Potential Activity of Micafungin and Amphotericin B Co-Encapsulated in Nanoemulsion against Systemic Candida auris Infection in a Mice Model.

The Candida auris species is a multidrug-resistant yeast capable of causing systemic and lethal infections. Its virulence and increase in outbreaks are a global concern, especially in hospitals where outbreaks are more recurrent. In many cases, monotherapy is not effective, and drug combinations are opted for. However, resistance to antifungals has increased over the years. In view of this, nanoemulsions (NEs) may represent a nanotechnology strategy in the development of new therapeutic alternatives. Therefore, this study developed a co-encapsulated nanoemulsion with amphotericin B (AmB) and micafungin (MICA) (NEMA) for the control of infections caused by C. auris. NEs were developed in previous studies. Briefly, the NEs were composed of a mixture of 10% sunflower oil and cholesterol as the oil phase (5:1), 10% Polyoxyethylene (20) cetyl ether (Brij® 58) and soy phosphatidylcholine as surfactant/co-surfactant (2:1), and 80% PBS as the aqueous phase. The in vivo assay used BALB/c mice weighing between 25 and 28 g that were immunosuppressed (CEUA/FCF/CAr n° 29/2021) and infected with Candida auris CDC B11903. The in vivo results show the surprising potentiate of the antifungal activity of the co-encapsulated drugs in NE, preventing yeast from causing infection in the lung and thymus. Biochemical assays showed a higher concentration of liver and kidney enzymes under treatment with AmB and MICAmB. In conclusion, this combination of drugs to combat the infection caused by C. auris can be considered an efficient therapeutic option, and nanoemulsions contribute to therapeutic potentiate, proving to be a promising new alternative.

Nanoemulsion Increases the Antifungal Activity of Amphotericin B against Four Candida auris Clades: In Vitro and In Vivo Assays.

Candida auris is an emerging yeast of worldwide interest due to its antifungal resistance and mortality rates. The aim of this study was to analyse the in vitro and in vivo antifungal activity of a nanoemulsion loaded with amphotericin B (NEA) against planktonic cells and biofilm of C. auris clinical isolates belonging to four different clades. In vivo assays were performed using the Galleria mellonella model to analyse antifungal activity and histopathological changes. The in vitro results showed that NEA exhibited better antifungal activity than free amphotericin B (AmB) in both planktonic and sessile cells, with >31% inhibition of mature biofilm. In the in vivo assays, NEA demonstrated superior antifungal activity in both haemolymph and tissue. NEA reduced the fungal load in the haemolymph more rapidly and with more activity in the first 24 h after infection. The histological analysis of infected larvae revealed clusters of yeast, immune cells, melanisation, and granulomas. In conclusion, NEA significantly improved the in vitro and in vivo antifungal activity of AmB and could be considered a promising therapy for C. auris infections.

Anti-Candida auris activity in vitro and in vivo of micafungin loaded nanoemulsions.

Fungi are becoming increasingly resistant, especially the new strains. Therefore, this work developed nanoemulsions (NE) containing micafungin (MICA), in order to improve its action against infections caused by Candida auris. The NEs were composed of the surfactants polyoxyethylene (20) cetyl ether (Brij 58®)/soy phosphatidylcholine at 10%, sunflower oil/cholesterol at 10%, and 80% PBS. The NEs were characterized by Dynamic Light Scattering (DLS). For the microbiological in vitro evaluation the determination of the minimum inhibitory concentration (MIC), ergosterol/sorbitol, time kill and biofilms tests were performed. Additionally, the antifungal activity was also evaluated in a Galleria mellonella model. The same model was used in order to evaluate acute toxicity. The NE showed a size of ∼ 42.12 nm, a polydispersion index (PDI) of 0.289, and a zeta potential (ZP) of -3.86 mV. NEM had an average size of 41.29 nm, a PDI of 0.259, and a ZP of -4.71 mV. Finally, both nanoemulsions showed good stability in a storage period of 3 months. Although NEM did not show activity in planktonic cells, it exhibited action against biofilm and in the in vivo infection model. In the alternative in vivo model assay, it was possible to observe that both, NEM and free MICA at 0.2 mg/l, was effective against the infection, being that NEM presented a better action. Finally, NEM and free MICA showed no acute toxicity up to 4 mg/l. NEM showed the best activities in in vitro in mature antibiofilm and in alternative in vivo models in G. mellonella. Although, NEs showed to be attractive for MICA transport in the treatment of infections caused by C. auris in vitro and in vivo studies with G. mellonella, further studies should be carried out, in mice, for example.

