|Year : 2019 | Volume
| Issue : 1 | Page : 1-7
Chemical characterization and evaluation of antioxidant and antimicrobial activities of Litchi chinensis sonn
Marisa De Oliveira Lopes1, Ana Flávia Da Silva1, Cláudio Daniel Cerdeira2, Ingridy Simone Ribeiro3, Isael Aparecido Rosa1, Luis Felipe Cunha Dos Reis1, Marcelo Aparecido Da Silva1, Marcos José Marques3, Jorge Kleber Chavasco2, Geraldo Alves Da Silva1
1 Department of Food and Drugs, Faculty of Pharmaceutical Sciences, Federal University of Alfenas, Alfenas, Minas Gerais, Brazil
2 Department of Microbiology and Immunology, Biomedical Science Institute, Federal University of Alfenas, Alfenas, Minas Gerais, Brazil
3 Department of Molecular Biology, Biomedical Science Institute, Federal University of Alfenas, Alfenas, Minas Gerais, Brazil
|Date of Web Publication||20-Feb-2019|
Ms. Luis Felipe Cunha Dos Reis
Department of Food and Drugs, Faculty of Pharmaceutical Sciences, Federal University of Alfenas, Alfenas, Minas Gerais
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background: Litchi chinensis is used in traditional Chinese medicine and by Indian medical system. Ethnopharmacological studies show anti-inflammatory, antidiabetic and analgesic activities, among others. However, there are few studies of antimicrobial activity. This study evaluates antimicrobial, antioxidant, and cytotoxicity properties of the lychee's leaves extract (LE) and fractions. Materials and Methods: Extracts were obtained using an exhaustive extraction method with ethanol: Water (7:3 v/v). Subsequently, LE was concentrated in a rotary evaporator. Finally, LE was dried via lyophilization. Fractions were obtained via the partition process. Bioactivity of the LE and fractions (hexane [Hex], ethyl acetate [EtOAc], n-butanol [BuOH], and aqueous [Aq]) from L. chinensis was evaluated through antimicrobial activity using broth microdilution, antioxidant activity via both 1,1-diphenyl-2-picryl-hidrazila assay and ferric reducing capacity and cytotoxicity through 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction assay. Furthermore, mass spectrometry technique electrospray ionization ion trap mass spectrometry was used to identify the chemical composition of the LE and fractions. Results: Phenolic compounds, such as flavonoids and condensed tannins were the main substances found. Total phenolic and flavonoid contents were higher in EtOAc (541.15 ± 2.4 mg/g and 31.06 ± 0.5 mg/g, respectively). This fraction showed the best results for antioxidant activity (IC50= 3.45 mg/mL) and ferric reducing capacity (20.27% ± 0.11). The LE and fractions showed considerable antimicrobial activity, chiefly against Bacillus subtilis, Bacillus cereus, Staphylococcus aureus, and Proteus mirabilis, with the minimum inhibitory concentration ranging from 50 to 1560 μg/ml. Conclusion: This study revealed that L. chinensis is a source of bioactive compounds potentially useful for pharmaceutical and food industries.
Abbreviations Used: LE: Leaves extract, Hex: Hexane fraction, EtOAc: Ethyl acetate fraction, BuOH: n-butanol fraction, Aq: Aqueous fraction, DPPH: 1,1-diphenyl-2-picryl-hidrazila, MTT: 3-(4,5-dimethyl-thiazol-2-yl)-2, 5-diphenyltetrazolium bromide, ESI-IT-MSn: Electrospray ionization ion trap mass spectrometry, IC50: Median Inhibition Concentration (concentration that reduces the effect by 50%), MIC: Minimum inhibitory concentration, MS/MS: Mass spectrometry, MSn: Tandem mass spectrometry, GAE: Equivalents of gallic acid, QE: Equivalents of quercetin, BHT: Butylated Hydroxy Toluene, UV: Ultraviolet, ATCC: American Type Culture Collection, MHB: Mueller Hinton broth, DMSO: Dimethyl sulfoxide, ANOVA: Analysis of variance, CC50: Cytotoxic concentration 50.
