INTRODUCTION

Infectious diseases caused by microorganisms have been one of the most important causes of death in humanity (^{Lozano et al., 2012}). Bacterial agents including Staphylococcus aureus, Escherichia coli, Pseudonomas aeruginosa, Bacillus subtilis and Proteus vulgaris have caused important infectious diseases within public health (^{Khan et al., 2013}).

The introduction of antimicrobial agents in medicine has been one of the most important interventions to control and reduce the prevalence of infectious diseases (^{Alós, 2015}); However, a growing threat in recent years that has diminished the effectiveness of these drugs is bacterial resistance to antibiotics; generated because the microorganisms have acquired the ability to prevent an antimicrobial from acting against it. As a result, the treatments of choice become ineffective, infections persist and can spread to other individuals (^{WHO, 2017}).

In most populations of different developing countries, humanity has used plants to treat common infectious diseases, which could be a potential alternative to produce new drugs of great health benefit (^{Renisheya et al., 2011}; ^{Khan et al., 2013}).

One of the plants considered important for the study of its photochemical properties is Salix babylonica, commonly known as weeping willow. This species belongs to the Salix genus of the Salicaceae family, Salix babylonica is one of the best known species in willows, distributed in some areas of Asia, Europe and America; commonly used as an ornamental and medicinal plant (^{Wahab et al., 2018}).

There are reports in which the pharmacological properties associated with the evaluation of leaf, bark and stem extracts are evidenced; obtained from Salix babylonica. Among the phytochemical properties attributable to Salix babylonica, there are: anthelmintic, antiseptic, antiarthritic, astringent, analgesic, anticancer, antipyretic, antimalarial, antioxidant, antifungal, anthelmintic and antibacterial activity; these properties are associated with its content of secondary compounds such as total phenolics, flavonoids, terpenes and lignans (^{Sulaiman et al., 2013}; ^{Wahab et al., 2018}).

Based on the aforementioned approaches, the objective of the present research work was to characterize and determine the antibacterial activity of the methanolic extract of Salix babylonica, on bacteria of importance in public health.

MATERIAL AND METHODS

RESULTS

The qualitative characterization carried out on the methanolic extract of Salix babylonica, indicates the presence of unsaturations, phenolic oxidriles, coumarins, lactones, flavonols, quinones, saponins, aromaticity and polyphenols; In addition to being positive to the Lieberman-Buchard test, which indicates the presence of triterpenes and steroidal compounds. (Table 2).

DISCUSSION

The presence of unsaturations, phenolic oxydriles, coumarins, lactones, flavonols, quinones, fluorataninos, triterpene steroids and saponins in the metonolic extract of Salix babylonica was determined by means of qualitative tests. In previous studies, compounds such as tritetracontane, 1,2,3-propanetriol ester, octadecanoic acid, methyl ester of hexadecanoic acid and 1,3-dioxane-4- (hexadecyl oxy) -2-pentadecyl have been identified; most of them classified as phenolic compounds, in addition to 7-O-β-D-glucopyranoside of luteolin, luteolin and crisoeriol; compounds classified as flavonoids (^{Salem et al., 2011}).

Biological activities such as anticancer, antiulcer, antimalarial, antidiarrheal, antifungal, antitussive, anti-inflammatory, anthelmintic and antibacterial have been reported; in studies conducted with phenolic compounds, alkaloids, glycosides and terpenes (^{Hernández-Alvarado et al., 2018}).

On the other hand, gas chromatography allowed to identify Timol and Carvacrol at concentrations of 0.5319 mg/ml, 0.4158 mg/ml respectively. These compounds are classified as essential oils of volatile nature, with some biological activities reported as: expectorant, antifungal, anti-inflammatory, analgesic, antiseptic, antioxidant, anti-rheumatic, antispasmodic, anti-hepatotoxic and antibacterial; both against Gram positive and Gram negative bacteria (^{Magi et al., 2015}).

When carrying out the antibacterial evaluation of the methanolic extract of Salix babylonica, it was determined that the extract exhibits better activity against Gram positive bacteria (Staphylococcus aureus, Listeria monocytogenes and Bacillus subtilis); which against Gram negative bacteria (Escherichia coli, Salmonella typhi, Salmonella cholerasuis, Pseudomonas aeruginosa). This effect is based on the structure of Gram negative bacteria, which by having a phospholipid membrane that prevents the cell wall from being penetrated by lipophilic solutes; while the porins constitute a selective barrier for hydrophilic solutes, so the bacteria are protected from being penetrated by compounds such as antibiotics or some secondary metabolites derived from plants (^{Kaye et al., 2004}; ^{Ndhlala et al., 2015}).

