In vitro antimicrobial activity of Luffa operculata Scalia, Rodolfo Alexander; Dolci, José Eduardo Lutaif; Ueda, Suely Mitoi Ykko ...
Brazilian journal of otorhinolaryngology,
07/2015, Volume:
81, Issue:
4
Journal Article
Peer reviewed
Open access
Luffa operculata is probably one of the most popular herbal medicines used in the treatment of rhinitis and rhinosinusitis. However, its specific mechanism of action is still unknown.
To evaluate in ...vitro antibacterial activity of L. operculata against three ordinary agents of upper respiratory tract infection: Staphylococcus aureus, Streptococcus pneumoniae and Streptococcus pyogenes.
Different concentrations of L. operculata alcoholic extract were applied to bacterial broth containing reference and community strains of the three described agents. After a 24-h incubation period, the bacterial culture turbidity was measured. The samples were then inoculated onto Mueller-Hinton and human blood agar plates. Bacterial growth was analyzed after 24- and 48-h incubation period. The test was considered negative when there was no environmental turbidity, confirmed by the absence of bacterial growth into the inoculated plates. Tests were considered positive when either turbidity changes were observed on the bacterial broth or when bacterial growth was detected on inoculated plates. Appropriate statistical analysis of the data was performed.
L. operculata extracts showed antibacterial activity mainly to S. pyogenes followed by S. pneumoniae and S. aureus.
L. operculata extract showed promising antibacterial activity in vitro against the studied agents.
A Luffa operculata é, provavelmente, o fitoterápico mais utilizado no tratamento das rinites e rinossinusites. Apesar de amplamente utilizada pela população, seus mecanismos de ação ainda não estão completamente estabelecidos.
Avaliar a atividade antimicrobiana in vitro da Luffa operculata em agentes causadores de infecções de vias aéreas superiores: Staphylococcus aureus, Streptococcus pneumoniae e Streptococcus pyogenes.
Foram utilizadas diferentes concentrações de extrato alcoólico de Luffa operculata em caldo de bactérias dos agentes avaliados. Após incubação de 24 horas foi realizada a leitura de turvação do meio, e posteriormente, semeadura em placas de ágar-sangue e ágar Muller-Hinton, após 24 e 48 horas de incubação. Foram considerados testes negativos aqueles em que não houve a turvação do meio, confirmados pela ausência do crescimento das bactérias nas semeaduras. Foram considerados positivos os testes que apresentaram turvação do caldo ou positividade nas semeaduras de 24 ou 48 horas. Os resultados foram submetidos à análise estatística pertinente.
Os extratos de Luffa operculata apresentaram atividade antimicrobiana, especialmente para Streptococcus pyogenes, seguido dos Streptococcus pneumoniae e Sthaphylococcus aureus.
O extrato de Luffa operculata apresentou promissora atividade antimicrobiana in vitro contra os agentes estudados.
We describe a high throughput method for screening up to 1728 distinct chemicals with protein crystals on a single microplate. Acoustic droplet ejection (ADE) was used to co-position 2.5nL of ...protein, precipitant, and chemicals on a MiTeGen in situ-1 crystallization plate™ for screening by co-crystallization or soaking. ADE-transferred droplets follow a precise trajectory which allows all components to be transferred through small apertures in the microplate lid. The apertures were large enough for 2.5nL droplets to pass through them, but small enough so that they did not disrupt the internal environment created by the mother liquor. Using this system, thermolysin and trypsin crystals were efficiently screened for binding to a heavy-metal mini-library. Fluorescence and X-ray diffraction were used to confirm that each chemical in the heavy-metal library was correctly paired with the intended protein crystal. A fragment mini-library was screened to observe two known lysozyme ligands using both co-crystallization and soaking. A similar approach was used to identify multiple, novel thaumatin binding sites for ascorbic acid. This technology pushes towards a faster, automated, and more flexible strategy for high throughput screening of chemical libraries (such as fragment libraries) using as little as 2.5nL of each component.