Candida auris is a fungus that can cause infections in the human body. As it is a microorganism with a high potential for resistance, it is extremely important to develop new therapeutic alternatives. Thus, nanotechnology, the science that studies materials with extremely small sizes, can be considered a promising method in the treatment of these infections.

Lycopene, Mesoporous Silica Nanoparticles and Their Association: A Possible Alternative against Vulvovaginal Candidiasis?

Commonly found colonizing the human microbiota, Candida albicans is a microorganism known for its ability to cause infections, mainly in the vulvovaginal region known as vulvovaginal candidiasis (VVC). This pathology is, in fact, one of the main C. albicans clinical manifestations, changing from a colonizer to a pathogen. The increase in VVC cases and limited antifungal therapy make C. albicans an increasingly frequent risk in women's lives, especially in immunocompromised patients, pregnant women and the elderly. Therefore, it is necessary to develop new therapeutic options, especially those involving natural products associated with nanotechnology, such as lycopene and mesoporous silica nanoparticles. From this perspective, this study sought to assess whether lycopene, mesoporous silica nanoparticles and their combination would be an attractive product for the treatment of this serious disease through microbiological in vitro tests and acute toxicity tests in an alternative in vivo model of Galleria mellonella. Although they did not show desirable antifungal activity for VVC therapy, the present study strongly encourages the use of mesoporous silica nanoparticles impregnated with lycopene for the treatment of other human pathologies, since the products evaluated here did not show toxicity in the in vivo test performed, being therefore, a topic to be further explored.

Cetyltrimethylammonium bromide in the synthesis of mesoporous silica nanoparticles: General aspects and in vitro toxicity.

Pharmaceutical nanotechnology has become a trend with incalculable advantages in the applicability of systems in the controlled, safe and effective release of drugs. Among the nanotechnological nanoparticles, the mesoporous silica nanoparticles stand out, a system with significant biocompatibility, good physical chemical stability, greater surface contact area with desirable and adjustable pore structure. Once developed and well defined, these pores can carry drugs and control their release. However, to create this type of nanoparticle is essencial to use surfactants since they act as pore template. Among the most important surfactants, cetyltrimethylammonium bromide (CTAB) highlights, a quaternary ammonium compound widely used as a surfactant in the synthesis of mesoporous silica nanoparticles (MSNs), hollow mesoporous silica (HMSNs) and core-shell MSNs. However, for achieving good results of drug-loaded pores it is necessary to remove CTAB by extraction techniques, which provides pores formation throughout the silica and the incorporation of molecules. During and after the removal process, it is possible that CTAB residues remains inside the pores, despide several removal processes are described as efficient in the complete removal of surfactants. In turn, the presence of CTAB residues can be advantageous, especially when considering its antimicrobial activity. Meanwhile, it should be noted that the presence of CTAB may present high toxicity risks. This review seeks to explore not only general aspects of the use of CTAB in the synthesis of MSNs, but also to assess its toxicity in prokaryotic and eukaryotic cells, in order to determine whether CTAB residues are acceptable in MSNs that will be used as drug delivery systems for further in vivo and clinical assays.

Natural product-based nanomedicine applied to fungal infection treatment: A review of the last 4 years.

Fungal infections are one of the main public health problems, especially in immunocompromised patients, nosocomial environments, patients with chronic diseases, and transplant recipients. These diseases are increasingly frequent and lethal because the microorganism has a high capacity to acquire resistance to available therapy. The main resistance factors are the emergence of new strains and the uncontrolled use of antifungals. It is, therefore, important to develop new methods that contribute to combating fungal diseases in the clinical area. Natural products have considerable potential for the development of new drugs with antifungal activity, mainly due to their biocompatibility and low toxic effect. This promising antimicrobial activity of natural products is mainly due to the presence of flavonoids, terpenes, and quinones, which explains their antifungal potential. Pharmaceutical nanotechnology has been explored to enhance the delivery, selectivity, and clinical efficacy of these products. Nanotechnological systems provide a safe and selective environment for various substances, such as natural products, improving antifungal activity. However, further safety experiments (in vivo or clinical trials) need to be carried out to prove the therapeutic action of natural products, since they may have undesirable, toxic, and mutagenic effects. Therefore, this review article addresses the main nanotechnological methods using natural products for effective future treatment against the main fungal diseases.