Keywords: Antimicrobial activity, antioxidant activity, medicinal plants, phenolic compounds, phytochemical screening
|How to cite this article:|
Lopes MD, Da Silva AF, Cerdeira CD, Ribeiro IS, Rosa IA, Reis LF, Da Silva MA, Marques MJ, Chavasco JK, Da Silva GA. Chemical characterization and evaluation of antioxidant and antimicrobial activities of Litchi chinensis sonn. Phcog Res 2019;11:1-7
|How to cite this URL:|
Lopes MD, Da Silva AF, Cerdeira CD, Ribeiro IS, Rosa IA, Reis LF, Da Silva MA, Marques MJ, Chavasco JK, Da Silva GA. Chemical characterization and evaluation of antioxidant and antimicrobial activities of Litchi chinensis sonn. Phcog Res [serial online] 2019 [cited 2020 Sep 28];11:1-7. Available from: http://www.phcogres.com/text.asp?2019/11/1/1/252557
- The greatest values of the total phenolics content and total flavonoid contents were found on ethyl acetate fraction (EtOAc) (541.15 ± 2.4 mg/g sample and 31.06 ± 0.5 mg EQ/g sample, respectively)
- It was observed that 1,1-diphenyl-2-picryl-hidrazila radical scavenging capacity of the EtOAc was as efficient or higher as compared with ascorbic acid, butylated hydroxy toluene , and quercetin standards. The ferric reducing capacity of extracts and fractions indicated that the antioxidant activity of EtOAc was greater when compared with the others fractions
- EtOAc fraction presented the most significant results demonstrated the lowest MIC values (50 μg/mL) against Bacillus subtilis, Bacillus cereus, and Staphylococcus aureus. The leaves extract (LE) was also significantly active against Gram-positive bacteria B. cereus and S. aureus and against Gram-negative bacteria Proteus mirabilis (50 μg/mL)
- The LE of Litchi chinensis and its fractions did not show cytotoxicity at the highest concentration used (160 μg/mL) on murine peritoneal macrophages by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide method.
| Introduction|| |
Litchi chinensis Sonn. is a native species from southern China that belongs to the Sapindaceae family. Its fruits are very popular in South and Southeast Asia. The exotic culture of this species occurs in the tropical and sub-tropical regions, also occurring in North and South America.
L. chinensis is used in traditional Chinese medicine  and is also used by the medical system of the Indian subcontinent, where ethnopharmacological studies have shown that the most popular uses found for litchi are as anti-inflammatory, antidiabetic, anti-obesity, analgesic, antitussive, and diuretic, as well as to treat neuralgic disorders, scurvy, orchitis, and sore throats.,,, The tea from the bark is used to treat diarrhea, and the seed is ground into a powder to be used for intestinal problems. Pharmacological studies have demonstrated its strong antioxidant activity ,,, and anticancer ,,, effects, mainly due to the presence of phenolic compounds and flavonoids.,,,,
The phenolic compounds present in fruits and vegetables have important bioactive properties and are well-known for their health benefits. These compounds are commonly found in parts of both edible and inedible plants and have been associated with multiple biological effects including antioxidant, antibacterial, antiviral, anti-inflammatory, anti-thrombotic, and vasodilatory activities.
Multi-drug resistance of micro-organism has led to serious public health problems, resulting in high rates of morbidity and mortality from infectious diseases.,,, Thus, it is extremely important the discovery of new compounds from natural sources that may support new therapeutic options. In this context, plants play a determinant role in the search for compounds with antimicrobial activity.,,
In this study, the in vitro antimicrobial, antioxidant, and cytotoxic activities of the leaves and fractions from L. chinensis were investigated. In addition, the quantification of the total phenolics and total flavonoids contents and the identification of the chemical constituents by mass spectrometry electrospray ionization ion trap mass spectrometry (ESI-IT-MS n) were carried out.
| Materials and Methods|| |
The plant material from L. chinensis Sonn. was collected in the city of Alfenas-MG (geographical data: Altitude of 808.0 m, latitude 21°20'27,3”S and longitude 45°53'29,2”). The plant was identified in the botany laboratory of the Federal University of Alfenas and a reference specimen (n° 2276) was deposited in the herbarium at UNIFAL-MG.
Preparation of hydroethanolic extract and fractions
The leaves were dried at 45°C and were then pulverized. The extracts were obtained using an exhaustive extraction method with a liquid mixture of ethanol: Water (7:3 v/v). Subsequently, leaves extract (LE) was concentrated in a rotary evaporator. Finally, the LE was dried via lyophilization. Fractions were obtained via a liquid–liquid partition process, using as solvents hexane (Hex), ethyl acetate (EtOAc) and n-butanol (BuOH) yielding the fractions Hex, EtOAc, and BuOH. The residual was named as an aqueous fraction (Aq).
Analysis of extract and fractions by mass spectrometry
The analysis of the extract and fractions was performed using a tandem mass spectrometry method (MS/MS or MS n) by direct insertion into the ESI-IT-MS n system. The samples were submitted to an electrospray ionization mode (ESI) and fragmentation was performed in an ion trap-type interface (IT). Negative mode was selected for generation of the first-order MS and MS n. The analysis range was between 50–2000 m/z. Mass spectra were obtained on a Thermo Scientific Linear 2D LTQ XL mass spectrometer. The Xcalibur software version 2.2 (Thermo Scientific®) was used for the acquisition and processing of spectral data.
The content of phenolic compounds in the extract and fractions was determined based on the Folin-Ciocalteau colorimetric method  using equivalents of gallic acid (GAE) as a standard. The absorbance of the samples was measured at 750 nm and the results were expressed in mg equivalents of GAE per gram of extract.
Determination of flavonoids was performed using the colorimetric spectrometric method of chelation with aluminum chloride in ultraviolet  using quercetin as the standard. Absorbance of the samples was measured at 425 nm corresponding to the absorption peak of quercetin aluminum chelate, and the results were expressed in mg equivalents of quercetin (QE) per gram of extract.
