Antimicrobial Effectiveness Test Research Articles

Study of the antimicrobial preservatives effectiveness test in development of the combined syrup composition

Мета: метою нашої роботи було обґрунтування та дослідження з вивчення ефективності антимікробних консервантів у розробці складу основи комбінованого сиропу на основі рослинної сировини, призначеного для лікування захворювань гепатобіліарної системи та жовчного міхура. Методи: застосовувався біологічний фармакопейний метод дослідження (ефективність антимікробних консервантів). Результати: за результатами проведених досліджень з вивчення ефективності низки антимікробних консервантів у зразках сиропів були отримані наступні дані. Зразки сиропу № 1 і № 2 (з ніпагін : ніпазолом (3 : 1) 0,1 і 0,2 %), зразки № 3 і № 4 (з калію сорбатом 0,1 % і 0,2 %), зразки № 5 і № 6 (з натрію бензоатом 0,1 % і 0,2 %) відповідали критеріям оцінки ефективності антимікробних консервантів, що рекомендовані ДФУ до лікарських препаратів для орального застосування. Встановлено, що усі експериментальні зразки розроблених сиропів з використанням у складі обраних консервантів є перспективними для подальших досліджень зі створення оральних лікарських форм. Однак, антимікробна ефективність калію сорбату дещо вища (lg зменшення кількості життєздатних клітин Candida albicans АТСС 885-653 дорівнює 3,55; Aspergillus brasiliensis АТСС 16404 – 2,3) і спектр антимікробної дії більш широкий, що сприятиме якості сиропу, який розробляється, також і у процесі його зберігання. Тому для подальших досліджень нами буде використовуватись консервант калію сорбат. Висновки: теоретично обґрунтовано і експериментально доведено доцільність використання у розробці основи комбінованого сиропу комплексної дії такого консерванту як калію сорбат

Lack of sporicidal activity in diallyl-disulphide-oxide and a nanotechnology-product for sterilizing medical and dental instruments

Objective: To evaluate sporicidal activity in products commercially available for sterilization of medical and dental instruments. Method: In simulated use and under worst-case scenario conditions, Bacillus atrophaeus spores were suspended in diallyl disulphide oxide (DDO), a nanotechnology product (NANO) or three SOW solutions. After 6 hour exposure, the spore suspensions were filtered, washed and incubated. Results: The DDO and NANO formulations evaluated lack sporicidal activity and are inadequate to sterilize instruments. The three SOW solutions evaluated are sporicidal but their activities are inhibited by protein. Their respective sterilization contact times remain to be determined. Conclusions: The results identify formulations that pose a risk to patients, and unveil failures in the regulatory evaluation of products. The absence of science-based regulatory standards results in the commercialization of unsafe products without the required sporicidal activity. To strengthen national and local health systems, the international infection prevention community must critically review the regulatory expectations for sporicidal products and disseminate information on the basics of antimicrobial effectiveness testing.

Development of a New Imaging Bulk Package for Multipatient Use in MDCT.

Multidose presentations of U.S. Food and Drug Administration (FDA)-approved radiographic contrast agents have been considered pharmacy bulk packages. However, the use of pharmacy bulk packages for multipatient dosing does not meet the U.S. Pharmacopeia definition of a pharmacy bulk package. The purpose of this study was to validate and gain FDA approval for a new multidose preparation of iopamidol for safe, compliant multipatient dosing in the CT suite. An FDA-approved development program was undertaken to determine whether multidose presentations of iopamidol used in combination with a transfer set remain free of chemical and microbiologic contamination during the labeled maximum hold time after container closure penetration and simulated worst-case handling conditions. The program comprised antimicrobial effectiveness testing of iopamidol-300 and iopamidol-370 containers with seven microbes. Microbial growth was evaluated at five time points up to 28 days after introduction. Microbial ingress testing involved inoculation of four challenge sites with each of four microorganisms for up to 14 hours. Chemical compatibility and extractable testing was performed to ensure chemical integrity. No growth of microorganisms occurred. All evaluated samples remained sterile, indicating no microbial contamination through 14 hours of simulated clinical use. No effect on chemical integrity was found in any of the drawn iopamidol samples meeting the chemical specifications for iopamidol, and no leachable compounds were detected. The absence of any chemical or microbiologic contamination led the FDA to approve the iopamidol multidose container and transfer set as a combination product for multipatient use. The approval resulted in a new U.S. Pharmacopeia category of multidose presentation-the imaging bulk package.