Reflection about the hemodialysis water microbiological quality in Brazil

Abstract Dialysis has been widely used in the treatment of patients with chronic kidney diseases and is considered a global public health issue. This treatment, which has changed the prognosis and quality of life in patients with chronic renal failure, can lead to complications that are often fatal. For this reason, there is a need for validation of alternative tests that favor the monitoring of treated water for dialysis in real-time to promote and prevent injuries to patients submitted to this procedure.

Validation of an innovative analytical method for simultaneous quantification of curcumin and fluconazole using high-performance liquid chromatography from nanostructured lipid carriers.

Vulvovaginal candidiasis is a public health problem with a high incidence among female patients. Currently, there is an increase in the identification of Candida spp. resistant to current therapy, making it necessary to search for new therapeutic alternatives. The synergistic potential of curcumin with fluconazole is described in the literature. However, due to its high lipophilicity, it is necessary to use drug-delivery systems to adequately explore its potential, among which is the nanostructured lipid carrier. However, to date, there is no validated method of high-performance liquid chromatography for simultaneous determination of fluconazole and curcumin in the literature. Thus, the present work developed a high-performance liquid chromatography method for simultaneous determination of fluconazole and curcumin co-encapsulated in nanostructured lipid carrier which was validated according to the International Conference on Harmonization (Technical Requirements for Registration of Pharmaceuticals for Human Use) - Q2 (R1) and the Food and Drug Administration - Guidance for Bioanalytical Method. The method was applied to determine the encapsulation efficiency and drug-loading of curcumin and fluconazole in nanostructured lipid carriers. The developed method proved to be selective, precise, accurate, and robust for the simultaneous determination of both drugs, enabling the quantification of encapsulation efficiency and drug-loading of curcumin and fluconazole in nanostructured lipid carriers.

Prevalence of vulvovaginal candidiasis in Brazil: A systematic review.

Vulvovaginal candidiasis (CVV) is a condition in which signs and symptoms are related to inflammation caused by Candida spp infection. It is the second leading cause of vaginitis in the world, representing a public health problem. The present systematic review comes with the proposal of analyze and identify the available evidence on CVV prevalence in Brazil, pointing out its variability by regions. For this, a systematic literature review was carried out with meta-analysis of cross-sectional and cohort studies, following the Preferred Reporting Items for Systematic Reviews and Meta-Analyzes (PRISMA) guide recommendations, and was registered in the International Prospective Register of Systematic Reviews (PROSPERO 2020 CRD42020181695). The databases used for survey were LILACS, Scielo, Scopus, PUBMED, Web of Science and CINAHL. Fifteen studies were selected to estimate CVV prevalence in the Brazilian territory. South and Southeast regions have higher prevalences than the North and Northeast regions, no data were found for the Midwest region. The estimated prevalence for Brazil is 18%, however, it is suggested that this number is higher due to underreporting and the presence of asymptomatic cases. Therefore, new epidemiological studies are recommended throughout Brazil, to elucidate the profile of this disease in the country, in addition to assisting in the elaboration of an appropriate prevention plan by state. LAY SUMMARY: Data found in the literature regarding the epidemiological profile of vulvovaginal candidiasis in Brazil are obsolete and incomplete, so the present systematic review has the proposal to analyze and identify the evidence on vulvovaginal candidiasis prevalence in Brazil. The estimated prevalence is 18%; however, this number can be higher.

Highlights in poloxamer-based drug delivery systems as strategy at local application for vaginal infections.

Infectious diseases related to the vagina include diseases caused by the imbalance of the vaginal flora and by sexually transmitted infections. Some of these present themselves as a public health problem due to the lack of efficient treatment that leads to their complete cure, and others due to the growing resistance to drugs used in therapy. In this sense, new treatment strategies are desirable, with vaginal administration rout being a great choice since can bypass first-pass metabolism and decrease drug interactions and adverse effects. However, it is worth highlighting limitations related to patient's discomfort at application time. Thereby, the use of poloxamer-based drug delivery systems is desirable due its stimuli-sensitive characteristic. Therefore, the present review reports a brief overview of poloxamer properties, biological behavior and advances in poloxamer applications in controlled drug release systems for infectious diseases related to the vagina treatment and prevention.