Evaluation of antioxidant activity
1,1-diphenyl-2-picryl-hidrazila radical scavenging capacity
Evaluation of scavenging activity of 1,1-diphenyl-2-picryl-hidrazila (DPPH) was performed according to the methodology previously described, using solutions of the samples and standards (quercetin, ascorbic acid and butylated hydroxy toluene [BHT]), which were prepared at concentrations between 12.5 μg/ml and 400 μg/ml. An aliquot of 0.5 ml of DPPH solution (0.5 mM) was added in 0.3 ml of samples and standards and was then diluted in 3 ml of ethanol. The mixture was stirred vigorously and was kept away from light exposure for 30 min. The readings were made at 517 nm. The IC50 value was determined for each sample. IC50 value is the concentration of the sample required to scavenge 50% of the free radicals present in the system. All experiments were performed in triplicate.
Ferric reducing capacity
The reduction capacity of the samples was calculated as previously described. Samples and standards (ascorbic acid and BHT) were prepared in triplicate at concentrations between 12.5 μg/ml and 400 μg/ml. An aliquot of 1.0 ml was added to a solution of 2.5 ml a phosphate buffer 0.2 M (pH 6.6) and 2.5 mL of K3[Fe (CN)6] (1%) to each tube. The mixture was kept at 50°C for 30 min. Subsequently, 2.5 mL of trichloroacetic acid (10%) was added, and the mixture was stirred. Finally, 2.5 mL of the mixture was transferred to a solution of 2.5 ml of distilled water and 0.5 mL FeCl3 (0.1%). The readings were done at 700 nm, and the results were expressed as a percentage of Fe 2+ chelating activity.
In vitro evaluation of antimicrobial activity
The strains of micro-organism used were from the American type culture collection (ATCC) and Microbiology and Immunology Laboratory (LMI) at UNIFAL-MG. These micro-organism are representative of the main groups of mycobacteria, bacteria, and fungi and demonstrate medical and environmental importance. The fungi included Candida albicans (ATCC 10231) and Saccharomyces cerevisiae (ATCC 2601). The Gram-positive bacteria included Bacillus subtilis (ATCC 6633), Bacillus cereus (ATCC 11778), Micrococcus luteus (ATCC 9341), Enterococcus faecalis (ATCC 51299), and Staphylococcus aureus (ATCC 6538). The Gram-negative bacteria included Escherichia More Details coli (ATCC 25922), Serratia marcescens (LMI-UNIFAL), Pseudomonas aeruginosa (ATCC 27853), Proteus mirabilis (ATCC 25922), Salmonella More Details typhimurium (ATCC 14028), and Enterobacter aerogenes (LMI-UNIFAL). The mycobacteria included Mycobacterium bovis (BCG strain, ATCC 27289) and Mycobacterium tuberculosis (H37Ra strain, ATCC 27294).
Broth microdilution method
The minimum inhibitory concentration (MIC) of L. chinensis extract and fractions was determined through broth microdilution against bacteria and fungi, according to methodologies established in the M7A6 (Clinical and Laboratory Standards Institute [CLSI], 2003) and M27A3 (CLSI, 2008) documents. Tests were performed on 96-well microplates. At first, the turbidity of microbial suspensions in sodium chloride 0.9%, cultured overnight at 35°C for 18 h, was adjusted according to a McFarland standard (0.5 tube). Next, 100 μl of Mueller Hinton broth (MHB) was added to the wells and after that, 100 μl of the LE and each fraction tested were added. Serial dilutions were made with the final concentrations of the LE or fractions ranging between 25 μg/mL to 1,250 μg/mL. Finally, 10 μl of micro-organism was added to each well. The reading was performed visually as previously determined (CLSI, 2003), wherein the presence of turbidity in the wells after incubation for 24 h at 37°C was considered indicative of bacterial growth. The MIC99.9 was established as the lowest concentration of the extract or fraction in which no turbidity had occurred. The growth control was comprised 100 μl of MHB and 10 μl of inoculum. The extract control was comprised 100 μl of MHB and 100 μl of the LE or fraction and the sterility control contained only 100 μl of MHB. Chlorhexidine 0.12% was also used as a positive control.
The LE, at a concentration of 50,000 μg/mL, was evaluated against both the M. bovis and M. tuberculosis via an agar diffusion assay, according to the M24A2 (CLSI, 2008) document., Regarding controls, Rifampicin 30 μg was used as a positive control and distilled water was used as a negative control.
