The Importance of Clinically Relevant Research When Making Comparisons

The Importance of Clinically Relevant Research When Making Comparisons We wish to thank the authors for their efforts in Australia to acquire and evaluate this product “PhaseOne” which is a form of pure hypochlorous acid (HOCl) not available in their country and that has never been marketed there.1 The authors of this study have previously published quite interesting and significant findings regarding dried biofilms on hard surfaces in the healthcare environment.2-4 In this particular study, though, standard methods that might be considered directly applicable to disinfection of soiled hard surfaces and equipment5 – involving addition of 5% bovine calf serum (BCS) to simulate surface “soiling” – were inappropriately applied to the evaluation of the potential performance of antiseptics in the surgical breast pocket at time of implant placement. Based on actual protein measurements of pocket fluid taken at the time of breast implant surgery (as noted below), the use of the 5% bovine serum in the experiments in this study, and the excessively high relative resultant levels of protein in relation to the amount of antiseptic tested, has produced invalid findings that have no relevance to the actual clinical setting. Although the results of this study appear at first glance to suggest that HOCl cannot kill bacteria in a clinical environment, the study upon closer evaluation has actually only confirmed the critical role that the “law of mass action “or more specifically the “mass action ratio” plays in the mechanism of HOCl in the body. HOCl is naturally produced by our white blood cells in a complex reaction associated with an oxidative burst. It is also cleared by the US FDA for use in various applications including as a skin and wound cleanser, with a quite substantial supporting body of published literature.6,7 White blood cells produce approximately 3 ppm each of hypochlorous acid and the reason we are alive to read these comments is because of the “law of mass action ratio.” Each time your body is invaded by bacteria and foreign material, or you have a surgical procedure, your normal body’s immune response is to overwhelm the proteinaceous material, serum, albumin, and bacteria associated with this environment. Typically, a healthy individual generates more than enough HOCl to carry out these critical functions. The key factor, here again, that cannot be overstressed is the amount of exogenous HOCl that is used in the surgical procedure. The problem with the standard hard surface/equipment testing methodology described by the authors, is that it does not take into account the critical variable of the adequate relative volume of HOCl, and thus total molar concentration of excess unbound HOCl, that occurs clinically. As detailed below, the authors’ chosen molar concentrations of protein as BCS relative to HOCl is not clinically relevant. The study noted that: “To test products with 5% BCS, 5 µL of BCS was added to 95 µL of neat or diluted test antiseptic before testing. Contact time was for 15 minutes and 24 hours.” If we assume that 5 µL of BCS contains 0.00325 g of protein or 0.0000325 moles of protein with 3 to 4 HOCl binding sites per mole of protein, and 95 µL of full strength (0.025%) HOCl contains 0.00361 moles HOCl, then assuming a 3:1 ratio of HOCl binding sites on BCS protein, then you might briefly have 0.009 moles of excess HOCl but further dilutions (50%, 25%, 12.5%, 6.25%, 3.13%, 1.56%, and 0.78%) guarantees the negative outcomes in antimicrobial activity observed under such conditions. Stoichiometric neutralization of proteins and thus HOCl, which the authors effectively achieved in their in vitro experiments, does not, however, relate to the real world clinical situation in breast implant surgery. The actual amount of functional HOCl that remains in the presence of clinically measured protein levels in the surgical pocket is more than enough to both penetrate a biofilm and kill any associated bacteria or residual material. In a recent in vitro study, PhaseOne was compared to three popular antimicrobial agents utilized in surgery today: betadine, 0.5% chlorhexidine gluconate and triple antibiotic solution. Pure HOCl in amber glass demonstrated superior antibiofilm capability against mature Ralstonia pickettii biofilms grown on three types of breast implant surfaces in current use. While triple antibiotic was dropped from the study for inability to show effectiveness against planktonic forms of R. pickettii, chlorhexidine did not penetrate any biofilms tested, while betadine took 2 hours of soak time in order to be effective, compared to PhaseOne which showed complete eradication in the first 5-minute time point.8 To assess clinical relevance of these findings within the surgical pocket, a secondary in vivo study was conducted to analyze the