Nanosystem functionalization strategies for prostate cancer treatment: a review.

Prostate cancer (PC) has a high morbidity and mortality rate worldwide, and the current clinical guidelines can vary depending on the stage of the disease. Drug delivery nanosystems (DDNs) can improve biopharmaceutical properties of encapsulated anti-cancer drugs by modulating their release kinetics, improving physicochemical stability and reducing toxicity. DDN can also enhance the ability of specific targeting through surface modification by coupling ligands (antibodies, nucleic acids, peptides, aptamer, proteins), thus favouring the cell internalisation process by endocytosis. The purposes of this review are to describe the limitations in the treatment of PC, explore different functionalization such as polymeric, lipid and inorganic nanosystems aimed at the treatment of PC, and demonstrate the improvement of this modification for an active target, as alternative and promising candidates for new therapies.

Chitosan-based systems aimed at local application for vaginal infections.

Vaginal administration is a promising route for the local treatment of infectious vaginal diseases since it can bypass the first-pass metabolism, drug interactions, and adverse effects. However, the commercial products currently available for topical vulvovaginal treatment have low acceptability and do not adequately explore this route. Mucoadhesive systems can optimize the efficacy of drugs administered by this route to increase the retention time of the drug in the vaginal environment. Several polymers are used to develop mucoadhesive systems, among them chitosan, a natural polymer that is highly biocompatible and technologically versatile. Thus, the present review aimed to analyze the studies that used chitosan to develop mucoadhesive systems for the treatment of local vaginal infections. These studies demonstrated that chitosan as a component of mucoadhesive drug delivery systems (DDS) is a promising device for the treatment of vaginal infectious diseases, due to the intrinsic antimicrobial activity of this biopolymer and because it does not interfere with the effectiveness of the drugs used for the treatment.

Highlighting the use of micro and nanoparticles based-drug delivery systems for the treatment of Helicobacter pylori infections.

Due to the high adaptability of Helicobacter pylori and the low targeting specificity of the drugs normally used in pharmacological therapy, the strains are becoming increasingly resistant to these drugs, making it difficult to eradicate the infection. Thus, the search for new therapeutic approaches has been considered urgent. The incorporation of drugs in advanced drug delivery systems, such as nano and microparticles, would allow the improvement of the retention time in the stomach and the prolongation of drug release rates at the target site. Because of this, the present review article aims to highlight the use of micro and nanoparticles as important technological tools for the treatment of H. pylori infections, focussing on the main nanotechnological systems, including nanostructured lipid carriers, liposomes, nanoemulsion, metallic nanoparticles, and polymeric nanoparticles, as well as microtechnological systems such as gastroretentive dosage forms, among them mucoadhesive, magnetic and floating systems were highlighted.

An Overview of Properties and Analytical Methods for Lycopene in Organic Nanocarriers.

Lycopene (LYC), a natural compound responsible for the red color of some fruits like pink grapefruit, red guava, watermelon, papaya and, mainly, present in tomatoes (Solanum lycopersicum). LYC has been extensively studied because of its various pharmacological properties such as antioxidant, cardioprotective, hypocholesterolemic, antineophasic, photoprotection, antidiabetic and antimicrobial activity. However, LYC uses in therapy is limited due to its insolubility in aqueous solvents, resulting in low bioavailability and stability. In order to overcome these drawbacks, it is essential to use of organic nanocarriers for LYC controlled release. Up to now, the description of LYC-loaded organic nanocarriers are scarce, mainly related to organic nanosystems based on lipid nanostructures such as nanoemulsions (NE), liposomes (LP), niosomes (NI), nanostructured lipid carriers (NLC) and solid lipid nanoparticles (SLN). Taking into account the development of new formulations, is indispensable the use of sensitive and suitable analytical methods previously validated. Among the analytical methods described here, high-performance liquid chromatography (HPLC) stands out due to its good accuracy, precision and desirable detection limit. In this review, we highlights the main biological and physicochemical properties of LYC, as well as LYC-based organic nanocarriers for controlled drug delivery and the analytical methods described in literature to determine LYC in any kind of matrix.

Highlights in Mesoporous Silica Nanoparticles as a Multifunctional Controlled Drug Delivery Nanoplatform for Infectious Diseases Treatment.