Quantitative evaluation of antimicrobial activity
The antimicrobial activity of plant extracts can be expressed in different ways. According to previous studies,, one can express antimicrobial effectiveness through numerical values beyond the MIC (μg/ml). In this study, two calculations were used to demonstrate the activity of the extracts tested: total activity and the values of the percentage activity (%). The use of the total activity values allows for comparing the bioactivity of different parts of the plant or between different plants on a more rational and standardized basis, allowing for the best interpretation of the results. These values would indicate the largest volume to which biologically active compounds in 1 g of plant material can be diluted and still inhibit microbial growth. The percentage of activity evaluates the antimicrobial potential of the tested L. chinensis extracts and represents the percentage of micro-organism that do not grow in the presence of these extracts. This index allows for a comparison between the statements that presented MIC of ≤100 μg/ml and determines the most active. These values were calculated as follows:
In vitro evaluation of cytotoxicity
Cytotoxicity was evaluated using the 3-(4,5-dimethyl-thiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) method. Murine peritoneal macrophages were used in RPMI 1640 medium and were kept at 37°C and 5% CO2. They were arranged in 24-well plates at the rate of 8 × 105 cells per well. The extract and fractions were added at concentrations ranging from 0.1 μg/ml to 160 μg/ml and were incubated for 72 h. After the incubation period, 50 μL of MTT was added to each well, with further incubation for 4 h. The cells were treated with dimethyl sulfoxide and evaluated at 570 nm to determine the cytotoxic concentration for 50% of the cells (CC50).
To evaluate the correlation between the amounts of total phenols and flavonoids with antioxidant and antimicrobial activities, the Spearman correlation coefficient (r) was employed using the BioEstat Software version 5.0. All of the results were submitted to analysis of variance followed by Tukey test (P < 0.05).
| Results and Discussion|| |
The mass spectra ESI-MS obtained from the LE [Figure 1] from 200 to 2000 m/z showed some major peaks, which were recognized as known phenolic compounds (catechin and quercetin), along with some tannins, which were confirmed via second order MS. The peaks of 863 and 1151 m/z corresponded to the polimeric condensed tannins known as procianidines, present in this species.,,, The MS 2 spectrum of the 447 m/z ion [Supplementary Figure 1 [Additional file 1]] showed a peak of 285 m/z, due to a loss of a 162 Da fragment of hexose, generating the aglycone form of catechin (C15H9O6). The ion 289 m/z represented the flavonoid epicatechin (C15H14O6), based on previous data., The MS 2 spectrum of the 609 m/z ion [Supplementary Figure 2 [Additional file 2]] showed a peak of 301 m/z, which represents a loss of two fragments from hexose and desoxi-hexose, with 162 Da and 146 Da, respectively, resulting in the aglycone form of quercetin (C15H10O7). The outstanding presence of the peaks of 289, 447, and 609 m/z in EtOAc indicates that this fraction exhibited a greater amount of phenolic compounds when compared with the other fractions [Supplementary Figure 3 [Additional file 3]].
|Figure 1: Mass spectrum, in full scan mode, obtained via direct injection of leaves extract of Litchi chinensis|
Click here to view
|Figure 2: DPPH radical scavenging activity of leaves extract and fractions of Litchi chinensis Sonn.|
Click here to view
In the present study, we found significant levels of total phenols in the extract and fractions of L. chinensis [Table 1]. The greatest value of total phenolics content was found on EtOAc (541.15 ± 2.4 mg/g sample). Similarly, the greatest value of total flavonoid contents was found in the EtOAc (31.06 ± 0.5 mg EQ/g sample). Similar studies indicate the presence of polyphenolics, flavonoids, condensed tannins, and proanthocyanidins in this species, especially in the fruit, flowers and seeds.,,,,,, This study confirmed the presence of these compounds in significant concentrations in the L. chinensis leaves, which have the advantage of being a renewable source and are available throughout the entire year. Phenolic compounds, especially flavonoids, have aroused great interest since recent discoveries have linked these compounds and their properties to the ability to scavenge oxidants, preventing chronic diseases such as cardiovascular disease, diabetes, kidney diseases, and cancer.,,,,
|Table 1: Antioxidant activity by 1,1-diphenyl-2-picryl-hidrazila method (median inhibition concentration) and ferric reducing capacity of leaves extract and fractions of Litchi chinensis Sonn., total phenolics content (values at milligram equivalent of gallic acid), and total flavonoids content (values at milligram equivalent of quercetin)|
Click here to view
The DPPH radical scavenging activity of LE and fractions of L. chinensis is shown in [Figure 2]. The antioxidant activity by DPPH method (IC50) of the extract and fractions showed the following profile: EtOAc > BuOH > Hex > Aq > LE [Table 1]. The highest antioxidant potential was shown on the EtOAc, which had the lowest concentration able to sequestering 50% of DPPH free radicals, with IC50= 3.45 ± 0.12 μg/mL. It was observed that DPPH radical scavenging capacity of the EtOAc was as efficient or higher as compared with ascorbic acid (6.49 ± 0.15 μg/mL), BHT (70.11 ± 0.10 μg/mL) and quercetin standards (4.57 ± 0.20 μg/mL). The antioxidant activity of EtOAc was greater than the observed by Yang et al., 2012 and Shahwar et al., 2010, in EtOAc fraction of flower acetone extract (16.73 ± 2.25 μg/mL) and in EtOAc fraction of stem-bark methanolic extract (15.30 μg/mL), respectively.
