavily and leads to the development of appreciable levels of gastritis closely mimicking human H. pylori
). The particular infection model has been successfully utilized in the past to evaluate potential anti-H. pylori
activity of a number of agents, such as antibiotics (14
) or lactic acid-producing bacteria (27
Our experiments showed that the mastic total extract could moderately reduce H. pylori
colonization in the antrum and corpus of the stomach. The reduction in colonization levels calculated was approximately 30-fold, in the range of 1.5 log CFU/g of tissue. These results were in concurrence with the visible reduction in H. pylori
colonization observed in the histopathology evaluations. However, such a moderate fall in H. pylori
colonizing numbers could not support any attenuation of the H. pylori
-associated chronic active gastritis. We have documented no such decline in either neutrophilic or lymphocytic infiltration within the lamina propria. Equally, no reduction was observed with reference to serum anti-H. pylori
IgG antibodies between the mastic-treated and nontreated animal groups. Collectively, these results are in line with earlier observations involving the same infection model, in which 7-day monotherapy with crude mastic tear diluted in 100% ethanol failed to eradicate H. pylori
). Nevertheless, we were able to document a moderate drop in colonization levels possibly by increasing the bioavailability of the active mastic constituents, following polymer removal. Another main advantage of our study was the application of a longer period of administration and observation. In this way we were able to administer 0.75 mg of TMEWP (the approximate equivalent of 1 mg total mastic gum) continuously through the animal water supply, for a period of 3 months, as opposed to only 1-week therapy regimens reported in other studies. Our mode of administration mimicked more closely real life conditions, where active mastic constituents can be released in a sustained release mode following daily consumption of mastic gum. Although 3 months proved too short a period for the mastic total extract to eradicate H. pylori
, results suggest that in real life, habitual daily consumption of mastic gum over a much longer period may well create conditions favoring a decrease in H. pylori
colonization levels. However, we observed no prophylactic activity of TMEWP on H. pylori
infection, although our observations depend on a small number of animals. Equally, to date no epidemiological data exist to support such a prophylactic effect among the mastic gum-consuming population of Chios Island.
A detailed in vitro investigation was performed in order to identify the potentially most active fractions in mastic extracts. The TMEWP was further separated into an acidic and a neutral fraction, and the antimicrobial activities of these fractions, as well as that of mastic total extract, were determined. We observed a moderate activity for the total mastic extract against a panel of 11 H. pylori
strains at a mean MBC of 0.256 mg/ml, which is higher than that previously reported against the H. pylori
SS1 strain for mastic gum preparations diluted in ethanol (17
). This could well be attributed to differences in methodology and the differential susceptibility of diverse H. pylori
isolates, as our in vitro data clearly demonstrated. The acidic fraction of the TMEWP was found to be the most active, with an MBC as low as 0.136 mg/ml, consistently in all strains tested. In order to identify the most active constituent among the contained triterpenic acids, the acid fraction was further submitted to several chromatographic separations that afforded the six major constituents. They were all tested against the same H. pylori
strains as mastic extracts, and the most active one was found to be isomasticadienolic acid. None of the pure compounds was more active than the whole acid fraction, suggesting that, although the anti-H. pylori
activity of mastic could be located particularly in the acidic fraction, probably the final activity is a result of synergy between all the acid constituents. Although it is the first case in which specific mastic acid triterpenic compounds have been shown to exhibit anti-H. pylori
activity, other tetracyclic acids similar to those contained in mastic have been reported to be active against Escherichia coli
). In our study, however, due to limitations in the isolation and purification steps, we were not able to test the potential antimicrobial in vivo activity of individual triterpenic compounds.
Emerging antibiotic resistance and reduced patient compliance are the main reasons for failed H. pylori eradication therapy. In addition, the considerable expense of the antibiotic regimen coadministered with a proton pump inhibitor creates the need for alternative antibiotics for combination therapy. The present study demonstrated that a mastic gum extract without the polymer constituent poly-β-myrcene was effective in reducing H. pylori colonization levels by 30-fold in infected mice over an administration period of 3 months and that the activity could be attributed to triterpenic acids within the acid fraction of mastic extracts. These results suggest that removal of the constituent poly-β-myrcene polymer can produce an enhanced therapeutic moiety which may exhibit anti-H. pylori activity, detectable over a shorter time period, bearing all the advantages of a nutritional product. We plan to evaluate further the true therapeutic potential of the acidic and neutral fractions of mastic gum in extensive animal studies following administration of the respective triterpenic constituents, alone and in combination, to assess synergic effects. The role of such mastic constituents will also be assessed with reference to potential synergy with currently used antibiotic eradication schemes.
Finally, our results also suggest that habitual long-term mastic consumption may be effective in moderating H. pylori colonization, although to date no epidemiological data exist to support the hypothesis of a reduced prevalence of H. pylori infection among the mastic gum-consuming population of Chios Island.