Comparative Studies Between Mauritia flexuosa and Mauritiella armata

Mauritia flexuosa and Mauritiella armata belong to the family Arecaceae and are widely found in Brazil. Aim: In this work were evaluated: the phytochemical profile of the secretion popularly known as M. flexuosa wine, antioxidant activity of leaf, root and petiole hydroethanolic extracts of the two species, as well as the quantification of flavonoids and the chromatographic profile by means of High Performance Liquid Chromatography. Materials and Methods: The chromatographic profile was determined by high performance liquid chromatography, quantification of flavonoids and antioxidant activity, were performed by spectrophotometric method. Results: Antioxidant activity and presence of flavonoids were observed in the extracts of all the analyzed structures of the two species. The phytochemical profile of the wine evidenced the presence of secondary metabolites reported in other structures of M. flexuosa. In the chromatographic analysis, it was observed that the extracts evaluated have between three and nine compounds. Conclusion: Further studies should be performed to identify the active compounds in the two species.


INTRODUCTION
The Arecaceae family is composed of 1500 species distributed in 200 genera, found mainly in tropical areas of the planet.They are part of this family the Mauritiella armata (buritirana ou xiriri) 1,2 and also the Mauritia flexuosa (buriti, miriti, muriti or palmeirado-brejo), 3 this last secretes a substance called wine, which has easy fermentation and is used in folk medicine as a fortifier and in the control of intestinal problems. 4,5 ese palm trees have wide distribution in Brazil. 6[10][11] Faced with this, the objective of this work was to carry out phytochemical prospecting in the M. flexuosa

Preparation of the extract
For each 10g of plant material 100 mL of 70% ethanol was added.The mixture was kept at controlled temperature at 35 o C (± 2) for seven days, then filtered and placed in a greenhouse at 40 o C (± 2) for three to five days for solvent drying, then they were stored in a refrigerator at -5°C until the time of analysis.

Phytochemical analysis
It was evaluated the presence of secondary metabolites: glycosides, 12 saponins, steroids and terpenes following the methodology of Royo et al. 13 anthraquinones were identified according to Mello et al. 14 and Falkenberg et al. 15 Tests to verify the presence of alcohols were made as described by Andrade. 16The acidity, aldehyde and ketone tests were reproduced according to the procedures suggested by Cruz and Galhardo. 17The verification of reducing sugars was done using the methodology described by Beserra et al. 18 and the thin layer chromatography (TLC) technique reported by Mercadante-Simões et al. 19 with modifications.

High Performance Liquid Chromatography (HPLC)
Each extract was diluted in 0.1% trifluoroacetic acid solution: acetonitrile (90:10) at 1 mg/mL concentration, taken ultrasonic bath for 10 min, then filtered on a millex filter and injected into the liquid chromatograph (Waters), equipped with binary pump 1525, automatic injector 717, automatic fraction collector III, diode arrangement detector 2996 and Soft Ware Empower 1, previously stabilized.The column used was type C18 250×4.6 mm 5 micron Spherisorb.The mobile phase used was a 0.1% trifluoroacetic acid solution diluted in acetonitrile (99:1).The injection volume of the samples was 20 μL, which ran in a flow of 0.6 mL/min.Readings of peak were taken at the wavelength of 220 nm.The analyzes occurred at a temperature of 30°C.

Quantification of flavonoids
The total flavonoid content was determined from the Alves and Kubota 20 spectrophotometric method with the use of aluminum chloride.The standard curve equations were obtained from reading the absorbances at the 425nm wavelength of the standard standard solution of rutin (20, 25.30, 35, 40, 45 μg/mL).In 100 μL of AlCl 3 methanolic solution (5% w/v), it was added 600 μL of samples, at the concentrations of 100, 200, 300, 400, 500 μg/mL and the volume was completed to 2.5 mL with methanol.

Antioxidant Activity
The evaluation of the antioxidant activity of the samples was measured by the DPPH free radical sequestering method (2,2-Diphenyl1-1-Pricrilhydrazyl) described by Brand-Williams, 21 with modifications.The stock solution of DPPH was prepared at the concentration of 40 μg/mL and stock solutions of the samples prepared at 500 μg/mL. 21,22 amples and DPPH were diluted in methanol.In separate test tubes were pipetted 3 mL of DPPH and 0.5 mL of the solutions of each sample in the concentrations according to Table 1.Then the samples stayed for 30 min under cover the light and then was proceeded the spectrophotometer read at a wavelength of 517 nm.For white, methanol was used and for the negative control it was pipetted 3mL of DPPH and 0.5mL of methanol.Gallic acid was used as a positive control at concentrations from 0.33 to 1.66 μg/mL.All the tests were performed in triplicate and the mean of the results was used for the calculation of percentage of antioxidant activity.The percentage of antioxidant activity (% AA) was calculated with equation 1: Where AbsCont is the absorbance value of the control and AbsAmos represents the absorbance value of the sample. 22