For the determination of antibacterial activity, it is important to determine Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC); defined as MIC at the lowest concentration of antimicrobial agent that inhibits the growth of the microorganism, detected visually (^{CLSI, 2012}). In the present experiment to determine the end point of the WCC, a colorimetric method was used, based on the use of tetrazolium salts (^{Balouiri et al., 2016}); which allows to observe a color change from yellow to pink, caused by the entry of this salt into the cell, which is reduced by the NAD (P)H-dependent oxidoreductases and the dehydrogenases of metabolically active cells, producing the change from color to pink (^{Berridge et al., 2005}).

^{Sulaiman et al., in 2013}, carried out a study in which they evaluated the antimicrobial activity of the ethanol extract of Sálix alba bark, belonging to the genus sálix and family salicaeae; same as Sálix babylonica. In this study they determined that Sálix alba has better antibacterial activity against Staphylococcus aureus; medium activity against Pseudomonas aeruginosa and had no effect against Escherichia coli and Klebsiella pneumoniae. The concentrations evaluated were 10, 20, 40, 60 and 80 mg/ml, using the agar diffusion technique; observing the greatest halos of inhibition at 80 mg/ml. The results of this study correspond to those observed in the present experiment, since the extract had a better effect against Gram positive bacteria (Staphylococcus aureus, 25 mg/ml; Listeria monocytogenes, 25 mg/ml and Bacillus subtilis, 12.5 mg/ml ), which against Gram negative bacteria (Escherichia coli, Salmonella typhi, Salmonella cholerasuis and Pseudomonas aeruginosa, 100mg/ml, for each one); however, the extract was obtained from the bark of Sálix alba. The concentrations change and the technique also, so the results are not 100 % comparable, although the trees belong to the same genus and family.

On the other hand, in a study conducted by ^{Wahab and colaboradores en 2018}, who evaluated the methanolic extracts of the leaves and bark of Sálix babylonica; in addition to its fractions of petroleum ether, methylene chloride and ethyl acetate (diluted in dimethylsulfoxide), to determine its antimicrobial activity against Gram positive (Staphylococcus aureus) and Gram negative (Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa) bacteria, using the agar diffusion technique and the extract at a concentration of 100 µg. The results showed that both the methanolic extract of the leaves and the bark have moderate or weak antimicrobial activity against the challenged microorganisms; observing the greatest halos of inhibition (10 mm) with Pseudomonas aeruginosa, followed by Klebsiella pneumoniae (9 mm), finally Escherichia coli, Staphylococcus aureus (8 mm).

Because in the mentioned studies only techniques are applied to determine the sensitivity of the microorganism to a certain compound by the agar diffusion method; It is not possible to compare the results with those obtained in the present experiment, since in this case the minimum inhibitory concentration was determined by the plate microdilution method; In addition to the study by Wahab et al., dimethyl sulfoxide was used to dilute extracts and fractions; compound that is used to increase the permeability of the bacterial membrane, increasing the activity of the compounds and reducing the concentrations of use (^{Borges et al., 2013}; ^{Sulaiman et al., 2013}; ^{Wahab et al., 2018}).

The Minimum Inhibitory Concentration is defined as the lowest concentration of antimicrobial agent necessary to kill 99.9% of the final inoculum, after incubation for 24 hours under a standardized set of conditions described by the CLSI (^{Balouiri et al., 2016}). The determination of the MIC is not a viable option to know 100 % the efficacy of a drug or compound, since within each well there may still be viable cells if the drug evaluated only had a bacteriostatic effect on the bacterial species under study (^{Wiegand et al., 2008}).

In the present experiment, the MBCs of the methanolic extract were determined against Escherichia coli, Salmonella typhi, Salmonella cholerasuis and Pseudomonas aeruginosa (200 mg/ml); Listeria monocytogenes and Staphylococcus aureus (50 mg/ml) and Bacillus subtillis (25 mg/ml); however, there are no studies reported with Sálix babylonica or another species of the genus Sálix, with which this activity has been reported.

CONCLUSION

In the present study it was demonstrated that the methanolic extract of Sálix babylonica has potential antibacterial activity on some bacterial pathogens of importance in public health; being an alternative for the treatment of diseases caused by bacteria resistant or multi-resistant to antibiotics.