The Efficacy of Essential Oils as Natural Preservatives in Vegetable Oil

ABSTRACTThe efforts for finding the natural preservatives with nontoxicity and nonirritancy have encouraged the scientists to research among the medicinal plants. The preservative efficacy of Daucus carota, Ferula gummosa, Eugenium caryophyllata, Oliveria decumbens, Pelargonium graveolens, Ziziphora tenuir, Acorus calamus, and Trachyspermum ammi essential oils on challenge test's pathogens and on pathogen's inoculated vegetable oil was evaluated by antimicrobial effectiveness test. Carotol (46%), β-pinene (62.7%), eugenol (78.4%), thymol (50.6%), cis-asarone (27.5%), thymol (50.1%), and α-terpineol (19.5%) were the primary main components of D. carota, F. gummosa, E. caryophyllata, T. ammi, A. calamus, O. decumbens, and Z. tenuir essential oils, respectively. A. niger was more sensitive microorganism to oils. The antimicrobial activity of O. decumbens oil was the highest. Different concentrations of essential oils were added to the vegetable oil. The results of test on the vegetable oil showed that the combination of O. decumbens and P. graveolens oils (0.5:0.5%) had enough efficacies as natural preservative in vegetable oil.

Efficacy of Antimicrobials Extracted from Organic Pecan Shell for Inhibiting the Growth of Listeria spp.

Growers and processors of USDA certified organic foods are in need of suitable organic antimicrobials. The purpose of the research reported here was to develop and test natural antimicrobials derived from an all-natural by-product, organic pecan shells. Unroasted and roasted organic pecan shells were subjected to solvent free extraction to produce antimicrobials that were tested against Listeria spp. and L. monocytogenes serotypes to determine the minimum inhibitory concentrations (MIC) of antimicrobials. The effectiveness of pecan shell extracts were further tested using a poultry skin model system and the growth inhibition of the Listeria cells adhered onto the skin model were quantified. The solvent free extracts of pecan shells inhibited Listeria strains at MICs as low as 0.38%. The antimicrobial effectiveness tests on a poultry skin model exhibited nearly a 2 log reduction of the inoculated cocktail mix of Listeria strains when extracts of pecan shell powder were used. The extracts also produced greater than a 4 log reduction of the indigenous spoilage bacteria on the chicken skin. Thus, the pecan shell extracts may prove to be very effective alternative antimicrobials against food pathogens and supplement the demand for effective natural antimicrobials for use in organic meat processing.

Antimicrobial Activity of a New Preservative for Multiuse Ophthalmic Solutions

The aim of this study was to examine the antimicrobial activity and the preservative efficacy of a novel preservative solution containing sodium hydroxymethyl glycinate (SHMG) and edetate disodium (EDTA), which is used for preservation of some commercial ophthalmic formulations. In vitro susceptibility assays were performed against several gram-positive (Staphylococcus aureus, Staphylococcus epidermidis, and Bacillus cereus) and gram-negative (Escherichia coli and Pseudomonas aeruginosa) bacteria representative of the microbial flora of epithelial surfaces or colonizing the conjunctiva, as well as against Candida albicans and Aspergillus niger. Using different concentrations of SHMG alone or in combination with EDTA, the minimal inhibitory and microbicidal concentrations against these organisms were assessed. In addition, 8 brands of multidose eye drops containing 0.002% SHMG and 0.1% EDTA as preservative were tested for antimicrobial activity using the antimicrobial effectiveness test recommended by the international pharmacopoeias. The minimal inhibitory and bactericidal/fungicidal concentration values of SHMG ranged from 0.0025% to 0.0125% for bacteria and from 0.125% to 0.50% for mold and yeast. Susceptibility testing demonstrated that the addition of EDTA substantially increased the SHMG activity against all bacterial and fungal strains. The preservative effectiveness test was applied to commercial eye drops. All the drop solutions met the criteria reported by the U.S. Pharmacopeia for parenteral and ophthalmic preparations. All products also satisfied the major acceptance criteria of the European Pharmacopeia with respect to the antifungal activity. With regard to the antibacterial activity, the less-stringent criteria of the European Pharmacopeia were fulfilled. The present study demonstrates the efficacy of a novel preservative for ophthalmic solutions (SHMG/EDTA) and its activity in protecting selected commercial artificial tears against microbial contamination.