total protein content of fluid within the surgical breast pocket in 28 consecutive human clinical samples in patients undergoing subglandular and submuscular augmentation. Following analysis by an independent laboratory, the molar comparison of total protein/albumin to available HOCl molecules (based on use of the undiluted product per label instructions) was calculated. This study was performed as a follow up to demonstrate the clinical effectiveness of a time tested, clinically proven antimicrobial molecule (HOCl) with robust evidence of its broad-spectrum effectiveness. In the majority of cases, an estimated 1000 times more hypochlorous acid than needed to carry out its critical functions of penetrating biofilm and killing bacteria was present. In one case of revision surgery following capsular contracture, in which the greatest amount of protein and albumin was measured, there was still over 250 times the amount required for clinical effectiveness. Results were also calculated for theoretical residual amounts of fluid that might be left in the pocket of 20, 40, and even 60 mL. Using 100 mL of 0.025% PhaseOne in all of these groups was still more than capable of providing relevant functional activity. Most importantly, PhaseOne was able to provide these antimicrobial benefits without the risk of cytotoxicity to fibroblasts and keratinocytes. We all have to thank Dr. John Tebbetts who years ago pointed out the importance of creating a dry surgical pocket during breast surgery. We cannot imagine any of one of us leaving 60 mL of blood and serum in the pocket prior to inserting an implant, but again even in this unlikely scenario, 100 mL of 0.025% PhaseOne is still more than sufficient to carry out its critical functions. It is important to not mix HOCl with saline prior to its use since this will dilute and compromise its efficacy. We understand that data on protein levels in the breast implant surgical pocket may not have been measured or available previously or known to the study authors, but this serves as another example of the critical importance of basing in vitro studies on actual in vivo conditions to the extent possible, and avoiding reliance on assumptions or test standards conditions that may not actually represent real world clinical situations. It is important to point out that Betadine has a long clinical history validating its use and efficacy in surgery. A great deal of data have been collected over the years documenting its numerous benefits. However, Betadine, too, is inactivated by organic macromolecules such as proteins in the breast pocket. The availability of sustained released iodophors in the chronic wound healing market are specifically designed to control the release of iodine into the wound bed to prevent rapid inactivation. Betadine is not produced by the body’s own white blood cells, while HOCl is. HOCl works by oxidation and the basic chemistry of the “law of mass action ratio” means that the volume used clinically is a critical factor and any in vitro experiments conducted must replicate clinically relevant conditions to provide valid findings. In summary, using real world clinical measurements of actual protein levels present in the surgical pocket during breast implant surgery – and not levels present in standard test methods used for disinfection of soiled hard surfaces/equipment – our data document that only a small portion of PhaseOne is likely to be neutralized by the minimal amount of exudative serum and proteinaceous material in a breast pocket, and more than sufficient active antiseptic is available to produce intended efficacy. In the end, all our work has one goal, which is to provide the best solutions for patients’ benefit. As PhaseOne has no known contraindications for its use, and Betadine is not tolerated in patients who have allergies to iodine and hyperthyroidism, among others, an alternative intraoperative irrigation is a beneficial option for the prevention of postsurgical complications. We look forward to continuing this discussion in upcoming published studies regarding the safe, rapid, and effective methods by which PhaseOne can benefit the surgeon and patient during breast surgery. Disclosures Dr. Fisher is a Founder, Chief Medical Officer, and Owner at Integrated Healing Technologies. Laboratory. Dr Brindle is the Chief Clinical Officer of Integrated Healing Technologies and has equity incentive ownership with the company. Dr Porter is the Chief Scientific Advisor for Integrated Healing Technologies; Chief Scientific Officer, Dragon Bio-Consultants, Ltd.; Chairman, President, and CEO of VDDI Pharmaceuticals; and President, PharmacoTherapy Consultants, LLC. Funding The authors received no financial support for the research, authorship, and publication of this article. REFERENCES 1. Hu H, Sleiman J, Johani K, Vickery K. Hypochlorous Acid Versus Povidone-Iodine Containing Irrigants: Which Antiseptic is More Effective for Breast Implant Pocket Irrigation? Aesthet Surg J . 