Safety, pitfalls, and misunderstandings about the use of ozone therapy as a regenerative medicine tool. A narrative review.

Despite various opinions and healthy controversy on Ozone Therapy (OT), the practices of this therapy have increased worldwide. Main areas of study with consistent scientific outcomes are the topical treatment of both disk herniation and periodontal disease. On the other hand, there is a net dissociation of the scientific resonance concerning systemic oxygen/ozone treatments. It is our intention to discuss in logical terms the numerous papers that commendably reported adverse reactions attributable to OT, focusing our attention mainly to the techniques of administration and not to the simple contact of ozone with biological material. The case reports on OT treatments safety concerns discussed on international journals, make it possible to state that most safety issues are secondary to infections or traumatic reactions due to malpractice. Commonly, the molecule of ozone itself is not responsible of severe reactions at the therapeutic modalities. The millions of patients treated so far from the thousands of physicians correctly practicing OT world widely in the last 40 years demonstrate the safety of this simple and cost-effective regenerative medicine tool. The promising therapeutic implications also for the current COVID-19 emergency are a further stimulus to the standardization of this therapeutic resource with multiple application specificities.

The use of oxygen ozone therapy in the treatment of cervicobrachial pain: case series study.

This retrospective, observational, uncontrolled case series study was carried out to evaluate the clinical efficacy and safety of intramuscular paravertebral injections of an oxygen-ozone (O2-O3) mixture in patients with cervicobrachial pain. One hundred and sixty-eight subjects affected by cervicobrachial pain, referred to Ozone Therapy Unit at San Pietro Fatebenefratelli Hospital in Rome (Italy), were enrolled in the study. All the subjects (n=168, 106 females and 62 males) completed the treatment and the follow-up visits. Subjects received 12 cervical intramuscular injections of an O2-O3 mixture (5 mL) with an O3 concentration of 16 µg/mL once a week. The overall reduction of pain was measured by the change in mean of Visual Analogue Scale (VAS) score from baseline to the end of treatment and from baseline to one, two, three, four and five years of follow-up. Patient satisfaction was assessed at the end of treatment, by modified MacNab Questionnaire. Possible adverse events related to the treatment were recorded. The mean (± standard deviation) VAS pain score at baseline, at the end of treatment, and during the follow-up at one, two, three, four and five years were 7.82 (±1), 1.6 (±1.5), 1.5 (±1.4), 1.4 (±1.3), 1.6 (±1.2), 1.5 (±1.3) and 1.60 (±1.2), respectively, showing a significant reduction in pain over time (p<0.001). Of 156 patients who responded to treatment, 128 (82.05%) were pain free at one year, 110 (70.51%) at second year, 103 (66.02%) at third year, 94 (60.25%) at fourth year and 86 (55.12%) at fifth year follow-up visit. According to pain distribution all subjects showed a significant reduction in pain over time in each group (p<0.05). No significant differences were observed between groups. No serious adverse events were observed during the entire study. We suggest the use of intramuscular paravertebral injections of an oxygen-ozone (O2-O3) mixture in patients with cervicobrachial pain as an effective and safe treatment option to consider before surgical intervention.

A critical evaluation of the use of ozone and its derivatives in dentistry.

OBJECTIVE: The therapeutic application of ozone and its derivatives in the dental field has been used for many purposes. However, there has yet to be a consistent evaluation of the outcomes, due to the lack of standardization of the treatment operating procedures. MATERIALS AND METHODS: The keywords "ozone", "ozonated", "ozonation" "ozonized", "ozonization", "dentistry", "periodontology", "oral surgery", "oxygen-ozone therapy" were used to perform a literature review using PubMed, Cochrane, Google Scholar, Zotero databases with the temporal restriction for manuscripts published between 2010 and 2020. Clinical trials and case reports of good, neutral, as well as negative results related to ozone treatment specifications were evaluated. DISCUSSION: A better understanding of the mechanisms of action of this bio-oxidative therapy could open new horizons related to the personalization of treatments and the quality of dental care. The critical condition to achieve these goals is an improved knowledge of the qualitative/quantitative characteristics of ozone and its derivatives.

Ozone: A Multifaceted Molecule with Unexpected Therapeutic Activity.

