1st International Symposium in O3 Ozone Applications
O3 Ozone in Biology
October 31st to November 3rd 1990
________________________________________ |
Effect of autohemotherapy with 03 ozone upon the levels of hemolysis.
With the aim to evaluate the possible hemolysis during the ozone therapy by autohaemotherapy, the liberation of hemoglobin was measured and the quantity of Heinz bodies formed after submitting stress.
Fourteen healthy donors were utilized to whose blood was administered the mixture of ozone/oxygen in a volume relation of ozone:blood from 0.5 up to 1.5 and ozone concentrations from 30 up to 70 mg/L.
To evaluate the defensive capacity of erythrocytes after being submitted to ozone treatment, we made a modification in the method of formation of Heinz bodies. We found significant difference only in a volume relation ozone:blood of 1.5 and with a gas concentration of 70 mg/L. At gas concentrations of 30 and 50 mg/L the formation of Heinz bodies was similar to the control samples.
These data suggest that the therapeutical dosages of ozone/oxygen mixture and the relation of volumes applied to the blood have no effect upon the normal levels of hemolysis.
This can be related to the fact that, in these conditions, the erythrocytes keep their defensive capacity against the oxidizing agents.
________________________________________ |
Teratogenic study of O3 ozonized oil.
The effects of ozonized oil upon pregnancy and the fetal development of rats were studied. The animals were treated by gastric intubation, from the sixth day to the fifteenth of pregnancy with dosages of 2.2 mg/d and 4.4 mg/d of ozonized oil, with an peroxides index of 500.
No significant differences in the weight average and fetal sizes were observed. The dosages did not cause maternal toxicity, nor loss of weight, abortions or deaths.
No significant differences were observed between the groups, which received ozone and the control as to the number of corpora lutea, death and alive fetus, resorptions and losses before and after implantation.
In the evaluation of visceral, skeleton and external fetal anomalies, 227 controls and 266 treated fetus were studied. Three deformed fetus were observed in a mother, in the group treated with the higher dose of ozone (15 times higher than the therapeutic one) and in the control group, 2 deformed fetus belonging to the same mother.
We concluded that in the study with ozonized oil, no teratogenic nor another adverse reproductive effect was observed.
________________________________________ |
Evaluation of O3 ozone genotoxicity by citogenetic tests.
The genotoxicity of ozone was evaluate, agent used in the treatment of diverse diseases which plays an important role in the therapy of Retinitis Pigmentosa and about which radiomimetic actions have been reported when the molecule of ADN reacts directly with it.
For this study, a group of patients with different diagnostics was studied, who were submitted to citogenetic tests of genotoxiclty. The analysis of chromosomic aberrations (CA) sister chromatids exchange (SCE) and micronucleus (MN) were done at the same time, in the same patients. They received ozone by autohemotherapy during 15 sessions, with a dose of 5 mg each time.
The mean values of the main types of CAs and gaps in the pre-treatment study were for: chromosomic aberrations 0.001, chromatids aberrations 0.01 and gaps 0.01 and in the post-treatment the means were of 0.007; 0.007 and 0.009 for chromosomic and chromatidics breaks and gaps respectively.
In the SCE test, the pre-treatment mean value was 6.91 per patient, while in post-treatment was of 6.33 per patient. The results of the MN test were: 0.0085 in the pre-treatment and 0.0103 in the post-treatment. No remarkable differences were observed in any of the 3 tests, between the results before and after ozone therapy.
We concluded that the ozone treatment by autohemotherapy in the established dosage (5 mg/L) has no genotoxical effect proved in a chromosomal level.
________________________________________ |
Citogenetic effects of medical treatment with O3 ozone.
The oxidating, disinfecting and deodorant effects of ozone, has determined the great interest in the development of preclinical investigations, which guarantee its further practical application.
Among these, its use in medicine as virucide and bactericide, as well as circulatory stimulator. Due to its properties as oxidizing agent and its mutagenic effect in bacterias and yeast it proves of great interest to know its genetic effect in humans. In this work we carried out a study about the citogenetic effect of ozone by means of micronucleus test and sister chromatids exchange in lymphocytes of peripheric blood, before and after the treatment with ozone. The results obtained are discussed.