The ferric reducing capacity of extract and fractions at 100 μg/mL concentration [Table 1], indicated that the antioxidant activity of EtOAc was greater (20.27% ± 0.11) when compared against the LE and others fractions (Hex = 12.30% ± 0.08, BuOH = 10.80% ± 0.31, LE = 8.74% ± 0.03, Aq = 7.85% ± 0.15). The EtOAc result was higher when compared with BHT standard (11.50% ± 0.17). However, the EtOAc result was found to be lower than that of ascorbic acid standard (36.85% ± 0.14) at the same concentration.
Total phenolics content and total flavonoids were correlated with antioxidant activity, and the Spearman coefficients were calculated [Table 2]. The Spearman correlation coefficient is a measure of the degree of linear relationship between two quantitative variables. This coefficient varies between the values-1 and 1, wherein value of 0 (zero) means that there is no linear relationship, a value of 1 indicates perfect linear relationship and the value of-1 indicates a perfect inverse linear relationship. The closer the Spearman coefficient is to 1 or-1, the stronger the linear association between the two variables. Based on the Spearman coefficient, the results of total phenols and flavonoids indicated a positive correlation between these compounds and antioxidant activity. The results showed the highest correlation between the concentration of flavonoids with ferric reducing capacity (r = 0.9). The Spearman coefficient was 0.8 when correlated the DPPH scavenging capacity with flavonoids.
|Table 2: Spearman coefficients for the correlation between total phenolics content, total flavonoids and 1,1-diphenyl-2-picryl-hidrazila, ferric reducing capacity and minimum inhibitory concentration|
Click here to view
It is known that the number of phenolic hydroxyl groups present in the substance is directly associated with the antioxidant capacity, as observed mainly in dimeric substances. Flavonoids such as catechin, epicatechin, and their dimmers, as well as procyanidins, exhibit strong antioxidant activity and can be cardioprotective, antimutagenic, and anticarcinogenics agents.
The presence of substances such as catechin, epicatechin, their dimers and polymers in L. chinensis have been previously reported ,,,,,, and it is possible to infer that flavonoids and condensed tannins may be partly responsible for the antioxidant activity of EtOAc, considering many studies that describe these different classes of compounds as potent antioxidants ,, and their use in the food and pharmaceutical industries. Even though in vitro assays on antioxidant activity are not able to predict in vivo activity, the in vitro tests are a convenient, fast, and stable means of screening for future in vivo trials.
In the evaluation of antimicrobial activity, many authors consider the antimicrobial activity of extracts to be significant if the MIC value is 100 μg/ml or lower, moderate if 100< MIC ≤625 μg/ml and weak if MIC >625 μg/ml., Thus, the EtOAc fraction that showed the most significant results demonstrated the lowest MIC values (50 μg/mL) against B. subtilis, B. cereus, and S. aureus [Table 3]. The LE was also significantly active against Gram-positive bacteria B. cereus and S. aureus and against Gram-negative bacteria P. mirabilis (50 μg/mL). The LE was inactive against mycobacteria (M. tuberculosis and M. bovis) and fungi (C. albicans and S. cerevisiae). Due to this, the fractions were not tested against these micro-organism.
|Table 3: Values of minimum inhibitory concentration (μg/mL) of leaves extract and fractions of Litchi chinensis across different micro-organism|
Click here to view
Other studies have investigated the antimicrobial activity of stem bark and seed of this species ,, with positive results. Singh et al., 2013, and Bath and al-Daihan, 2014, reported that the L. chinensis seeds exhibited moderate growth inhibition against Proteus vulgaris, Klebsiella pneumoniae, S. aureus, Streptococcus pyogenes, Bacilllus subtillis, E. coli, and P. aeruginosa. Luteolin, (–)-epicatechin, procyanidin A2, and quercetin-3-O-rutinoside were identified from the EtOAc-soluble extract of litchi leaves. These compounds possessed strong antimicrobial activity towards S. aureus, E. coli, S. dysenteriae, Salmonella, and B. thuringiensis.
The MS analysis of the extract and fractions allowed for the indication of catechin and quercetin [Supplementary Figure 1], [Supplementary Figure 2], [Supplementary Figure 3], as well as polymeric proanthocyanidins. The presence of these compounds justifies the satisfactory MIC values found for antimicrobial activity and is supported by previous studies., Moreover, in quantitative terms, total phenolics and flavonoids compounds showed a positive correlation with MIC results of r = 0.75 and r = 0.5, respectively [Table 2].
The search for new antibiotics to treat infectious diseases caused by multiresistant micro-organism is necessary. Thus, plants are presented as a viable source of new herbal substances or as an alternative to this problem. In this study, we observed excellent MIC values (?<100 μg/mL) against S. aureus and B. cereus for LE and EtOAc fractions. S. aureus is one of the most important agents of opportunistic and nosocomial infections and is considered a significant causative agent for infections in the community, in addition to multi-drug resistance to currently used antibiotics. This results in increased morbidity and mortality rates.