A comprehensive outline for understanding and recommending the therapeutic use of ozone in combination with established therapy in diseases characterized by a chronic oxidative stress is currently available. The view of the absolute ozone toxicity is incorrect, because it has been based either on lung or on studies performed in artificial environments that do not correspond to the real antioxidant capacity of body compartments. In fact, ozone exerts either a potent toxic activity or it can stimulate biological responses of vital importance, analogously to gases with prospective therapeutic value such as NO, CO, H2S, H2, as well as O2 itself. Such a crucial difference has increasingly become evident during the last decade. The purpose of this review is to explain the aspects still poorly understood, highlighting the divergent activity of ozone on the various biological districts. It will be clarified that such a dual effect does not depend only upon the final gas concentration, but also on the particular biological system where ozone acts. The real significance of ozone as adjuvant therapeutic treatment concerns severe chronic pathologies among which are cardiovascular diseases, chronic obstructive pulmonary diseases, multiple sclerosis, and the dry form of age-related macular degeneration. It is time for a full insertion of ozone therapy within pharmaceutical sciences, responding to all the requirements of quality, efficacy and safety, rather than as either an alternative or an esoteric approach.

What is the best strategy for enhancing the effects of topically applied ozonated oils in cutaneous infections?

Owing to diabetes, atherosclerosis, and ageing, there are several million patients undergoing skin lesions degenerated into infected ulcers with very little tendency to heal and implying a huge socioeconomical cost. Previous medical experience has shown that the daily application of ozonated oil eliminates the infection and promotes a rapid healing. The purpose of the study is the optimization of the antimicrobial effect of ozonated oils by testing in vitro four bacterial species and one yeast without or in the presence of different amounts of human serum. The results obtained suggest that a gentle and continuous removal of debris and exudate is an essential condition for the potent bactericidal effect of ozonated oils. In fact, even small amounts of human serum inactivate ozone derivatives and protect bacteria. The application of ozonated oil preparations is very promising in a variety of skin and mucosal infections. Moreover, ozonated oils are far less expensive than antibiotic preparations.

Ozonated oils as functional dermatological matrices: effects on the wound healing process using SKH1 mice.

Wound tissue repair is a complex and dynamic process of restoring cellular structures and tissue layers. Improvement of this process is crucial for several pathologies characterized by chronic delayed wound closure such as diabetes, and the investigation of new approaches aimed to ameliorate the wound healing process is under continuous evolution. Recently, the usage of vegetable matrices in the form of ozonated oils has been proposed and several researchers have shown a positive effect in the wound, based on their bactericidal, antiviral, and antifungal properties. The present study was undertaken to compare the effect that different ozonated oils (olive, sesame and linseed) with the same level of ozonation have on wound healing rate in SKH1 mice. Several histological parameters and the level of key proteins such as VEGF and PCNA have been analyzed. Only treatment with ozonated sesame oil shows a faster wound closure in the first 7 days. This effect paralleled with the increased VEGF and PCNA levels, NFκB nuclear translocation and 4-HNE formation. The present study shows that not only the ozonation grade is of importance for the improvement of wound healing process but also the typical composition of the oil.

Ozone and ozonated oils in skin diseases: a review.

Although orthodox medicine has provided a variety of topical anti-infective agents, some of them have become scarcely effective owing to antibiotic- and chemotherapeutic-resistant pathogens. For more than a century, ozone has been known to be an excellent disinfectant that nevertheless had to be used with caution for its oxidizing properties. Only during the last decade it has been learned how to tame its great reactivity by precisely dosing its concentration and permanently incorporating the gas into triglycerides where gaseous ozone chemically reacts with unsaturated substrates leading to therapeutically active ozonated derivatives. Today the stability and efficacy of the ozonated oils have been already demonstrated, but owing to a plethora of commercial products, the present paper aims to analyze these derivatives suggesting the strategy to obtain products with the best characteristics.

Packing properties of starch-based powders under mild mechanical stress.

This study reports the ability to settle of commercial pharmaceutical grade starch samples, both native and pregelatinized. The experiments were carried out under different relative humidity (RH%) conditions and the packing properties were evaluated using both official pharmacopoeial monograph conditions and also modified conditions in order to give a deeper knowledge of tapping under mild mechanical stress. The technique adopted, simulating common pharmaceutical operating practices, appears to be useful to estimate some technologically relevant features of diluent powder materials. Moreover, a general mathematical function has been applied to the experimental data; this could be appropriate for adequately describing material settling patterns and offers practical parameters for characterizing starch powders within the context of a pharmaceutical quality system.

How much ozone bactericidal activity is compromised by plasma components?