________________________________________ |
Effect of O3 ozone applied by rectal insufflation in rats.
Forty rats of the SHR strain which develop a primary hypertension were studied and applied ozone and oxygen for 15 days by rectal insufflation. They were divided into 4 groups of 10:
Group 1 was applied 4 mL of ozone (equivalent to the highest therapeutical dose in human beings).
Group 2, 8 mL of ozone; group 3, 8 mL of oxygen and group 4, negative control (without treatment).
The rats were sacrificed the 16th day and samples of the rectum, colon and small intestine were taken. These were fixed in formaldehyde (10 %) and were processed for optical microscopy. The cuts were colored with hematoxyne and eosine. Any changes in the tissue structure were observed in the microscope.
It is conclude that ozone by this way of administration and in these dosages does not produce alterations in rats.
________________________________________ |
Histological behavior of duodenal and gastro-intestinal mucous and liver in gerbils treated with O3 ozone, infected and not infected by Giardia lamblia.
Evaluating the efficacy of ozone upon diseases which affect the digestive tract such as peptic and gastroduodenal ulcers, idiopathic ulcerative colitis, Giardia lamblia, bacterial affections, etc., it proved to be of great interest to assess the effect of ozone in different forms, upon such mucous with the aim to use it as therapy. Gerbils (meriones unguiculatus) infected with Giardia lamblia were used along with another non infected group.
This treatment was carried out with ozonized water and oil, using concentrations higher and lower than therapeutic dosage. In the case of oil, it was administered during 8 days, sacrificing the animals after 15 days. In the case of ozonized water, treatment extended for 30 days and the same period after it, animals were sacrificed. Studies by optical and electronic microscopy were performed.
There were alterations only in animals receiving high overdoses and no significant differences were found between other doses, including therapeutic and non treated controls.
________________________________________ |
Effect of the intreperitoneal O3 ozone administration upon the aseptic inflammation.
Sprague Dawley rats received ozone by intraperitoneal injections with a dose of 0.12 mg/kg during 15 days. At the end of the treatment, to every rat were implanted 2 glass sheets (cover-objects). In aseptic conditions, which were in cavity during 1; 3 and 5 day according to groups of evolution.
At the end of every period, those sheets were extracted and the inflammation cells were fixed and colored. Other similar groups received intraperitoneal injections of colloidal carbon solution 2 hours before extracting the sheets, which allowed the evaluation of phagocytosis in such cells.
In the group treated with ozone, the results indicated a smaller development in the inflammatory cytology, as well as in the phagocytical capacity of cells from 1 and 3 day of inflammation, what seems to compensate towards the fifth day of evolution.
________________________________________ |
Evaluation of the renal function in rats treated with O3 ozone by rectal insufflation.
Sprague Dawley rats, 250 ±10 g weight, were treated with ozone, dally, by rectal insufflation with dose of 0.03 mg, during periods of 15; 30 and 45 d in different groups. Others received a similar treatment with a dose of 0.33 mg. All groups had parallel controls, which received oxygen
In similar doses and frequency. At the end, the animals were studied as to: arterial systolic pressure, effective plasmatic renal flux, glomerular filtration intensity, p-aminehypurate maximum, as well as, sodium, potassium and creatinine plasmatic clearings. This was performed with the purpose to assess the influence ozone could have upon the renal function. If applied in the aforementioned form.
The analysis of results did not show any difference between experimental and control animals for any of the groups studied. It was concluded there was no alteration of the renal function under experimental conditions.
On the basis of these results and the high doses used, there is a large margin of safety for the use of this agent on human beings according to its medical usefulness.
________________________________________ |
Study of renal function in animals treated with O3 ozone intraperitoneally.
The renal functions of rats Sprague Dawley, of roughly 250 g of weight, were studied after having been submitted in different groups, to intraperitoneal administration of ozone with doses of 0.03 and 0.33 mg during 15; 30 and 45 d.
The study was carried out similarly in the parallel controls, which received in the sane way oxygen. The assessed variants were: arterial pressure, effective plasmatic renal flow, glomerular filtration intensity, p-aminehypurate maximum and sodium, potassium and creatinine plasmatic clearings.