The LE and the EtOAc showed the percentage of activity of 40% and 75%, respectively, against the different micro-organism used. These results demonstrate that the partitioning process can increase the antimicrobial activity. However, when one takes into account the yield factor in the calculation of the total activity, it has been observed that the LE displays better results in terms of total activity [Table 4] compared to the EtOAc (5384 and 2576, respectively). This fact can be explained by the synergy between the constituent compounds, increasing the antimicrobial action. Hex, BuOH, and Aq fractions did not show satisfactory activity against micro-organism in the evaluation using the microdilution broth method (MIC >100 μg/ml).
|Table 4: Total activity of the extract and fractions from the leaves extract of Litchi chinensis with minimum inhibitory concentration ≤100 μg/ml|
Click here to view
Of great relevance, the LE of L. chinensis and its fractions did not show cytotoxicity at the highest concentration used (160 μg/mL) on murine peritoneal macrophages by MTT method.
| Conclusion|| |
The phytochemical profile of the extract and fractions of L. chinensis Sonn. presented phenolic compounds as the main chemical constituents, such as flavonoids (quercetin and catechin) and condensed tannins. The EtOAc showed high levels of total phenolics and flavonoids and the best results of antioxidant tests, while LE showed the best result for antimicrobial activity, suggesting the possibility of the compounds act synergistically to this biological activity. Therefore, we demonstrated the feasibility of the leaves from this species as a source of bioactive compounds, being it renewable and available throughout the entire year. However, additional studies should be conducted aimed at the isolation of new compounds from leaves of L. chinensis and further evaluation of their potential use as an antimicrobial and/or an antioxidant agent that may be useful for the pharmaceutical and food industries.
The authors are grateful to Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) and Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG) and Federal University of Alfenas (UNIFAL-MG) for financial support and to Prof. Dr. Wagner Vilegas (Projeto Biota/FAPESP) and Dr. Marcelo José Dias for the analyzes with mass spectrometry.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Orwa C, Mutua A, Kindt R, Jamnadass R, Simons A. Agroforestree Database: A Tree Reference and Selection Guide Version 4.0. Kenya: World Agroforestry Centre; 2009.
Zhou HC, Lin YM, Li YY, Li M, Wei SD, Chai WM, et al
. Antioxidant properties of polymeric proanthocyanidins from fruit stones and pericarps of Litchi chinensis
Sonn. Food Res Int 2011;44:613-20.
Kilari EK, Putta S. Biological and phytopharmacological descriptions of Litchi chinensis
. Pharmacogn Rev 2016;10:60-5.
Ibrahim SR, Mohamed GA. Litchi chinensis
: Medicinal uses, phytochemistry, and pharmacology. J Ethnopharmacol 2015;174:492-513.
Faysal MM. Justification of use of some medicinal plants to treat various diseases in Khulna, Bangladesh. Ethnobotanical Lealf 2008;12:1231-5.
Zhao M, Yang B, Wang J, Liu Y, Wang W, Jiang Y. Identification of polisaccharides from pericarp tissues of litchi (Litchi chinensis
Sonn.) fruit in relation to their antioxidant activities. Carbohydr Res 2006;341:634-8.
Yang DJ, Chang YZ, Chen YC, Liu SC, Hsu CH, Lin JT, et al.
Antioxidant effect and active components of litchi (Litchi chinensis
Sonn.) flower. Food Chem Toxicol 2012;50:3056-61.
Duan X, Jiang Y, Su X, Zhang Z, Shi J. Antioxidant properties of anthocianins extracted from litchi (Litchi chinensis
Sonn.) fruit pericarp tissues in relation to their role in the pericarp browning. Food Chem 2007;101:1365-71.
Su D, Zhang R, Hou F, Zhang M, Guo J, Huang F, et al.
Comparison of the free and bound phenolic profiles and cellular antioxidant activities of litchi pulp extracts from different solvents. BMC Complement Altern Med 2014;14:9.
Li J, Jiang Y. Litchi flavonoids: Isolation, identification and biological activity. Molecules 2007;12:745-58.
Wen L, You L, Yang X, Yang J, Chen F, Jiang Y, et al.
Identification of phenolics in litchi and evaluation of anticancer cell proliferation activity and intracellular antioxidant activity. Free Radic Biol Med 2015;84:171-84.
Roya S, Besra SE, De T, Banerjee B, Mukherjee J, Vedasiromoni JR. Induction of apoptosis in human leukemic cell lines U937, K562 andHL-60 by Litchi chinensis
leaf extract via activation of mitochondria mediated caspase cascades. Open Leuk J 2008;1:1-14.
Bhoopat L, Srichairatanakool S, Kanjanapothi D, Taesotikul T, Thananchai H, Bhoopat T, et al.
Hepatoprotective effects of lychee (Litchi chinensis
Sonn.): A combination of antioxidant and anti-apoptotic activities. J Ethnopharmacol 2011;136:55-66.