2017. doi: 10.1093/asj/sjx213. [Epub ahead of print]. 2. Vickery K, Deva A, Jacombs Aet al.   Presence of biofilm containing viable multiresistant organisms despite terminal cleaning on clinical surfaces in an intensive care unit. J Hosp Infect . 2012; 80( 1): 52- 55. Google Scholar CrossRef Search ADS PubMed  3. Hu H, Johani K, Gosbell IBet al.   Intensive care unit environmental surfaces are contaminated by multidrug-resistant bacteria in biofilms: combined results of conventional culture, pyrosequencing, scanning electron microscopy, and confocal laser microscopy. J Hosp Infect . 2015; 91( 1): 35- 44. Google Scholar CrossRef Search ADS PubMed  4. Johani K, Abualsaud D, Costa DMet al.   Characterization of microbial community composition, antimicrobial resistance and biofilm on intensive care surfaces. J Infect Public Health . 2017;pii. doi: 10.1016/j.jiph.2017.10.005. 5. Rutala WA, Peacock JE, Gergen MF, Sobsey MD, Weber DJ. Efficacy of hospital germicides against adenovirus 8, a common cause of epidemic keratoconjunctivitis in health care facilities. Antimicrob Agents Chemother . 2006; 50( 4): 1419- 1424. Google Scholar CrossRef Search ADS PubMed  6. Wang L, Bassiri M, Najafi Ret al.   Hypochlorous acid as a potential wound care agent: part I. Stabilized hypochlorous acid: a component of the inorganic armamentarium of innate immunity. J Burns Wounds . 2007; 6: e5. Google Scholar PubMed  7. Robson MC, Payne WG, Ko Fet al.   Hypochlorous Acid as a Potential Wound Care Agent: Part II. Stabilized Hypochlorous Acid: Its Role in Decreasing Tissue Bacterial Bioburden and Overcoming the Inhibition of Infection on Wound Healing. J Burns Wounds . 2007; 6: e6. Google Scholar PubMed  8. Brindle CT, Porter S, Bijlani Ket al.   Preliminary Results of the Use of a Stabilized Hypochlorous Acid Solution in the Management of Ralstonia Pickettii Biofilm on Silicone Breast Implants. Aesthet Surg J . 2017 Dec 12. doi: 10.1093/asj/sjx229. [Epub ahead of print]. © 2018 The American Society for Aesthetic Plastic Surgery, Inc. Reprints and permission: journals.permissions@oup.com http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Aesthetic Surgery Journal Oxford University Press

The Importance of Clinically Relevant Research When Making Comparisons

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Publisher
Mosby Inc.
Copyright
© 2018 The American Society for Aesthetic Plastic Surgery, Inc. Reprints and permission: journals.permissions@oup.com
ISSN
1090-820X
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1527-330X
D.O.I.
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Abstract

We wish to thank the authors for their efforts in Australia to acquire and evaluate this product “PhaseOne” which is a form of pure hypochlorous acid (HOCl) not available in their country and that has never been marketed there.1 The authors of this study have previously published quite interesting and significant findings regarding dried biofilms on hard surfaces in the healthcare environment.2-4 In this particular study, though, standard methods that might be considered directly applicable to disinfection of soiled hard surfaces and equipment5 – involving addition of 5% bovine calf serum (BCS) to simulate surface “soiling” – were inappropriately applied to the evaluation of the potential performance of antiseptics in the surgical breast pocket at time of implant placement. Based on actual protein measurements of pocket fluid taken at the time of breast implant surgery (as noted below), the use of the 5% bovine serum in the experiments in this study, and the excessively high relative resultant levels of protein in relation to the amount of antiseptic tested, has produced invalid findings that have no relevance to the actual clinical setting. Although the results of this study appear at first glance to suggest that HOCl cannot kill bacteria in a clinical environment, the study upon closer evaluation has actually only confirmed the critical role that the “law of mass action “or more specifically the “mass action ratio” plays in the mechanism of HOCl in the body. HOCl is naturally produced by our white blood cells in a complex reaction associated with an oxidative burst. It is also cleared by the US FDA for use in various applications including as a skin and wound cleanser, with a quite substantial supporting body of published literature.6,7 White blood cells produce approximately 3 ppm each of hypochlorous acid and the reason we are alive to read these comments is because of the “law of mass action ratio.” Each time your body is invaded by bacteria and foreign material, or you have a surgical procedure, your normal body’s immune response is to overwhelm the proteinaceous material, serum, albumin, and bacteria associated with this environment. Typically, a