Aristolochia longa is widely used in traditional medicine in Algeria to treat breast cancer. The aim of the present study was to investigate the response of bone resorption markers to A. According to the A. Serum and urinary creatinine, uric acid, and urea were measured. The intake of A. The pyridinium cross-links pyridinoline PYD and deoxypyridinoline DPD are established markers of bone resorption measured in blood and urine and are used to investigate bone metabolism and manage bone diseases [ 1 ].
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Aristolochiacea is a native plant of Algeria used in traditional medicine. This study was devoted to the determination of polyphenols, flavonoids, and tannins contents of A. Extracts were prepared from aerial parts stems and leaves , fruits and tubers by using various solvents with different polarities such as acetone, methanol and distilled water. Acetone extracts from the aerial parts presented the highest contents of polyphenols For antimicrobial activity, the fruit methanol extract was too efficient against the bacterial strains tested, whereas no effect was observed when these extracts were tested against fungi.
The protein denaturation was found in the aerial parts acetone extract to be These preliminary results could be used to justify the traditional use of this plant and its bioactive substances could be exploited for therapeutic purposes.
Aristolochia longa L. They are widely distributed in practically all continents except Australia, a continent for which only few species are known 1. Aristolochia species contain secondary metabolites that are important natural toxins and traditional medicines 1. The data obtained by Heinrich et al.
Tubers of this plant were used such as astringent, antirheumatic, antitumor, anti-inflammatory and antiseptic 3, 4. The aim of this study was to assess the biological activity of the different extracts of Aristolochia longa L.
Plant material and extracts preparation: Aristolochia longa L. Setif1, Algeria. The aerial parts stem and leaves , fruits and tubers were shadow-dried and pulverized to dry powder. All chemicals were purchased from Sigma. Three extracts acetone, methanol and water were prepared from aerial parts stem and leaves , fruits and tubers. Second successive extraction with 50 ml of the same hydro alcoholic solution was carried out at room temperature for 24 h. The ethyl acetate fraction is dried with anhydrous sodium sulphate, and then evaporated to dryness using a rotary evaporator.
The same parts of plant were extracted with acetone in a Soxhlet apparatus within a period of 6 h. The solvent was removed under vacuum and the crude acetone extract obtained. For the preparation of water extracts, the plant, 20g, was extracted with boiling distilled water ml for 10min. Plant extract yield EY : The yield of the extraction was calculated from the following equation.
Where, W 1 is the weight of extract after evaporation of solvent and W 2 is the dry weight of the plant sample. Samples were incubated at room temperature for 2 h, the absorbance of all samples were measured at nm against a methanol blank using a spectrophotometer. The standard calibration curve was obtained using gallic acid. Flavonoids content: The determination of total flavonoids content was conducted according to the AlCl 3 method 4.
The absorbance of mixture was measured at nm after 30 min of incubation at room temperature. Flavones and flavonols content: Flavones and flavonols were estimated according to the protocol developed by Kosalec et al.
Tannins content: For the determination of tannin content in extracts, a method proposed by Bate-Smith 9 was followed. All tests were carried out in triplicate. The absorbance of the filtrate was measured at , , and nm. DPPH radical scavenging assay : The potential antioxidant activity of plant extracts was assessed on the basis of the scavenging activity of the stable 1, 1-diphenylpicrylhydrazyl DPPH free radical according to the previous described procedures.
Different concentrations of test samples were prepared. The reaction mixtures, consisting of 1ml test sample and 1ml methanolic solution of DPPH, 6.
Methanol was used to zero the spectrophotometer. The ability to scavenge the DPPH radical was calculated using the following equation:. A lower IC 50 value corresponds to a higher antioxidant activity of sample. BHT butylated hydroxy toluene was used as a standard antioxidant Briefly, a solution of 2. From the upper layer, 2. Finally, the absorbance was measured at nm against a blank. Quercetin and BHA butylated hydroxy anisole were used as a control.
A higher absorbance indicates a higher reducing power. In brief, 0. The emulsion obtained was freshly prepared before each experiment. An aliquot 2. Thereafter, the absorbance of each sample was measured at nm. A control consisted of 0. BHA was used as positive standard 19, Tests were carried out in triplicate. The relative antioxidant activity was calculated according to the following formula:.