Phytochemical analysis of wine
They were identified: Glycosides, anthraquinones and saponins in the most concentrated dilutions of wine and still steroids and triterpenoids in extraction I, but not in extraction II.The M. flexuosa also has these metabolites in other structures, for the ethanol extract from its leaves has already been reported presence of steroids / triterpenoids, saponins, flavonoids and tannins. 23In the pulp there are phenolic compounds, flavonoids and phytosteroids. 24And in the roots there are flavonoids and triterpenes 25,26 (Table 2).Aldehydes were not detected as well as reducing sugars in both TLC and Fehling reactions.However, it was detected the presence of ketones and alcohols in a most intense way in the potassium dichromate test than in ferric chloride, which is directed to identification of phenols. 27The verification of ketones, alcohols and the pH = 4 may be due to the fermentative process that consumed the sugars, where the primary alcohols produce acids and the secondary alcohols, the end product is the ketone. 28,29romatographic analysis In the extracts of the two species, the compound with retention time from 6.545 to 6.574 min was the majority for all analyzed parts.The compound with retention time between 5.257 and 5.351 min was the second most found in all structures, except for the root of M. armata where the second most prevalent compound is the one with retention time from 4.231 to 4.227 min, being this the third major in: leaves of both species and in the petiole of M. flexuosa (Table 3).
In the leaves of M. flexuosa were identified eight compounds, whereas this structure in M. armata has only three compounds, which coincide in prevalence in the two species.In the M. flexuosa root were detected six compounds and in M. armata seven.It is possible to verify that the compound with retention time 6.545 to 6.574 min is majority in the roots of both species and the compound with retention time 5.928 to 6.098 min is not present in the roots of M. flexuosa.The other compounds vary in quantity in the roots of the two plants.Among all the extracts the petiole of M. armata was the one with the highest number of compounds (9) (Table 3).In M. flexuosa wine there are six compounds, the retention time of these substances was different from the retention times of the compounds found in the plant structures, as can be observed in Table 3.

Antioxidant activity and quantification of flavonoids
All the samples had antioxidant activity and it was possible to verify the presence of flavonoids, however when compared, the structures of M. armata showed higher antioxidant activity and amount of flavonoids than in the corresponding samples of M. flexuosa (Table 4).
In both palms, the leaf extracts were the ones with the highest EC 50 value as can be seen in Table 4.In M. armata and M. flexuosa the leaves exhibited greater antioxidant potential and flavonoid content than the leaves of Calamus rotang (EC 50 of 387,948 μg/mL and flavonoid content of 2.68 EQ mg/g) 30 which is popularly used as antihelmintic and in the treatment of eye problems. 31  12.61 μg/mL), 32 parts of this plant are used in traditional medicine for the treatment of disorders of memory, fever, inflammation, paralysis and its leaves in particular possess antibacterial activity. 33All these species mentioned belong to the family Arecaceae.
The best antioxidant performance of the leaves of M. armata (EC 50 of 50.6 μg/mL) may be due to the amount of flavonoids (7.92 mg/g.E) that was higher than that found in M. flexuosa (4.85 mg/g.E), in addition M. armata possesses only three compounds while M. flexuosa has eight (Table 4), this smaller amount of constituents in its extract reduces the possibility of interaction between the metabolites and potentiates the antioxidant activity of the extract. 34e roots of M. armata and M. flexuosa had intermediate values of antioxidant activity when compared to the other analyzed structures, with EC 50 of 75.6 and 98.7 μg/mL respectively and although they presented antioxidant activity, the values are lower than that described for roots of Borassus flabellifer (15.75 μg/mL) that also belongs to the Arecaceae family. 35The roots of the two species presented contents of flavonoids higher than those found in petioles, as can be observed in Table 4.
It was not possible to realize the tests of antioxidant activity of the wine at any of the concentrations.Initially, the samples were cloudy, not being possible the spectrophotometric analysis.Faced with this concentrations were reduced and it was centrifuged at 14,000 rpm and it was made white with the same centrifuged dilutions of the sample, however the absorbance was still higher than that of the control.The solvent methanol was changed to ethanol, but anyway there was no success.
The two species have phyto-constituents of interest, mainly flavonoids.It is important to perform analyzes to verify biological activities for both species, especially with M. armata, once the tests showed that the species has flavonoids and antioxidant activity higher than M. flexuosa and also because there are few studies of this plant related to the chemistry composition, besides, the prospection of these species and the proof of potential for use, can contribute to the valorization and conservation of the biome cerrado.

CONCLUSION
The presence of flavonoids allows the results of the antioxidant activity found for both species.The M. flexuosa wine evidenced secondary metabolites reported in other plant structures.Further studies should be performed to identify the active compounds.
Samples of petiole, leaf and root of three different individuals of M. flexuosa and M. armata were collected in region of vereda in Bonito of Minas Gerais, the collection was carried out by the Doctor Maria Olivia Mercadante Simões.The plant material was oven dried at 40°C (±2) until it presents a constant mass, later it was pulverized and stored in paper bags in the refrigerator (5°C).Exsicates were deposited in the Montes Claros Herbarium: MCMG and identified by Rubens Manoel dos Santos (Mauritia flexuosa Mart.(Buriti) n. 5777; Mauritiella armata (Mart.)Burret (Xiriri) n.5778a.

Table 1 : Sample concentrations in the antioxidant activity tests.
However, they have less antioxidant activity than the leaves of Phoenix dactylifera (EC 50 corresponding to 7.44 to Pharmacognosy Journal, Vol 11, Issue 1, Jan-Feb, 2019