Stability and preservation of a new formulation of epoprostenol sodium for treatment of pulmonary arterial hypertension

BackgroundThe aim of this study was to evaluate the stability and microbiological properties of a formulation of epoprostenol sodium with L-arginine and sucrose excipients (epoprostenol AS).MethodsThe stability of the reconstituted solutions after storage at 5°C and 25°C, diluted solutions (3000–60,000 ng/mL) at controlled room temperature, and diluted solutions (3000–60,000 ng/mL) stored at 5°C and then at room temperature were evaluated. Solutions were prepared using sterile water for injection or sterile saline (sodium chloride 0.9%) for injection. Shelf-life was assessed by determining potency over time relative to initial potency. In this context, potency is synonymous with content. The antimicrobial activity of reconstituted (100,000 ng/mL for 0.5 mg vial, 300,000 ng/mL for 1.5 mg vial) and diluted (3000 ng/mL) epoprostenol AS was measured using an antimicrobial effectiveness test after inoculation with six species of bacteria, yeast, and mold.ResultsReconstituted epoprostenol AS was stable for up to one day’s storage at 25°C or 7 days’ storage at 5°C. Epoprostenol AS was stable for up to 72 hours when diluted, depending on temperature and concentration. The maximum shelf-life of the diluted solution if the reconstituted solution had been stored for up to one day at room temperature or up to 7 days at 5°C, was between 24 and 72 hours, depending on concentration. Following storage of diluted solutions at 5°C for up to 8 days, maximum shelf-life was between one and 2 days, depending on temperature and concentration. Potency was not dependent on diluents. Preservative testing confirmed no microbial growth for any of six organisms tested for at least 14 days at 5°C or 25°C for the reconstituted solution and for at least 16 days at 5°C followed by one day at 25°C for the diluted solutions.ConclusionEpoprostenol AS has favorable thermal stability and does not support the growth of any micro-organism tested for up to 17 days. This extended stability under ambient conditions has the potential to improve convenience further for patients.

Preservative of choice for Prev(e)nar 13™ in a multi-dose formulation

Development of a Prev(e)nar 13™ multi-dose vaccine, in support of vaccinating populations against pneumococcal disease, required the addition of a preservative to the vaccine formulation that met antimicrobial effectiveness tests based on the European Pharmacopoeia (EP) requirements, including deliberate multiple challenge studies and recommendation by the WHO Open Vial Policy. In this study, the antimicrobial effectiveness of several preservatives in Prev(e)nar 13™ formulations was evaluated. A Prev(e)nar 13™ formulation containing 2-Phenoxyethanol (2-PE) at a concentration of 5.0mg/dose was stable and met EP recommended criteria for antimicrobial effectiveness tests when the formulation was kept over a 30 month period. In contrast, a recommended dose of Thimerosal, as a comparator, or other preservatives did not meet EP antimicrobial effectiveness acceptance criteria. The rate of growth inhibition of Thimerosal compared to 2-PE on Staphylococcus aureus, a resilient organism in these tests, was significantly slower in single and multi-challenge studies. These results indicate that 2-PE provides a superior antimicrobial effectiveness over Thimerosal for this vaccine formulation.

Comparison of Compendial Antimicrobial Effectiveness Tests: A Review

The antimicrobial effectiveness or preservative effectiveness test is described in the tripartite compendia for sterile parenteral multi-dose formulated products. The execution of the test is essentially harmonized with respect to inoculum preparation and test execution but not the acceptance criteria. This article describes how a single test can be performed that procedurally satisfies all of the compendia and their acceptance criteria.