healthy individual generates more than enough HOCl to carry out these critical functions. The key factor, here again, that cannot be overstressed is the amount of exogenous HOCl that is used in the surgical procedure. The problem with the standard hard surface/equipment testing methodology described by the authors, is that it does not take into account the critical variable of the adequate relative volume of HOCl, and thus total molar concentration of excess unbound HOCl, that occurs clinically. As detailed below, the authors’ chosen molar concentrations of protein as BCS relative to HOCl is not clinically relevant. The study noted that: “To test products with 5% BCS, 5 µL of BCS was added to 95 µL of neat or diluted test antiseptic before testing. Contact time was for 15 minutes and 24 hours.” If we assume that 5 µL of BCS contains 0.00325 g of protein or 0.0000325 moles of protein with 3 to 4 HOCl binding sites per mole of protein, and 95 µL of full strength (0.025%) HOCl contains 0.00361 moles HOCl, then assuming a 3:1 ratio of HOCl binding sites on BCS protein, then you might briefly have 0.009 moles of excess HOCl but further dilutions (50%, 25%, 12.5%, 6.25%, 3.13%, 1.56%, and 0.78%) guarantees the negative outcomes in antimicrobial activity observed under such conditions. Stoichiometric neutralization of proteins and thus HOCl, which the authors effectively achieved in their in vitro experiments, does not, however, relate to the real world clinical situation in breast implant surgery. The actual amount of functional HOCl that remains in the presence of clinically measured protein levels in the surgical pocket is more than enough to both penetrate a biofilm and kill any associated bacteria or residual material. In a recent in vitro study, PhaseOne was compared to three popular antimicrobial agents utilized in surgery today: betadine, 0.5% chlorhexidine gluconate and triple antibiotic solution. Pure HOCl in amber glass demonstrated superior antibiofilm capability against mature Ralstonia pickettii biofilms grown on three types of breast implant surfaces in current use. While triple antibiotic was dropped from the study for inability to show effectiveness against planktonic forms of R. pickettii, chlorhexidine did not penetrate any biofilms tested, while betadine took 2 hours of soak time in order to be effective, compared to PhaseOne which showed complete eradication in the first 5-minute time point.8 To assess clinical relevance of these findings within the surgical pocket, a secondary in vivo study was conducted to analyze the total protein content of fluid within the surgical breast pocket in 28 consecutive human clinical samples in patients undergoing subglandular and submuscular augmentation. Following analysis by an independent laboratory, the molar comparison of total protein/albumin to available HOCl molecules (based on use of the undiluted product per label instructions) was calculated. This study was performed as a follow up to demonstrate the clinical effectiveness of a time tested, clinically proven antimicrobial molecule (HOCl) with robust evidence of its broad-spectrum effectiveness. In the majority of cases, an estimated 1000 times more hypochlorous acid than needed to carry out its critical functions of penetrating biofilm and killing bacteria was present. In one case of revision surgery following capsular contracture, in which the greatest amount of protein and albumin was measured, there was still over 250 times the amount required for clinical effectiveness. Results were also calculated for theoretical residual amounts of fluid that might be left in the pocket of 20, 40, and even 60 mL. Using 100 mL of 0.025% PhaseOne in all of these groups was still more than capable of providing relevant functional activity. Most importantly, PhaseOne was able to provide these antimicrobial benefits without the risk of cytotoxicity to fibroblasts and keratinocytes. We all have to thank Dr. John Tebbetts who years ago pointed out the importance of creating a dry surgical pocket during breast surgery. We cannot imagine any of one of us leaving 60 mL of blood and serum in the pocket prior to inserting an implant, but again even in this unlikely scenario, 100 mL of 0.025% PhaseOne is still more than sufficient to carry out its critical functions. It is important to not mix HOCl with saline prior to its use since this will dilute and compromise its efficacy. We understand that data on protein levels in the breast implant surgical pocket may not have been measured or available previously or known to the study authors, but this serves as another example of the critical importance of basing in vitro studies on actual in vivo conditions to the extent possible, and avoiding reliance on assumptions or test standards conditions that may not actually represent real world clinical situations. It is important to point out that