The results showed there were no significant differences for any of the studied variants among animals treated with ozone and controls. It was concluded that intraperitoneally ozone administration in doses mentioned, show a high safety margin for its application in humans, due to its medical use.
________________________________________ |
Acute and chronic toxicological study of O3 ozone applied intraperitoneally in different species.
ith the aim to incorporate ozone therapy in the therapeutic model for the treatment of reservoirs of the virus which produces hepatitis B (HVB) and taking into account the advantages of the use of intraperitoneal (IP) way, ozone was applied to Wistar rats and rams with a dose of up to ten times the highest equivalent used on human beings, for acute studies and the same dose in the same way, daily during 1 month, for chronic studies.
The clinical evaluation for both groups of animals, as well as the study by means of optical microscopy of the liver and lungs of the rats were performed.
No undesirable effects were detected in these animals due to ozone application. The microscopic morphological study did not evidence any alteration.
The toxic effect of ozone, from the clinical and microscopic standpoint was corroborated when it is applied even in small doses through respiratory tract.
Taking into account these results, it was concluded that ozone, applied trough IP is innocuous for the studied species. Its use within the therapeutic model proposed by our group still ought to be proved.
________________________________________ |
Structural and ultra-structural study of organs of rats submitted to the intraperitoneal application of O3 ozone.
Sprague Dawley rats of roughly 250 g of weight were submitted to daily application of ozone with doses of 0.03 mg during different periods of 15; 30 and 45 d. Other groups received the gas in dally doses of 0.33 mg in similar periods. All the groups had parallel control groups, which received oxygen in similar dose and administration.
After the sudden death by heart hemorrhage, the kidneys, liver, heart, lungs and spleen were extracted, fixed and processed for their study with light microscopy. Fragments of these organs were processed by standard techniques of electronic microscopy.
The results showed that in different organs, no obvious lesions occurred in their structure, or ultra-structure, which could be ascribed to the ozone action.
________________________________________ |
Structural and ultra-structural study of organs of rats submitted to O3 ozone therapy by rectal insufflation.
Sprague Dawley rats of roughly 250 g of weight were submitted to dally application of ozone with doses of 0.03 and 0.33 mg during periods of 15; 30 and 45 d. All the groups had parallel control groups, which received oxygen in similar doses and administration.
After the sudden death caused by heart hemorrhage, the kidneys, liver, heart, lungs and spleen were taken out and later fixed and processed for their study by optical microscopy. Besides, fragments of such organs were processed by electronic microscopy.
In the different organs studied, no harm in their structure or ultra-structure that could be ascribed to the action of ozone was observed.
________________________________________ |
Evaluation of the influence of O3 ozone in the superoxide dismutase in renal tissue of rats.
The superoxide dismutase is distributed, almost totally, in the cells where oxygen plays an important metabolic role, exerting upon them a protecting action against the superoxide radicals.
It is logical to think that there are differences given by intrinsical properties of organs and cells by themselves and at the same time, by those factors which exert influence in their biosynthesis, as for instance, increase in the partial pressure of oxygen, flow of radicals or liability to oxidants.
The influence of ozone application in renal tissue in rats of 250 g of average weight is assessed, applying different doses of gas (0.33 and 0.03 mg/day) in different periods (15, 30 and 45 d) and through two different ways: intraperltoneal and rectal.
The results obtained are analyzed and discussed, considering among others possibilities of applications in medical practice. Its use in renal transplants, due to the outstanding role of incidence of free radicals in reperfusion of organs.
________________________________________ |
Protective effect of O3 ozone upon vascular endothelium of rats.
The contradictory statements related to the action of ozone at a cell level, motivated the assessment of its action upon the integrity of vascular endothelium. The effect of the mixture O3/O2 at doses of 0.25 and 1 mg/kg through rectum was studied in two models of induction of vascular damage: by trisodic citrate and by arterial hypertension in rats.
The results obtained showed that ozone exerts a protective effect upon the vascular endothelium of rats in the dose and in the way of administration applied.
________________________________________ |
Preoperative treatment with O3 ozonized autohemotransfunsions in horses under general anaesthesia by halotane.