Infectious diseases are a major global concern being responsible for high morbidity and mortality mainly due to the development and enhancement of multidrug-resistant microorganisms exposing the fragility of medicines and vaccines commonly used to these treatments. Taking into account the scarcity of effective formulation to treat infectious diseases, nanotechnology offers a vast possibility of ground-breaking platforms to design new treatment through smart nanostructures for drug delivery purposes. Among the available nanosystems, mesoporous silica nanoparticles (MSNs) stand out due their multifunctionality, biocompatibility and tunable properties make them emerging and actual nanocarriers for specific and controlled drug release. Considering the high demand for diseases prevention and treatment, this review exploits the MSNs fabrication and their behavior in biological media besides highlighting the most of strategies to explore the wide MSNs functionality as engineered, smart and effective controlled drug release nanovehicles for infectious diseases treatment. Graphical Abstract Schematic representation of multifunctional MSNs-based nanoplatforms for infectious diseases treatment.

Recent advances in hydrogels as strategy for drug delivery intended to vaginal infections.

Vaginal infections represent a clear women health problem due to the several issues as high recurrence rate, drug resistence and emergence of persistent strains. However, achieving improvements in therapeutic efficacy by using conventional formulations intended to vaginal drug delivery remains as a challenge due to anatomy and physiology of the vagina, since the secretion and renewal of vaginal fluids contribute to the removal of the dosage form. Hydrogels have been widely exploited aiming to achieve drug delivery directly into vaginal mucosa for local therapy due to their attractive features as increased residence time of the drug at the action site and control of drug release rates. Some polymers can aggregate specific properties to hydrogels as mucoadhesive, stimuli-responsive and antimicrobial, improving their interaction with the biological interface and therapeutic response. In this review, we highlight the advances, advantages and challenges of the hydrogels as drug and/or nanocarrier vehicles intended to the treatment of vaginal infections, emphasizing also the polymers and their properties more explored on the design these systems to improve the therapeutic effect on the vaginal tissue. In addition, this review can contribute for better exploitation these systems in search of new local treatments for bacterial vaginosis, candidiasis and trichomoniasis.

Coqueluche na infância: revisão integrativa da doença / Pertussis in children: an integrative review / Tos ferina en la infancia: revisión integrativa de la enfermedad

Justificativa e Objetivos: Apesar dos esforços despendidos pelo Programa Nacional de Imunizações, a coqueluche é um agravo que ainda gera grandes preocupações para a saúde pública. Assim, o objetivo deste estudo foi descrever as evidências na literatura sobre coqueluche na infância que indicassem as práticas de assistência à saúde necessárias para sua prevenção, promoção e controle. Método: Revisão integrativa da literatura nas bases de dados LILACS, MedLine, PubMed, SciELO, Embase, Cumulative Index to Nursing and Allied Health Literature (CINAHL) utilizando os descritores Whooping Cough, Epidemiology, Vaccination. A revisão foi realizada em maio de 2020 e abrangeu a literatura disponível na íntegra, publicada em português, inglês ou espanhol, sem delimitação de ano. Resultados: Dentre os 1.248 artigos selecionados apenas nove foram analisados (a maioria norte-americana), dos quais 55% eram estudos realizados para prevenção de coqueluche. Quanto à autoria dos estudos, em 67% das publicações havia participação de um epidemiologista. Conclusão: Esta investigação mostra as evidências científicas sobre a coqueluche na infância, necessárias para prevenir, controlar, promover a assistência e definir o perfil epidemiológico desse agravo nessa população. Os resultados revelaram nível de evidência cinco, sugerindo que as pesquisas desenvolvidas na área não retratam fortes evidências.(AU)

Background and Objectives: Despite the efforts of the National Immunization Program, pertussis remains a serious health problem. Thus, this study described the evidence in the literature on pertussis during childhood, which elucidates the health-care practices necessary for prevention, promotion, and control of the disease. Methods: This is an integrative literature review conducted in the LILACS, MedLine, PubMed, SciELO, Embase, Cumulative Index to Nursing and Allied Health Literature (CINAHL) databases, using the following descriptors: "Whooping Cough", "Epidemiology", and "Vaccination". The search was restricted to humanities, in English, Portuguese, or Spanish, without year delimitation. The search was completed on May 2020, Results: in total, 1,248 articles were selected, of which only nine articles were analyzed, mostly from North America, and 55% of the analyzed studies were about preventing pertussis. Regarding the authorship of the studies, epidemiologists participated in 67% of the publications. Conclusion: This study presents the necessary scientific evidence on pertussis during childhood is necessary for prevention, control, promotion, and epidemiological profile of this disease in this population. The results revealed a level of evidence of five, suggesting that the research in this area does not show strong evidence.(AU)