Jiang G, Lin S, Wen L, Jiang Y, Zhao M, Chen F, et al.
Identification of a novel phenolic compound in litchi (Litchi chinensis
Sonn.) pericarp and bioactivity evaluation. Food Chem 2013;136:563-8.
Wang L, Lou G, Ma Z, Liu X. Chemical constituents with antioxidant activities from litchi (Litchi chinensis
Sonn.) seeds. Food Chem 2011;126:1081-7.
Prasad KN, Yang B, Yang S, Chen Y, Zhao M, Ashraf M, et al
. Identification of phenolic compounds and appraisal of antioxidant and antityrosinase activities from litchi (Litchi chinensis
Sonn.) seeds. Food Chem 2009;116:1-7.
Li S, Xiao J, Chen L, Hu C, Chen P, Xie B, et al
. Identification of A-series oligomeric procyanidins from pericarp of Litchi chinensis
by FT-ICR-MS and LC-MS. Food Chem 2012;135:31-8.
Su D, Ti H, Zhang R, Zhang M, Wei Z, Deng Y, et al.
Structural elucidation and cellular antioxidant activity evaluation of major antioxidant phenolics in lychee pulp. Food Chem 2014;158:385-91.
Babbar N, Oberoi HS, Uppal DS, Patil RT. Total phenolic content and antioxidant capacity of extracts obtained from six important fruit residues. Food Res Int 2011;44:391-6.
Alan LH. Medicines from nature: Are natural products still relevant to drug discovery? Trends Pharmacol Sci 1999;20:196-8.
Dall'Agnol R, Ferraz A, Bernardi AP, Albring D, Nör C, Sarmento L, et al.
Antimicrobial activity of some Hypericum
species. Phytomedicine 2003;10:511-6.
Ríos JL, Recio MC. Medicinal plants and antimicrobial activity. J Ethnopharmacol 2005;100:80-4.
Cushnie TP, Lamb AJ. Antimicrobial activity of flavonoids. Int J Antimicrob Agents 2005;26:343-56.
Noldin VF, Cechinel Filho V, Delle Monache F, Benassi JC, Christmann IL, Pedrosa RC, et al
. Chemical composition and biological activities of the leaves of Cynara scolymus
L. (artichoke) cultivated in Brazil. Quim Nova 2003;26:331-4.
Leitão SG, Castro O, Fonseca EN, Julião LS, Tavares ES, Leo RR, et al
. Screening of central and South American plant extracts for antimycobacterial activity by the alamar blue test. Rev Bras Farmacogn 2006;16:6-11.
Denise OG, Luciano SM, Mônica TP. Antibiotics: Therapeutic importance and perspectives for the discovery and development of new agents. Quim Nova 2010;33:667-9.
Singleton VL, Orthofer R, Lamuela-Raventos RM. Analysis of total phenols and other oxidation substrates and antioxidants by means of Folin-Ciocalteau reagent. Methods Enzymol 1999;299:152-78.
Kalia K, Sharma K, Singh HP, Singh B. Effects of extraction methods on phenolic contents and antioxidant activity in aerial parts of Potentilla atrosanguinea
lodd. And quantification of its phenolic constituents by RP-HPLC. J Agric Food Chem 2008;56:10129-34.
Brand-Wiliams W, Cuvelier ME, Berset C. Use of a free radical method to evaluate antioxidant activity. Food Sci Technol 1995;28:25-30.
Yildirim A, Mavi A, Kara AA. Determination of antioxidant and antimicrobial activities of Rumex crispu
s L. extracts. J Agric Food Chem 2001;49:4083-9.
Clinical and Laboratory Standards Institute. Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically. Approved Standard. M7-A6. 6th
ed. Wayne, PA, USA: Clinical and Laboratory Standards Institute; 2003.
Clinical and Laboratory Standards Institute. Reference Method for Broth Dilution Antifungal Susceptibility Testing of Yeasts. Approved Standard. M27-A3. 3rd
ed. Wayne, PA, USA: Clinical and Laboratory Standards Institute; 2008.
Singh R, Hussain S, Verma R, Sharma P. Anti-mycobacterial screening of five Indian medicinal plants and partial purification of active extracts of Cassia sophera
and Urtica dioica
. Asian Pac J Trop Med 2013;6:366-71.
Chavasco JM, Prado E Feliphe BH, Cerdeira CD, Leandro FD, Coelho LF, Silva JJ, et al.
Evaluation of antimicrobial and cytotoxic activities of plant extracts from Southern Minas Gerais Cerrado. Rev Inst Med Trop Sao Paulo 2014;56:13-20.
Fabri RL, Nogueira MS, Moreira JR, Bouzada ML, Scio E. Identification of antioxidant and antimicrobial compounds of Lippia
Species by bioautography. J Med Food 2011;14:840-6.
Bonjar GH. New approaches in screening for antibacterials in plants. Asian J Plant Sci 2004;3:55-60.
Eloff JN. Quantification the bioactivity of plant extracts during screening and bioassay guided fractionation. Phytomedicine 2004;11:370-1.