Stability of Fortified Cefazolin Ophthalmic Solutions Prepared in Artificial Tears Containing Surfactant-Based Versus Oxidant-Based Preservatives

The aim of this study was to investigate the stability of fortified cefazolin sodium ophthalmic solutions (50 mg mL⁻¹) extemporaneously prepared in commercial artificial tears containing 2 different types of preservatives, namely the surfactants and oxidants. Fortified cefazolin sodium solutions were prepared by reconstituting cefazolin for injection with sterile water and further diluted with Tears Naturale II or Natear, 2 commercial artificial tears containing polyquaternium-1 and sodium perborate, respectively, as preservatives. The solutions were then kept at room temperature (28°C) or in the refrigerator (4°C). During the 28-day period, the formulations were periodically examined for the physical appearance, pH, and the remaining drug concentrations. The antibacterial potency was evaluated as the minimal inhibitory concentration against Staphylococcus aureus strain ATCC 29923 by broth dilution technique. The activity of the preservatives was demonstrated by antimicrobial effectiveness tests. On day 28, the microbial contamination in the preparations was tested. The stability profiles of cefazolin solutions prepared in Tears Naturale II, Natear, and water were not different, but they were significantly influenced by the storage temperature. The refrigerated formulations showed no loss of drug and antibacterial potency as well as alteration of physical appearance and pH throughout the 28 days. In contrast, those kept at room temperature showed gradual change in color and odor. The degradation of drug exceeded 10% from day 3 and the decrease of antibacterial potency could be observed at week 3. All cefazolin solutions prepared in artificial tears retained the antimicrobial activity of preservatives and were free from bacterial and fungal contamination throughout the 28-day period of study. Cefazolin sodium ophthalmic solutions can be extemporaneously prepared in Tears Naturale II or Natear without the influence from different types of preservatives used in these vehicles. The formulation is physically and chemically stable and retains the antibacterial potency for at least 28 days in the refrigerator.

Stability and antimicrobial effectiveness of treprostinil sodium in Sterile Diluent for Flolan

Long-term intravenous infusion of epoprostenol and treprostinil for treatment of pulmonary arterial hypertension (PAH) via a central venous catheter is associated with the risk of bloodstream infection (BSI). While several potential explanations exist for possible differences in BSI incidence among intravenous prostanoids, one hypothesis suggests that the alkaline pH of epoprostenol in Sterile Diluent for Flolan (SDF) has greater antimicrobial activity compared with the neutral pH of other common diluents such as sterile saline or water, which have been used for treprostinil. The chemical stability and antimicrobial activity of 4 microg/ml and 130 microg/ml treprostinil in SDF were assessed according to United States and European Pharmacopeia. At both concentrations, treprostinil in SDF remained stable after incubation at 40 degrees C and ambient relative humidity for up to 52 h. Solution pH also remained stable (range 10.4-10.6), and the solutions were essentially free of particulate at all time points examined. Antimicrobial activity was measured using an antimicrobial effectiveness test after inoculation with five species of bacteria, yeast and mould. The antimicrobial activity of both concentrations of treprostinil met United States Pharmacopeia requirements. Further, the antimicrobial activity of treprostinil in SDF against gram-negative bacteria (> 4 log(10) reductions) exceeded that previously described for treprostinil in sterile saline. These results suggest that dilution of treprostinil with the alkaline solution SDF may reduce the risk of infection from inadvertent patient contamination compared with dilution of treprostinil in sterile saline.

Preservative Evaluation of Novel 2,4-Hexadienoic Acid Derivatives in Aluminium Hydroxide Gel – USP

The potential derivatives of sorbic acid (2,4-hexadienoic acid) from our previous study were subjected to preservative efficacy testing. Aluminium Hydroxide Gel – USP was used as a pharmaceutical product and Staphylococcus aureus MTCC 2901, Bacillus subtilis MTCC 2063, Escherichia coli MTCC 1652, Candida albicans MTCC 227 and Aspergillus niger MTCC 8189 were used as representative challenging microorganisms for antimicrobial effectiveness testing as per USP 2004. The hexadienoic acid derivative, m-nitrosorbanilide has better preservative efficacy than sorbic acid as well as the standard preservatives, methyl paraben and propyl paraben.