Justificación y Objetivos: La tos ferina todavía es un grave problema de salud a pesar de los esfuerzos realizados por el Programa Nacional de Inmunización. El objetivo de este estudio fue identificar las evidencias científicas en la literatura sobre la tos ferina en la infancia que indican las prácticas de asistencia sanitaria necesarias a la prevención, promoción y control de la enfermedad. Métodos: Revisión integrativa de la literatura en las bases de datos LILACS, MedLine, PubMed, SciELO, Embase, Cumulative Index to Nursing and Allied Health Literature (CINAHL) utilizando las palabras clave: Whooping Cough, Epidemiology, Vaccination. Se llevó a cabo en mayo de 2020 la búsqueda de artículos publicados en portugués, inglés y español, disponibles en su totalidad y sin delimitación de año. Resultados: Se seleccionaron un total de 1.248 artículos, de los cuales solo se analizaron nueve, en su mayoría norteamericanos; el 55% de los estudios analizados fueron sobre la prevención de la tos ferina. Respecto de la autoría de los estudios, un epidemiólogo participó en el 67% de las publicaciones. Conclusiones: Esta investigación muestra evidencias científicas sobre la tos ferina en la infancia necesarias para la prevención, el control, la promoción de la asistencia y la definición del perfil epidemiológico de la enfermedad en esa población. Los resultados revelaron que el nivel de evidencia se remite a cinco, lo que sugiere que la investigación en el área no muestra evidencias sólidas.(AU)

Nanosystems against candidiasis: a review of studies performed over the last two decades.

The crescent number of cases of candidiasis and the increase in the number of infections developed by non-albicans species and by multi-resistant strains has taken the attention of the scientific community, which has been searching for new therapeutic alternatives. Among the alternatives found the use of nanosystems for delivery of drugs already commercialized and new biomolecules have grown, in order to increase stability, solubility, optimize efficiency and reduce adverse effects. In view of the growing number of studies involving technological alternatives for the treatment of candidiasis, the present review came with the intention of gathering studies from the last two decades that used nanotechnology for the treatment of candidiasis, as well as analysing them critically and pointing out the future perspectives for their application with this purpose. Different studies were considered for the development of this review, addressing nanosystems such as metallic nanoparticles, mesoporous silica nanoparticles, polymeric nanoparticles, liposomes, nanoemulsion, microemulsion, solid lipid nanoparticle, nanostructured lipid carrier, lipidic nanocapsules and liquid crystals; and different clinical presentations of candidiasis. As a general overview, nanotechnology has proven to be an important ally for the treatment against the diversity of candidiasis found in the clinic, whether in increasing the effectiveness of commercialized drugs and reducing their adverse effects, as well as allowing exploring more effectively properties therapeutics of new biomolecules.

Validation and applicability of an alternative method for dialysis water and dialysate quality analysis.

INTRODUCTION: In hemodialysis, patients are exposed to a large volume of water, which may lead to fatal risks if not meeting quality standards. This study aimed to validate an alternative method for monitoring microbiological quality of treated water and assess its applicability in dialysis and dialysate analysis, to allow corrective actions in real-time. METHODS: Validation and applicability were analyzed by conventional and alternative methods. For validation, E. coli standard endotoxin was diluted with apyrogenic water in five concentrations. For the applicability analysis, treated water for dialysis was collected from different points in the treatment system (reverse osmosis, drainage canalization at the storage tank bottom, reuse, and loop), and dialysate was collected from four machines located in different rooms in the hemodialysis sector. RESULTS: The validation results were in accordance with the Brazilian Pharmacopoeia acceptance criteria, except for the last two concentrations analyzed. In addition, the ruggedness criterion performed under the US Pharmacopoeia was in agreement with the results. DISCUSSION: A limiting factor in the applicability analysis was the absence of the endotoxin maximum permitted level in dialysate by the Brazilian legislation. When comparing the analysis time, the alternative method was more time-consuming than the conventional one. This suggests that the alternative method is effective in the case of few analyses, that is, real-time analyses, favoring corrective actions promptly. On the other hand, it does not support the implementation of the alternative method in a laboratory routine due to the high demand for analyses.