Pereira IO, Marques MJ, Pavan AL, Codonho BS, Barbiéri CL, Beijo LA, et al.
Leishmanicidal activity of benzophenones and extracts from Garcinia brasiliensis
mart. Fruits. Phytomedicine 2010;17:339-45.
Liu L, Xie B, Cao S, Yang E, Xu X, Guo S. A-type procyanidins from Litchi chinensis
pericarp with antioxidant activity. Food Chem 2007;105:1446-51.
Le Roux E, Doco T, Sarni-Manchado P, Lozano Y, Cheynier V. A-Type proanthocyanidins from pericarp of Litchi chinensis
. Phytochemistry 1998;48:1251-8.
Guo J, Chen J, Lin L, Xu F. Five chemical constituents of the ethyl acetate fraction from ethanol extract of semen litchi. J Med Plants Res 2012;6:168-70.
Yang D, Chang W, Hsu C, Liu C, Wang Y, Chen Y. Protective effect of a litchi (Litchi chinensis
Sonn.)-flower-water-extract on cardiovascular health in a high- fat/cholesterol- dietary hamsters. Food Chem 2010;119:1457-64.
Chen YC, Lin JT, Liu SC, Lu PS, Yang DJ. Composition of flavonoids and phenolic acids in lychee (Litchi chinensis
Sonn.) flower extracts and their antioxidant capacities estimated with human LDL, erythrocyte, and blood models. J Food Sci 2011;76:C724-8.
Zhao M, Yang B, Wang J, Liu Y, Yu L, Jiang Y, et al.
Immunomodulatory and anticancer activities of flavonoids extracted from litchi (Litchi chinensis
sonn) pericarp. Int Immunopharmacol 2007;7:162-6.
Kroon PA, Iyer A, Chunduri P, Chan V, Brown L. The cardiovascular nutrapharmacology of resveratrol: Pharmacokinetics, molecular mechanisms and therapeutic potential Curr Med Chem 2010;17:2442-55.
Ghasemzadeh A, Ghasemzadeh N. Flavonoids and phenolic acids: Role and biochemical activity in plants and human. J Med Plants Res 2011;5:6697-703.
Rodrigues AM, Marcilio Fdos S, Frazão Muzitano M, Giraldi-Guimarães A. Therapeutic potential of treatment with the flavonoid rutin after cortical focal ischemia in rats. Brain Res 2013;1503:53-61.
Shahwar D, Raza MA, Mughal MA, Abbasi MA, Ahmad VU. Comparative study on the antioxidant and antimicrobial activities of stem-bark extract of Litchi chinensis
and its organic fractions. J Chem Soc Pak 2010;32:357-62.
Pessuto MB, Costa IC, Souza AB, Nicoli FM, Palazzo de Mello JC, Petereit F, et al.
Antioxidant activity of extracts and condensed tannins from leaves of Maytenus ilicifolia
Mart. ex Reiss. Quím Nova 2009;32:412-6.
Yao LH, Jiang YM, Shi J, Tomás BF, Datta N, Singanusong R, et al
. Flavonoids in food and their health benefits. Plant Foods Hum Nutr 2004;59:113-22.
Dutra RC, Leite MN, Barbosa NR. Quantification of phenolic constituents and antioxidant activity of Pterodon emarginatus
vogel seeds. Int J Mol Sci 2008;9:606-14.
Rosa EA, Silva BC, Silva FM, Tanaka CM, Peralta RM, Oliveira CM, et al
. Flavonoids and antioxidant activity in Palicourea rigida
Kunth, Rubiaceae. Rev Bras Farmacogn 2010;20:484-8.
Qi S, Huang H, Huang J, Wang Q, Wei Q. Lychee (Litchi chinensis
Sonn.) seed water extract as potential antioxidant and anti-obese natural additive in meat products. Food Control 2015;50:195-201.
Kuete V. Potential of Cameroonian plants and derived products against microbial infections: A review. Planta Med 2010;76:1479-91.
Singh JP, Singh SK, Chandel R, Mishra B, Suneetha V. Evaluation of antimicrobial and antioxidant property of lychee's seed for therapeutic purpose. Int J Pharm Sci Rev Res 2013;19:72-6.
Bhat RS, Al-Daihan S. Antimicrobial activity of Litchi chinensis
and Nephelium lappaceum
aqueous seed extracts against some pathogenic bacterial strains. J King Saud Univ Sci 2014;26:79-82.
Wen L, Wu D, Jiang Y, Prasad KN, Lin S, Jiang G, et al
. Identification of flavonoids in litchi (Litchi chinensis
Sonn.) leaf and evaluation of anticancer activities. J Funct Foods 2014;6:555-63.
Powell JP, Wenzel RP. Antibiotic options for treating community-acquired MRSA. Expert Rev Anti Infect Ther 2008;6:299-307.
[Figure 1], [Figure 2]
[Table 1], [Table 2], [Table 3], [Table 4]