Scrollavezza G., Botti A., Zanichelli F., Poldi R. and Pezzoli G.
Veterinary Science Surgical Clinics Institute, Parna University, Italy.
A group of 10 horses to be operated were pretreated with ozone, by ozonized big auto-hemotransfusion (15 mg). The results were compared with a control group of other 10 operated horses.
The most evident result was the difference in same hematochemical parameters. Particularly, it was observed how ozone treatment prevents the decrease in intraoperation PO2 values.
________________________________________ |
Positive effect of O3 ozonation in cardiac activity restoration after clinical death.
Kontorshchicova C.N., Buchneva E.A., Peretyagin S.P., Mukhina I.V. and Britova E.I.
Medical Institute, Gorky, USSR.
White nonlinear rats were used in the experiments to induce clinical death by sharp bloodless due to carotid artery cut. 5 minutes after the respiration and contractions stopped, hearts were isolated and perfused by Krebs-Henzelite solution during 60 minutes with gas mixture. The latter contained 95 % O2 and 5 % CO2 in control group. As for experimental series 4 sessions of perfusate ozonation were carried out during 5 minutes with ozone concentrations of 0.047 mg/L of air-oxygen mixture.
During perfusation cardiac contraction function was established by electrophysiological data. Creatin phosphate (CP) and adenosyl nucleotide content in myocardial were determined at the end of experiment.
Myocardial ischemia, being developed during the clinical death, caused the change of direction for synthesis, transport and consumption of macroergic phosphates. It resulted in the decrease of CP and ATP levels as well as, arterial pressure decreasing the blood flow (reduction of blood volume/minute). Cardiac activity was restored in 60 min perfusion both in control and experimental series.
However, much higher recordings of developed pressure and maximum velocity of cardiac contractions were reliably received. In similar, reduction of adenosyl nucleotide levels and CP content unchanged in control group, while increased of 47 % in ozonation. A high positive ATP-CP correlation was established, being + 0.754; CP-Cardiac contraction rate correlation being + 0.759.
It can be concluded that in ozonation much faster CP re-synthesis takes place. It may be assumed that ozone oxidating the final glycolisis products, accumulated due ischemia, contributed to a more marked activation of oxidative phosphorilation compared to oxygen.
As a result, ATP and CP increase is being observed, which leads to more quick and complete cardiac activity restoration.
________________________________________ |
The application of 03 ozone in cardiac surgery for the correction structural and metabolic changes in the organs under cardiopulmonary bypass.
Zelenov D.H., Peretyagin S.P., Boyarinov G.A. and Tarasova A.I.
Comfi-Center, Gorky, USSR.
The cardiopulmonary bypass (CB) on the open heart surgery, as rule. Is accompanied by development of hypoxic changes in organs and tissues, specially in microcireulation (MC). That is why the problem of elaboration of preventive methods of bloodmicroreology disorders is still actual in theses conditions. Besides, the positive influence of small ozone dose to MC with hypoxia is well-known fact.
The aim of this work is to study the influence of perfusate oxygenation, by doses of 03/02 between 0.05 - 0.1 mg/L, to M.C. in the Jejunelleum intestine and abdomen organs (liver, kidney) under CB. Experiments on dogs showed that under CB, with usual oxygenation of perfusate (control series) deep disorders in MC of organs are observed: blood ecstasy in venules and capillaries; decrease of functioning capillaries; increase of plasmatic capillaries and diffuse blood stroke.
The destructive change of erythrocytes: their swelling; destruction of membrane; regrouping of electronic dense material take place too. The results of researches on animals, in experimental series, indicated that ozone has positive influence in the state of MC: greater number of functioning capillaries are revealed; destruction of permeability of vessels and deep destructive changes of erythrocytes is not determined.
Ozone, improving MC in organs, intensifies the transfer of oxygen to tissue, preventing the hypoxia. The dystrophic changes in hepato-nephrocytes were reduced; normalization of glycometabolism and preservation of ATP are presented in liver and kidneys.
Thus, ozone, under CB, prevents the development of irreversible changes in the abdomen organs. The method, perfusate ozonating under the CB, is presently used in clinical practice with the aim to prevent the complications in the liver and kidneys after open heart surgery.