Aplicabilidade de métodos alternativos no monitoramento da qualidade microbiológica da água tratada para diálise / Applicability of alternative methods for the monitoring microbiological quality of treated water for dialysis

A fim de garantir a qualidade final de produtos os laboratórios de análise microbiológica fornecem dados sobre a qualidade dos mesmos em todas as suas etapas de produção. A crescente preocupação com a saúde dos pacientes conduz à busca de métodos que forneçam resultados precisos e rápidos, pois possibilitam que ações corretivas sejam tomadas em tempo real. O presente trabalho teve por objetivo avaliar o potencial de tecnologia alternativa no monitoramento de endotoxina bacteriana na água tratada para diálise e dialisato e avaliar o potencial da citometria de fluxo na análise de água. Para isso utilizou-se Portable Test System (PTS®) como método alternativo para detecção de endotoxina bacteriana no monitoramento da água tratada para diálise e dialisato, o qual foi validado frente ao método convencional farmacopeico. Paralelamente realizou-se revisão narrativa da literatura a fim de avaliar a aplicabilidade da citometria de fluxo em análises de água. A análise dos diferentes parâmetros de validação para endotoxina bacteriana no método alternativo mostrou que, exceto para a menor diluição analisada, houve linearidade e precisão nos resultados. Por outro lado a concentração de 0,25 UE/mL foi a menor que apresentou exatidão e especificidade. Observou-se ainda, que o limite de detecção foi de 0,125UE/mL e o de quantificação de 0,25 UE/mL, portanto o intervalo foi de 0,25-1,0 UE/mL. Adicionalmente pela análise de resistência pode-se perceber que ao variar analistas não houve diferença significativa. Em relação ao tempo de análise em uma condição de rotina laboratorial com muitas amostras, o PTS® mostrou-se demorado. Ressalta ainda, que seria importante que a legislação vigente determinasse a análise mensal de endotoxinas no dialisato. A revisão da literatura evidencia o potencial da tecnologia de citometria de fluxo, pois a mesma mostrou-se satisfatória quando comparada a metodologias convencionais para análise de água. O trabalho desenvolvido permitiu concluir que o PTS®) mostrou-se adequado para analisar amostras in loco, permitindo análises em tempo real, que para as quais haja a expectativa de ausência de endotoxinas ou de concentração respeitando o intervalo de 0,25 UE/mL a 1,0 UE/mL. Quanto a citometria de fluxo, esta mostrou-se uma tecnologia promissora em analisar amostras de água, sendo portanto recomendável proceder a estudos de validação e aplicabilidade

In order to guarantee the final quality of products, the microbiological analysis laboratories provide data about their quality at all production stages. The growing concern for patients' health leads to the search for methods that provide accurate and fast results, as they enable corrective actions to be taken in real time. The present work aimed to evaluate the alternative technology potential in the monitoring of bacterial endotoxin in treated water for dialysis and dialysate and to evaluate the potential of flow cytometry in water analysis. The different validation parameters analysis for bacterial endotoxin in alternative method showed that, except for the lowest dilution analyzed, there was linearity and precision in the results. On the other hand, the concentration of 0.25 EU / mL was the lowest that presented accuracy and specificity. It was further observed that the detection limit was 0.125UE / mL and the quantification limit was 0.25 EU / mL, so the range was 0.25-1.0 EU / mL. Additionally by the ruggedness analysis it was possible to perceived that when varying analysts there was no significant difference. Regarding the analysis time in a laboratory routine condition with many samples, the PTS® was was time consuming. It was also observed that it would be important to determine monthly analysis of endotoxins in dialysate. The literature review evidence the flow cytometry technology potential of the because it was satisfactory when compared to conventional methodologies for water analysis. The research showed that the PTS® was suitable for analyzing samples in loco, allowing real-time analyzes, for which there is expectation of endotoxins absence or concentration respecting the range of 0.25 EU / mL to 1.0 EU / mL. For the flow cytometry, it was shown to be a promising technology for analyzing water samples, and it is therefore advisable to carry out validation and applicability studies