Preservative Evaluation of Caprylic Acid Derivatives in Aluminium Hydroxide Gel – USP

The potential derivatives of caprylic acid were subjected to preservative efficacy testing in Aluminium Hydroxide Gel – USP using Staphylococcus aureus MTCC 2901, Bacillus subtilis MTCC 2063, and Escherichia coli MTCC 1652 as representative challenging microorganisms for antimicrobial effectiveness testing as per USP 2004. The caprylic acid derivative, capryl hydrazide exhibited better preservative efficacy than caprylic acid as well as the standard preservatives, methyl paraben and propyl paraben.

Antimicrobial preservative use in parenteral products: Past and present

The following review provides a comprehensive summary of antimicrobial preservatives that are commonly used in licensed parenteral products to date. The information reviewed includes the general properties of the preservatives, the doses and frequency of their use, the classes of the preserved products (peptide, protein, vaccine, and small molecule products), the interactions with other formulation components, and the criteria commonly used for their selection in parental product formulations. It was revealed that phenol and benzyl alcohol are the two most common antimicrobial preservatives used in peptide and protein products, while phenoxyethanol is the most frequently used preservative in vaccines. Benzyl alcohol or a combination of methylparaben and propylparaben are generally found in small molecule parenteral formulations. The key criteria for antimicrobial preservative selection are the preservative's dose, antimicrobial functionality, and effect on the active ingredient. Additionally, the use of spectroscopic techniques (circular dicroism (CD) and fluorescence) and differential scanning calorimetry (DSC) were identified as common techniques used in evaluating an antimicrobial preservative for its impact on the conformational stability of peptide, protein, and vaccine antigens. The future use of preservatives is also discussed, including antimicrobial agents such as peptides, and regulatory requirements for antimicrobial effectiveness testing.

Alteplase as a Catheter Locking Solution: In Vitro Evaluation of Biochemical Stability and Antimicrobial Properties

To reduce potential complications of fibrin deposition to catheter surfaces, there is increasing empiric use of alteplase as a catheter lock solution. The purpose is to evaluate the properties of alteplase when reconstituted in sterile water (SW) or bacteriostatic water (BW) for prolonged periods. Alteplase in glass vials was reconstituted (1 mg/mL) with SW or BW (0.9% benzyl alcohol) in duplicates and stored at 37 degrees C. Biochemical assays were performed at days 0 and 7 and included optical clarity, protein concentration, percent protein monomer, and in vitro clot lysis activity. Microbiologic assays were performed on days 7 through 28 with use of a standardized antimicrobial effectiveness test (pass/fail) and pour-plate methods incubated at 22.5 degrees C (fungus, 3-7 days) or 32.5 degrees C (bacteria, 3-5 days). Organisms tested included Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Candida albicans, and Aspergillus niger. Biochemical assay results were as follows: on day 0, all samples were clear/colorless; protein concentrations were 1.10 mg/mL +/- 0 in SW and 1.11 mg/mL +/- 0 in BW; percent protein monomer was 8.2% +/- 0.07 in SW and 98.6% +/- 0.07 in BW; and in vitro clot lysis activity (in percent of relative activity) was 100% in all samples. On day 7, all samples were clear/colorless, protein concentrations were 1.11 mg/mL +/- 0.07 in SW and 1.11 mg/mL +/- 0.07 in BW; percent protein monomer was 97.4% +/- 0.21 in SW and 96.1% +/- 0.21 in BW; and in vitro clot lysis activity (relative activity compared with day 0) was 91% +/- 2.8 in SW and 90% +/- 2.8 in BW. Microbiologic assays (US Pharmacopeia [USP] antimicrobial effectiveness test) yielded a failing result for alteplase reconstituted in SW and a passing result for alteplase reconstituted in BW. Alteplase reconstituted with SW or BW remains relatively stable with retained bioactivity when stored at 37 degrees C for as long as 7 days. Despite the biochemical similarities of the two solutions, only alteplase in BW met USP criteria as an effective antimicrobial solution. Further clinical evaluation is warranted.

Characterization of an extemporaneous liquid formulation of lisinopril.