Acoustic droplet ejection (ADE) is a powerful technology that supports crystallographic applications such as growing, improving and manipulating protein crystals. A fragment‐screening strategy is ...described that uses ADE to co‐crystallize proteins with fragment libraries directly on MiTeGen MicroMeshes. Co‐crystallization trials can be prepared rapidly and economically. The high speed of specimen preparation and the low consumption of fragment and protein allow the use of individual rather than pooled fragments. The Echo 550 liquid‐handling instrument (Labcyte Inc., Sunnyvale, California, USA) generates droplets with accurate trajectories, which allows multiple co‐crystallization experiments to be discretely positioned on a single data‐collection micromesh. This accuracy also allows all components to be transferred through small apertures. Consequently, the crystallization tray is in equilibrium with the reservoir before, during and after the transfer of protein, precipitant and fragment to the micromesh on which crystallization will occur. This strict control of the specimen environment means that the crystallography experiments remain identical as the working volumes are decreased from the few microlitres level to the few nanolitres level. Using this system, lysozyme, thermolysin, trypsin and stachydrine demethylase crystals were co‐crystallized with a small 33‐compound mini‐library to search for fragment hits. This technology pushes towards a much faster, more automated and more flexible strategy for structure‐based drug discovery using as little as 2.5 nl of each major component.
Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this ...technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal‐improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s−1) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from the inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl‐shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high‐throughput screening.
Improvements needed for automated crystallography include crystal detection and crystal harvesting. A technique that uses acoustic droplet ejection to harvest crystals was previously reported. Here a ...method is described for using the same acoustic instrument to detect protein crystals and to monitor crystal growth. Acoustic pulses were used to monitor the progress of crystallization trials and to detect the presence and location of protein crystals. Crystals were detected, and crystallization was monitored in aqueous solutions and in lipidic cubic phase. Using a commercially available acoustic instrument, crystals measuring ~150 µm or larger were readily detected. Simple laboratory techniques were used to increase the sensitivity to 50 µm by suspending the crystals away from the plastic surface of the crystallization plate. This increased the sensitivity by separating the strong signal generated by the plate bottom that can mask the signal from small protein crystals. It is possible to further boost the acoustic reflection from small crystals by reducing the wavelength of the incident sound pulse, but our current instrumentation does not allow this option. In the future, commercially available sound-emitting transducers with a characteristic frequency near 300 MHz should detect and monitor the growth of individual 3 µm crystals.
Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this ...technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal-improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s-1) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from the inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl-shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high-throughput screening.
An acoustic high-throughput screening method is described for harvesting protein crystals and combining the protein crystals with chemicals such as a fragment library. Acoustic droplet ejection (ADE) ...is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal-improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s{sup −1}) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from the inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl-shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high-throughput screening.
A method is presented for screening fragment libraries using acoustic droplet ejection to co-crystallize proteins and chemicals directly on micromeshes with as little as 2.5 nl of each component. ...This method was used to identify previously unreported fragments that bind to lysozyme, thermolysin, and trypsin. Acoustic droplet ejection (ADE) is a powerful technology that supports crystallographic applications such as growing, improving and manipulating protein crystals. A fragment-screening strategy is described that uses ADE to co-crystallize proteins with fragment libraries directly on MiTeGen MicroMeshes. Co-crystallization trials can be prepared rapidly and economically. The high speed of specimen preparation and the low consumption of fragment and protein allow the use of individual rather than pooled fragments. The Echo 550 liquid-handling instrument (Labcyte Inc., Sunnyvale, California, USA) generates droplets with accurate trajectories, which allows multiple co-crystallization experiments to be discretely positioned on a single data-collection micromesh. This accuracy also allows all components to be transferred through small apertures. Consequently, the crystallization tray is in equilibrium with the reservoir before, during and after the transfer of protein, precipitant and fragment to the micromesh on which crystallization will occur. This strict control of the specimen environment means that the crystallography experiments remain identical as the working volumes are decreased from the few microlitres level to the few nanolitres level. Using this system, lysozyme, thermolysin, trypsin and stachydrine demethylase crystals were co-crystallized with a small 33-compound mini-library to search for fragment hits. This technology pushes towards a much faster, more automated and more flexible strategy for structure-based drug discovery using as little as 2.5 nl of each major component.