AIMS: Evaluation of bactericidal effect of different concentrations of ozone when used (a) as a gas, or (b) dissolved in saline. The addition of hydrogen peroxide or 4-hydroxynonenal dissolved in saline was also tested, as well as the effect of human plasma. METHODS AND RESULTS: Staphylococcus aureus, methicillin-resistant Staph. aureus (MRSA), and Pseudomonas aeruginosa, suspended in their culture media were tested. While all bacteria suspended in protein-free saline were killed at high ozone concentrations, they survived when as little as 5% human plasma was present. Hydrogen peroxide was 100-fold less active than ozone and needed to remain in contact with bacteria for at least 60 min. 4-hydroxynonenal (2 micromol l(-1)) was inhibitory for proliferation of both Staph. aureus and MRSA, but not for Ps. aeruginosa. CONCLUSIONS: Ozone and the cascade of its derivative products are potent bactericidal agents, but even small amounts of human plasma, hence of hydro- and liposoluble antioxidants, in bacterial suspensions inhibit oxidation and protect bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: Any substantial in vivo cytocidal effect of ozone and its derivatives can be excluded. On the other hand, topical and continuous action of various ozone preparations remains valuable in a variety of skin and mucosal infections.

Commercial pregelatinized starches: ability to settle and rheological evaluation.

The ability to settle and the tribological properties of three samples of pregelatinized starch widely available commercially were investigated with reference to their technological implications. In particular, we evaluated the influence of storage conditions, storage time, moisture content, equipment, handling, and mechanical treatment on the apparent volume, poured density and tapped density, as well as on tests of flow parameters such as basic flowability energy. The results obtained highlight the fact that the technologically relevant behaviour of pregelatinized starches in terms of apparent volume and tribological properties may be influenced not only by relative humidity (RH%) conditions, but also by common pharmaceutical operating practices. In particular, previous mechanical treatment can lead to statistically different behaviour in tests of ability to settle according to pharmacopoeial specifications, depending on both RH% conditioning and the amount of powder, although the materials under investigation all conform to the respective monograph in the Pharmacopoeia. All these aspects are of special interest with regard to the various pharmaceutical formulations which include pregelatinized starch, both on an industrial scale and in small-scale production.

[Cleaning validation of the prefermentation-fermentation group for production of biomedical products]. / Cleaning validation di un gruppo prefermentatore-fermentatore per la produzione di prodotti biomedicali.

The fermentation-group is largely employed to produce pharmaceuticals by means of biotechnology techniques. The operative functions of these plants are surely kept under control by the validation process, while its cleaning is less investigated. The aim of this work is the cleaning validation of a fermentation-group. Furthermore, Standard Operative Procedures (SOPs) have been validated relating to the production necessities and to the allowed safety limits.

Effects of surfactants on the spectral behaviour of calcein (II): a method of evaluation.

The spectral behavior of calcein, a water-soluble self quenching fluorescent marker often used in biomedical analysis, can be considerably affected by the presence of surfactants. With this study we intend to obtain further information on the photophysical properties of calcein, in the presence of surfactants and in the concentration range commonly used to investigate the release of such marker from vesicle dispersions. The experiments were carried out both in water and in a physiological buffer (HEPES, pH 7.5), in the presence of Triton X-100, sodium dodecyl sulphate and centyltrimethylammonium bromide, both below and above their critical micelle concentration (c.m.c.). The obtained results confirm that calcein fluorescence can be affected by the presence of surfactants. Thus, environmental conditions must always be carefully checked for the actual quantitative evaluation of this dye. Furthermore, this study sheds some light on the nature and mechanism of calcein quenching.

Effects of surfactants on the spectral behaviour of calcein.

The spectral behaviour of calcein, a water-soluble self-quenching molecule that is often used as a marker in biomedical analysis, can be considerably affected by the presence of surfactants. The aim of this work was to investigate the spectral properties of calcein under conditions that are particularly significant in liposome studies. For this purpose the fluorescence and absorbance of this dye were determined in solutions of ionic and non-ionic surfactants (cetyltrimethylammonium bromide, sodium dodecyl sulphate and Triton X-100), at different concentrations below and above their critical micelle concentrations (c.m.c.) as well as in the presence of phospholipids in the form of liposomes and/or mixed micelles. Cationic surfactants induced changes in lambda max, absorbance and fluorescence but these changes were less noticeable in the phospholipid dispersions. The anionic and the non-ionic surfactants induced mainly changes in fluorescence intensity.