The stability of lisinopril in an extemporaneously prepared suspension stored at or below 25 degrees C for 28 days under ambient light exposure was studied. A formulation of 1-mg/mL oral suspension was prepared from commercially available 20-mg lisinopril tablets, using Bicitra and Ora-Sweet SF as the compounding vehicles to make a final volume of 200 mL. Individual samples, stored in 8-oz amber polyethylene terephthalate bottles, were used for each test performed. All samples were stored at 25 degrees C. Appropriateness of the extemporaneous preparation method was performed by shaking three lots of each suspension for 30, 60, and 90 seconds. To test the robustness and reproducibility of the method, two chemists prepared the suspensions from the same three lots of lisinopril tablets. Chemical and physical stability were established by analyzing duplicate samples at time zero and after one, two, four, and six weeks. The solubility of lisinopril was tested from suspensions stored for four weeks. In-use stability was also examined over four weeks. Photochemical stability was examined by exposing three batches of the suspension to maximum light stress in accordance with the International Conference on Harmonization. Antimicrobial-effectiveness testing was also conducted with freshly prepared suspensions and suspensions stored for six weeks. The preparation method used was appropriate and effective. Lisinopril is fully dissolved in the suspension matrix. Satisfactory chemical, physical, and microbiological results were obtained after the suspensions were stored for six weeks at 25 degrees C and 35% relative humidity. Lisinopril suspensions extemporaneously prepared from tablets are stable for at least four weeks when stored at or below 25 degrees C under ambient light exposure.

Pramlintide injection drug product robustness studies.

The article examines the effects of temperature excursions and actual dose withdrawal on the quality of pramlintide injection, a multidose liquid parenteral formulation. Studies were designed to demonstrate product robustness under conditions that may occur during patient use. Pramlintide %Purity was determined by two high-performance liquid chromatography (HPLC) methods, a reversed-phase (RP-HPLC) and a strong-cation exchange (SCX-HPLC) method. A second RP-HPLC method was used to determine pramlintide potency and the concentration of the m-cresol preservative. Antimicrobial preservative effectiveness testing was per USP and European Pharmacopeia (Ph. Eur.). Short-term stability studies were undertaken to probe the effects of the following conditions: 5 degrees C to 40 degrees C and 5 degrees C to -20 degrees C temperature cycling over 10 days; once daily or four-times daily dose withdrawal over 12 or 42 days; and combined 30 degrees C storage and four-times daily dose withdrawal over 42 days. In all cases, pramlintide %Purity and potency values remained essentially unchanged or unchanged relative to controls. Similarly, product appearance, and m-cresol concentration and preservative effectiveness were not significantly affected by the stress conditions used in the 5 studies. Pramlintide injection drug product is extremely robust to challenging stress conditions that may occur during patient use of this multidose product for chronic administration.

Long-Term Storage of Microorganisms Used in Antimicrobial Effectiveness Tests

Liquid nitrogen storage of organisms designated in the U.S. Pharmacopeia (XIX) for the assessment of antimicrobial effectiveness is described. By use of simple apparatus and procedures, 85-95% viabilities of pre-frozen cells are attained immediately upon freezing. Viability levels have remained constant over a 3-year period and the use of such inocula has considerably improved the reliability of the test. The superiority of liquid nitrogen storage for microbial test inocular over other types of storage is discussed.

Methods for Storage of Antimicrobial Effectiveness Test Inoculum Suspensions below Freezing

Studies were conducted to determine the suitability of storage below freezing of some antimicrobial effectiveness test inoculum organisms: Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, and Cattdida albicans. Several solutions used for protecting microorganisms subjected to storage below freezing were compared. Comparison of 10% dextrose with other solutions (distilled water; 0.07M phosphate buffer, pH 7.0, with 15% glycerol; and 0.07M phosphate buffer, pH 7.0, with 7.5% dimethyl sulfoxide) demonstrated that inoculum suspensions were most stable when prepared with 10% dextrose. After storage for 6 months at −50° in a freezer, inoculum suspensions prepared with 10% dextrose retained viability and demonstrated suitability for use in antimicrobial effectiveness tests of dosage forms containing preservatives.