________________________________________ |
O3 Ozone: Its sterilising capacity and effects on the interaction of the surface immunological system of the virus which produces hepatitis.
With the aim to eliminate the frequent contamination in our environment of biological reagents, produced in our Institution, with diagnostic purposes for HVB reservoirs, ozone was applied in different concentrations to biological samples, belonging to the diagnostic set ELISA HBs GIRON, which were treated previous incubation to environmental temperature with sodium azide for the HBs antigen and for antibody of covering, as well as timerosal for the antibody conjugated to peroxidase.
All the samples were applied microbiological studies with antibiogram and determination of the different components of the immune system studied, before and after the treatment with ozone.
It was demonstrated that the contaminating agent was Pseudomona aureoginosa, which was eliminated with ozone. No antigenicity of HBsAg was evidenced and the sensitivity of the conjugate decreased 14.1 times after the treatment with ozone.
________________________________________ |
Study of the behavior of different species of microoorganisms with the O3 ozone treatment.
The power of ozone in its disinfecting action is widely known and used. This paper deals with the study of different bacterias in presence of ozone as to: Pseudomonas aeruginosa (ATCC 27853); Pseudomonas aeruginosa wild strain and Pseudomonas fluorescens, isolated from natural and waste waters respectively; E. coli (ATCC 25922) and E. coliisolated from sewage.
All these microorganisms were suspended in buffer saline phosphate to pH 7. The ozone concentration used in experiments was increasing from 0.1 to 0.5 mg/L and with times of permanence in reactor between 0 and 33 min, depending on the case.
Significant differences among the bacterial species studied were observed in regard to their behavior in presence of the inactivation with ozone.
________________________________________ |
Study in vitro of antibacterial action of O3 ozonized sunflower oil.
In Cuba, ozonized oil has been successfully used in microbial etiology affections. Its biocidal nature has been equally proved before, in pathogenicity strains P2 like <1>Candida tropicalis. The aim of this work consisted in prove the antibacterlal action of ozonized oil.
In ATCC strains recomended for these studies; from which we know its behavior before numerous antibiotics and chemical therapeutical agents, as well as its action spectrum before isolated strains in different hospitals. The macrodilution-in-tube and radial diffusion techniques were applied in the Mueller-Hinton environment. All the experiences were developed twice at 37 oC.
The results suggested that ozonized sunflower oil is more effective against GRAM positive microorganisms than in GRAM negative Microorganisms, increasing the inhibition halo as peroxides index of ozonized oil peroxides increase.
he bacteria most resistant was Pseudomonas aeruginosaand the most sensitive one was Staphylococcus aureus.
________________________________________ |
Evaluation of fungicide action of ozone in presence of Aspergillus flavus and in vegetable products contaminated with this fungus.
The fungus contamination of raw materials and fodder, in special by toxicogenic fungi, is one of the main calamities in the economic aspect, since many control methods affect the nutritional properties of the food and others are not sufficiently efficacious.
The objective of this study was to evaluate the effect of ozone in the elimination of Aspergillus flavus in aqueous suspension of conidium and vegetable products contaminated.
With an ozone concentration of 60 mg/L, the micocidal effect was achieved in 4 min. In vegetables products as corn, barley and oats, a concentration of 11 mg/L during 60 min was recommended, in order to obtain a maximum reduction of the fungi concentration from 107 to 103.
________________________________________ |
Disinfection of the air in a specialized climatization area.
Experiments in 2 areas were carried out, measuring the microbial concentration of the air and the surfaces at the beginning and at the end of each employed treatment. Moreover, a control of survival of strains in a third area was performed.
For the contamination, microbial strains of reference Bacillus subtilis, Staphylococcus aureus,Pseudomonas aeruginosa, Aspergillus versicolum were used. The equipments used for the experiments were a formalizer-neutralizer (Paragerms) and an ozonator SOLOZONO NORTE (Spain), both with a programmer of fixed time.
The results were satisfactory through both methods with a reduction of the contaminating flora of more than 10 % in the air. In the surfaces a reduction factor bigger than 3 logarithms was obtained. Also, an interdependence for the strains and contaminated surfaces was observed.
________________________________________ |
Source: Ozone Research Center
|