Methods for in vitro evaluating antimicrobial activity： A review

In recent years, there has been a growing interest in researching and developing new antimicrobial agents from various sources to combat microbial resistance. Therefore, a greater attention has been paid to antimicrobial activity screening and evaluating methods. Several bioassays such as disk-diffusion, well diffusion and broth or agar dilution are well known and commonly used, but others such as flow cy- tofluorometric and bioluminescent methods are not widely used because they require specified equip- ment and further evaluation for reproducibility and standardization, even if they can provide rapid re- sults of the antimicrobial agent＇s effects and a better understanding of their impact on the viability and cell damage inflicted to the tested microorganism. In this review article, an exhaustive list of in vitro antimicrobial susceptibility testing methods and detailed information on their advantages and limita- tions are reported.

Development of Topical Antimicrobial Gel Containing Beehive Extract

This study aimed to determine the antimicrobial activity of beehive extract and developed topical antimicrobial gel. The beehives were extracted with three different solvents： distilled water, 50% ethanol and 95% ethanol, then each filtrate was evaporated by vacuum rotary evaporator to obtain crude extracts： water extract （W）, water-ethanol extract （WE） and ethanol extract （E）. These extracts were tested for their antimicrobial activity against five bacterial strains： Escherichia coli （ATCC 25922）, Psuedomonas aeuruginosa （ATCC 27853）, Staphylococcus aureus （ATCC 25923）, Methicillin Resistance Staphylococcus aureus （MRSA） and β-hemolytic Streptococcus group A （GAS）, then they were evaluated for their MIC and MBC values. The results revealed that the yield of the W, WE and E extracts amounted to 16.25%, 14.42% and 6.67%, respectively. Whereas, all of the samples were found to be effective against P. aeuruginosa, S. aureus, MRSA and GAS, but they were ineffective against E. coli. The W extract exhibited the highest antimicrobial activity with MIC values of 31.25-62.50 mg/mL and MBC values of 62.50 mg/mL. Then the gel bases using different gelling agents such as carbopol, hydroxyethylcellulose （HEC） and hydroxypropyl methylcellulose （HPMC） were formulated and evaluated for their physical properties： appearance, texture, pH, spread ability and stability. The formula which contained HEC was selected to incorporate with the W extract due to its smooth texture, good spreadability on the skin and good stability. The W gel exhibited a promising antimicrobial activity in both before and after stability test and also expressed no skin irritation.

Antimicrobial and cytotoxic potential of seeds and flowers crude extracts of sunflower

Oil-producing crops,including Helianthus annuus L.(H.annuus,Sunflower),have been used for the treatment of different human diseases since ancient times.This study aims to determine the antimicrobial and cytotoxic potential of different polarities of seeds and flower extracts of locally grown H.annuus by well-established bioassays.To achieve the objects,the plant extracts were prepared from both flowers and seeds powder samples individually with methanol solvent by using a Soxhlet extraction method.The antimicrobial and cytotoxic potential of the flowers and seed extracts were determined by the in vitro diffusion and brine shrimp method(BSL).The results of the antimicrobial potential of both prepared flower and seed extracts of various polarities at different concentrations showed promising potential against the gram-positive and gram-negative bacterial strains,within the range of inhibition zone 0–20 mm.Among the flower extracts,the highest potential was obtained in water extract and the lowest potential was in butanol extract followed by water>ethyl acetate>hexane>methanol>chloroform>butanol extracts.However,from the seed extracts,the highest potential was obtained in water extract and the lowest potential was in the methanol extract followed by water>hexane>butanol>ethyl acetate>chloroform>methanol extract.On the other hand,the results of cytotoxicity among the flowers extracts against the brine shrimp method where the highest LC50 against brine shrimp nauplii were found to be 31.25μg/mL in water extract and the lowest LC50 was 1345.86μg/mL in chloroform extract followed by water>ethyl acetate>hexane>butanol>methanol>chloroform extract.Similarly,different polarities of seed extracts,the highest LC50 against brine shrimp nauplii was shown to be 109.56μg/mL in butanol extract and the lowest LC50 was 1513.56μg/mL in ethyl acetate extract followed by butanol>hexane>water>methanol>chloroform>ethyl acetate extract.In conclusion,the results showed that both flowers and seeds extracts at various concentrations have promising potential against the positive and negative bacterial strains and the highest potential water extract could be used as natural broad-spectrum antibiotics as herbal medicine to treat different human infectious diseases.

Antitumor and antimicrobial activities of endophytic fungi from medicinal parts of Aquilaria sinensis

The purpose of this study was to isolate and characterize endophytic fungi from the stem tissue which can produce fragrant ingredients in Aquilaria sinensis(also called agarwood) to determine their antitumor and antimicrobial activities.Twenty-eight fungal endophytes were isolated from agarwood by strict sterile sample preparation and were classified into 14 genera and 4 taxonomic classes(Sordariomycetes,Dothideomycetes,Saccharomycetes,and Zy-gomycetes) based on molecular identification.Of the 28 isolates,13(46.4%) showed antimicrobial activity against at least one of the test strains by the agar well diffusion method,and 23 isolates(82.1%) displayed antitumor activity against at least one of five cancer cell lines by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) assay.The diameters of inhibition zones of YNAS07,YNAS14,HNAS04,HNAS05,HNAS08,and HNAS11 were equal to or higher than 14.0 mm against Staphylococcus aureus,Escherichia coli,Bacillus subtilis,B.subtilis,Aspergillus fumigatus,and B.subtilis,respectively.The inhibition rates of YNAS06,YNAS08,and HNAS06 were not less than 60% to 293-T,293-T,and SKVO3 cells,respectively.These results suggest that the endophytic fungi associated with agarwood will provide us with not only useful micro-ecological information,but also potential antimicrobial and anti-tumor agents.