In vitro anti-inflammatory activity: Anti-inflammatory activity of Aristolochia longa extracts was evaluated by protein denaturation method In this experiment, four solutions were prepared : test solution consist of 0. All above solutions were adjusted to pH 6. After cooling, 2. The percentage inhibition of protein denaturation was calculated by using the following formula:.
Each experiment was done in triplicate and the average was taken. Anti-bacterial activity: Extracts were tested against the reference strains for their inhibitory activity, using two methods: agar diffusion method 22 and the microdilution method minimum inhibitory concentrations MIC The bacteria were grown on Mueller Hinton agar.
Gentamycin was used as positive control since it is commonly used as antibiotic against gram positive and gram negative bacterial sp. A volume 20 mL of each medium was poured into 90 mm diameter Petri dishes. All the experiments were performed in triplicate. Minimum inhibitory concentration MIC : Minimum inhibitory concentration MIC was determined using a common broth microdilution method, in 96 multiwell microtiter plates, in triplicate The 10 th well was considered as growth control, since no extracts solutions were added.
After incubation period, the vials were checked for turbidity and the lowest concentrations of the extract showing no turbidity no growth were regarded as the MIC of the test substance 24, One-way analysis of variance followed by the Tukey test was performed to assess differences between groups. Extracts yields: The extraction yields were obtained after removal of solvent, which ranged from 1.
In our study, the results showed that the aqueous extracts gave us highest yields Table 1. For example, the highest yield in extracts was achieved by the aerial part aqueous extract, where it had considerable proportion of phenols content. The aqueous extraction is carried out by high temperature decoction for 10 min, in fact Su et al. This explains that water high-temperature causes disruption of cells facilitating the penetration of the solvent and solubilizing molecules Martins et al.
Koruthu et al. It has been reported that the solvent polarity highly affects extraction rate and antioxidant activity of plant extracts 31 , and the efficiency of the extraction depends on many parameters, including the extraction time and temperature, the volume and type of the solvents used Total phenolics content: Phenolic compounds such as flavonoids, phenolic acids, and tannins are considered to be major contributors to the antioxidant capacity of plants 7.
It has been suggested that polyphenolic compounds have inhibitory effect on mutagenesis and carcinogenesis in humans, when ingested at up to1g daily from a diet rich in fruits and vegetables 4. The content of total polyphenols, flavonoids and tannins in aqueous, methanol and acetone extracts of Aristolochia longa L. The results showed that the total phenols were characterized by highest values compared with other ingredients in all extracts used Table 1 , but the highest amounts of total phenolics were found in aerial parts acetone extract Flavonoids content: The content of flavonoids expressed in quercetin equivalents varied from 4.
We also noticed that the aqueous extracts contained remarkably lower amounts of these compounds in comparison with acetone and methanol extracts where we observed the highest amounts Previously, Djeridane et al. Tannins content: Determining the tannins content is made by the hemoglobin precipitation test. According to Seidel, water and methanol are both polar solvents which extract particularly glycosylated flavonoids and tannins 34 , and we could interpret these results by the occurrence of different types of tannins.
It is possible to divide the tannins into 2 groups according to their structure: condensed tannins and hydrolysable tannins We conclude that the difference in results is due to the type of the solvent and the part of the plant.
Phenolic compounds possess scavenging ability due to their hydroxyl groups and are known to be powerful antioxidant Phenolic compounds such as tannins and flavonoids are considered to be the major contributors to the antioxidant capacity of plants. Some of diverse biological activities of plants, such as antibacterial activity, may also be related to phenolic compounds Aerial parts methanol and acetone extracts have the highest content 1.
Upon reduction, solution of DPPH fades from purple to yellow. Thus, a lower absorbance at nm indicates a higher radical scavenging activity of extract The scavenging effects of different concentrations from different extracts of Aristolochia longa L.
As it is known, the lower the IC 50 value means the higher the antioxidant capacity of the plant extract. This scavenging activity especially assigned to phenolic compounds i. Our study are in accordance with previous studies which indicate that not only total content, but the type of phenolics and their relative distribution is important for biological activity Previous published papers 4 demonstrated that the root methanol extract of Aristolochia longa L.
In fact, it is widely accepted that higher absorbance at nm is correlated to power reducing 20,