Betadine has a long clinical history validating its use and efficacy in surgery. A great deal of data have been collected over the years documenting its numerous benefits. However, Betadine, too, is inactivated by organic macromolecules such as proteins in the breast pocket. The availability of sustained released iodophors in the chronic wound healing market are specifically designed to control the release of iodine into the wound bed to prevent rapid inactivation. Betadine is not produced by the body’s own white blood cells, while HOCl is. HOCl works by oxidation and the basic chemistry of the “law of mass action ratio” means that the volume used clinically is a critical factor and any in vitro experiments conducted must replicate clinically relevant conditions to provide valid findings. In summary, using real world clinical measurements of actual protein levels present in the surgical pocket during breast implant surgery – and not levels present in standard test methods used for disinfection of soiled hard surfaces/equipment – our data document that only a small portion of PhaseOne is likely to be neutralized by the minimal amount of exudative serum and proteinaceous material in a breast pocket, and more than sufficient active antiseptic is available to produce intended efficacy. In the end, all our work has one goal, which is to provide the best solutions for patients’ benefit. As PhaseOne has no known contraindications for its use, and Betadine is not tolerated in patients who have allergies to iodine and hyperthyroidism, among others, an alternative intraoperative irrigation is a beneficial option for the prevention of postsurgical complications. We look forward to continuing this discussion in upcoming published studies regarding the safe, rapid, and effective methods by which PhaseOne can benefit the surgeon and patient during breast surgery. Disclosures Dr. Fisher is a Founder, Chief Medical Officer, and Owner at Integrated Healing Technologies. Laboratory. Dr Brindle is the Chief Clinical Officer of Integrated Healing Technologies and has equity incentive ownership with the company. Dr Porter is the Chief Scientific Advisor for Integrated Healing Technologies; Chief Scientific Officer, Dragon Bio-Consultants, Ltd.; Chairman, President, and CEO of VDDI Pharmaceuticals; and President, PharmacoTherapy Consultants, LLC. Funding The authors received no financial support for the research, authorship, and publication of this article. REFERENCES 1. Hu H, Sleiman J, Johani K, Vickery K. Hypochlorous Acid Versus Povidone-Iodine Containing Irrigants: Which Antiseptic is More Effective for Breast Implant Pocket Irrigation? Aesthet Surg J . 2017. doi: 10.1093/asj/sjx213. [Epub ahead of print]. 2. Vickery K, Deva A, Jacombs Aet al.   Presence of biofilm containing viable multiresistant organisms despite terminal cleaning on clinical surfaces in an intensive care unit. J Hosp Infect . 2012; 80( 1): 52- 55. Google Scholar CrossRef Search ADS PubMed  3. Hu H, Johani K, Gosbell IBet al.   Intensive care unit environmental surfaces are contaminated by multidrug-resistant bacteria in biofilms: combined results of conventional culture, pyrosequencing, scanning electron microscopy, and confocal laser microscopy. J Hosp Infect . 2015; 91( 1): 35- 44. Google Scholar CrossRef Search ADS PubMed  4. Johani K, Abualsaud D, Costa DMet al.   Characterization of microbial community composition, antimicrobial resistance and biofilm on intensive care surfaces. J Infect Public Health . 2017;pii. doi: 10.1016/j.jiph.2017.10.005. 5. Rutala WA, Peacock JE, Gergen MF, Sobsey MD, Weber DJ. Efficacy of hospital germicides against adenovirus 8, a common cause of epidemic keratoconjunctivitis in health care facilities. Antimicrob Agents Chemother . 2006; 50( 4): 1419- 1424. Google Scholar CrossRef Search ADS PubMed  6. Wang L, Bassiri M, Najafi Ret al.   Hypochlorous acid as a potential wound care agent: part I. Stabilized hypochlorous acid: a component of the inorganic armamentarium of innate immunity. J Burns Wounds . 2007; 6: e5. Google Scholar PubMed  7. Robson MC, Payne WG, Ko Fet al.   Hypochlorous Acid as a Potential Wound Care Agent: Part II. Stabilized Hypochlorous Acid: Its Role in Decreasing Tissue Bacterial Bioburden and Overcoming the Inhibition of Infection on Wound Healing. J Burns Wounds . 2007; 6: e6. Google Scholar PubMed  8. Brindle CT, Porter S, Bijlani Ket al.   Preliminary Results of the Use of a Stabilized Hypochlorous Acid Solution in the Management of Ralstonia Pickettii Biofilm on Silicone Breast Implants. Aesthet Surg J . 2017 Dec 12. doi: 10.1093/asj/sjx229. [Epub ahead of print]. © 2018 The American Society for Aesthetic Plastic Surgery, Inc. Reprints and permission: journals.permissions@oup.com

Journal

Aesthetic Surgery JournalOxford University Press

Published: Apr 1, 2018

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