The first herpes virus to be described was types 1 and 2, whose denomination is herpes simplex 1 and 2 or HSV -1 and HSV -2. These viruses have specific biological characteristics, such as the ...ability to cause different kinds of diseases, as well as to establish host's latent or persistent lifetime infections and also of being reactivated, causing lesions that can be located at the same site of the initial primary infection or close to it. It is suggested that this virus reactivation in the geniculate ganglion may be related to Bell's palsy. In this situation, the viruses that would be latent in this ganglion, would suffer reactivation and replication, then be diffused through the facial nerve and its branches, among them the chorda tympani nerve, which by stimulating salivary secretion would enable the identification of the viral DNA in the patients’ saliva. Until recently, a great number of patients was diagnosed as holders of this kind of paralysis, named idiopathic or Bell's palsy. With the introduction of the technique studying the viral DNA by Polymerase Chain Reaction (PCR), several authors have found herpes simplex virus type I DNA in the cerebrospinal fluid, in the lachrymal secretion, in the saliva and in the geniculate ganglia of patients with Bell's palsy.
observe the occurrence of herpes simplex type I virus using PCR technique in the saliva of patients with Bell's palsy and relating it to the clinical evolution of these cases.
We evaluated 38 patients with Bell's palsy submitted to anamnesis, clinical and ENT examination and saliva sampling for viral DNA detection by PCR technique. The control group was ten normal adults.
We found positive viral DNA in 11 cases out of the 38, which corresponded to 29% of the sample. This result was statistically significant if compared to the control group, in which we did not find any positive case.
The end result was that the presence of HSV -1 in the saliva of patients with Bell's palsy indicating that the viral reactivation can be the etiology of this disease. The detection of the virus in these patients’ saliva does not influence the disease prognosis.
Os primeiros herpes-vírus a serem descritos foram os tipos 1 e 2, cuja denominação é herpes simplex 1 e 2 ou HSV-1 e HSV-2. Estes vírus possuem características biológicas particulares, tais como a ...capacidade de causar diferentes tipos de doenças, assim como estabelecer infecções latentes ou persistentes por toda a vida dos hospedeiros e de serem reativados causando lesões que podem se localizar no sítio da infecção primária inicial ou próxima a ele. Postula-se que a reativação deste vírus no gânglio geniculado esteja relacionada com a paralisia de Bell. Nesta situação, os vírus, que estariam latentes neste gânglio, sofreriam reativação e replicação difundindo-se pelo nervo facial e seus ramos, dentre eles o nervo corda do tímpano, que ao estimular a secreção salivar possibilitaria a identificação do DNA viral na saliva dos pacientes. Até recentemente, um grande número de pacientes eram diagnosticados como portadores de uma forma desta paralisia, chamada de idiopática ou de paralisia de Bell. Com o advento da técnica de estudo do DNA viral pelo método da reação da polimerase em cadeia (PCR), diversos autores encontraram DNA do vírus herpes simplex tipo I no líquido cefalorraquidiano, na secreção lacrimal, na saliva e nos gânglios geniculados de pacientes com paralisia de Bell. OBJETIVO: observar a prevalência do vírus herpes simplex tipo I pela técnica de PCR, na saliva de pacientes com PFP de Bell, relacionando-a com a evolução clínica destes casos. METODOLOGIA: Avaliamos 38 pacientes portadores de Paralisia Facial Periférica de Bell, que foram submetidos a anamnese, exame médico geral e otorrinolaringológico e coleta de saliva para detecção do DNA viral pela técnica de PCR. O grupo controle correspondeu a 10 adultos normais. RESULTADOS: Obtivemos positividade para o DNA viral em 11 casos dos 38 avaliados, o que corresponde a 29% da amostra. Este resultado foi estatisticamente significante se comparado ao grupo controle, no qual não foi obtido nenhum caso de positividade. CONCLUSÃO: Concluiu-se que a presença do HSV-1 na saliva de pacientes portadores de PFP de Bell indica que a reativação viral pode ser a etiologia desta doença. A detecção do vírus na saliva destes pacientes não influencia o prognóstico da doença.
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