Next Article in Journal
Biosorption of Precious Metals Present at Dilute Concentrations on Fungal Pellets
Next Article in Special Issue
From Foods to Chemotherapeutics: The Antioxidant Potential of Dietary Phytochemicals
Previous Article in Journal
Successful Manufacturing Protocols of N-Rich Carbon Electrodes Ensuring High ORR Activity: A Review
Previous Article in Special Issue
Curcumin Improved Glucose Intolerance, Renal Injury, and Nonalcoholic Fatty Liver Disease and Decreased Chromium Loss through Urine in Obese Mice
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Processes
Volume 10
Issue 4
10.3390/pr10040646
processes-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Advances on the Antioxidant Activity of a Phytocomplex Product Containing Berry Extracts from Romanian Spontaneous Flora

by
Elena Moroșan
1,†,
Ana Andreea Secăreanu
2,†,
Adina Magdalena Musuc
3,*,†,
Magdalena Mititelu
1,*,
Ana Corina Ioniță
1,
Emma Adriana Ozon
2,*,
Adriana Maria Dărăban
4 and
Oana Karampelas
2
1
Department of Clinical Laboratory and Food Safety, Faculty of Pharmacy, “Carol Davila” University of Medicine and Pharmacy, 6 Traian Vuia Street, 020945 Bucharest, Romania
2
Department of Pharmaceutical Technology and Biopharmacy, Faculty of Pharmacy, “Carol Davila” University of Medicine and Pharmacy, 6 Traian Vuia Street, 020945 Bucharest, Romania
3
“Ilie Murgulescu” Institute of Physical Chemistry, 202 Spl. Independentei, 060021 Bucharest, Romania
4
Pharmaceutical Science Department, “Vasile Goldiș” Western University of Arad, Dermatocosmetology and Cosmetic Products, 86 Liviu Rebreanu Street, 310045 Arad, Romania
*
Authors to whom correspondence should be addressed.
†
These authors contributed equally to this work.
Processes 2022, 10(4), 646; https://0-doi-org.brum.beds.ac.uk/10.3390/pr10040646
Submission received: 28 December 2021 / Revised: 20 March 2022 / Accepted: 23 March 2022 / Published: 25 March 2022
(This article belongs to the Special Issue Chemical Characterization and Bioactivity Evaluation of Medicinal Plants)
Browse Figures
Versions Notes

Abstract

:
The present study aimed to evaluate the antioxidant properties of a phytocomplex product obtained using 10% hydroalcoholic extractive solutions, in equal proportions, from Ribes nigrum, Rubus idaeus, Rubus fruticosus and Fragaria moschata fresh fruits harvested from the spontaneous flora of Romania. These plant products were recognized for their rich antioxidant content. The phytochemical profile was assessed using HPLC chromatography and UV-Vis spectrometry. The obtained results highlighted the presence of complex bioactive compounds with antioxidant actions, namely anthocyanins, proanthocyanins and vitamin C. The antioxidant actions of the hydroalcoholic extractive solutions and the phytocomplex product were evaluated using chemiluminescence, electrochemical and superoxide dismutase (SOD) methods. The experimental results showed evident antioxidant activity in both the hydroalcoholic extracts and the phytocomplex product.

1. Introduction

Recently, numerous studies have been conducted on free radicals formed in the human body as a result of endogenous metabolic activity and the influence of various agents with higher or lower degrees of toxicity on the cells and tissues of living organisms [1,2,3,4]. The attack produced by free radicals is manifested either locally on the component cells of an organ or simultaneously on several tissues, gradually leading to serious local or general disorders. In the case of analyses performed on the potency of different free radical types, it was established that, in most cases, the evolution of biochemical processes and reactions triggers (simultaneously or at different times) some factors which favor the effects of oxidative stress [5,6,7,8].
Oxidative stress represents the aggression produced at the molecular level by an imbalance between pro-oxidants and antioxidants—in favor of the first—that is manifested as negative effects at the tissue and organ level. Reactive oxygen species (ROS) are produced in aerobic organisms under physiological conditions. These intermediate metabolic products induce pathological changes only when they can no longer be neutralized by endogenous antioxidants [9,10,11,12,13,14,15]. Numerous recent studies have highlighted the main mechanisms occurring at a cellular level through which reactive oxygen species start to show harmful and toxic effects: (i) the peroxidation of membrane lipids, which causes changes in the functionality of cell membranes and cell organs; (ii) damage to DNA molecules, which induces mutagenic processes and cell death; (iii) the alteration of protein and polycarbohydrate structures (hyaluronic acid); and (iv) the disruption of calcium homeostasis in the vascular wall [16,17,18,19].
In cases where oxidative stress leads to pathological conditions, therapeutic intervention with exogenous, synthetic or natural antioxidants is necessary. Current trends in medicine focus on the use of natural products from a plant or animal origin [20,21,22,23,24,25,26].
The most important mechanisms through which oxidative stress occurs are: (i) an increase in the formation of free oxygen and nitrogen radicals following exposure to large amounts of oxygen in the presence of toxins; (ii) a decrease in endogenous, physiological antioxidants, and especially of some enzymes, such as glutathione-peroxidase (GSH), manganese or copper-zinc superoxide-dismutase, with a role in the protection of organisms; and (iii) a decrease in antioxidants in the human diet.
Oxidative stress induces apoptosis. Antioxidants may reduce the ability of ROS to participate in processes that lead to apoptosis, such as the infiltration and adhesion of neutrophils, by activating myocyte receptors or altering cell membranes; by altering the homeostasis of calcium ions, which affects the sarcoplasmic reticulum membrane; and by the oxidation of nucleic acids [27,28].
During the processes by which the formation of free radicals takes place, an increase in vascular permeability and the extravasation of macromolecules, manifested as edema, inflammation, etc., has been observed. The considerable formation of oxygen free radicals during the inflammatory process, as a result of oxidative toxins, produces increases in vascular permeability [29,30,31].
Oxidative stress can also occur intracellularly (i) after intoxication with certain chemicals, such as ethanol, paracetamol, acetanilide derivatives and adriamycin, or (ii) after the reperfusion of ischemic tissues and organs [25,32]. In this case, the antioxidants used must be able to cross the cell membrane. For this purpose, the antioxidants must be lipophilic or amphipathic, such as vitamin E or β-carotene [16,33,34].
The use of hydrophilic antioxidant compounds leads to a longer time requirement for the restoration of intracellular antioxidant capacity [35,36].
Since the compounds that are richest in vitamin C, an especially effective antioxidant, are fruits, it is a challenge to obtain natural pharmaceutical products with improved therapeutic qualities that contain the whole range of active properties. Phytocomplex technology represents a promising alternative; it incorporates phytoconstituents that are soluble in water (terpenoids or flavonoids) or standardized plant and fruit extracts. In this context, we obtained a phytocomplex with antioxidant activity by combining 10% hydroalcoholic extractive solutions obtained from the fresh fruits of Ribes nigrum, Rubus idaeus, Rubus fruticosus and Fragaria moschata in equal proportions. In general, berry fruits are one natural resource of antioxidants, and contain a high concentration of phenolic compounds (benzoic acid, cinnamic acid, tannins, stilbenes and flavonoids, such as anthocyanins, flavanols and flavonols), vitamin C and other antioxidant compounds [37,38,39]. However, the amount of bioactive compounds is determined by diverse factors, such as the variety, species, region, cultivation, ripeness, harvesting time, weather conditions and storage time [40,41,42,43,44,45]. Therefore, this study aimed to evaluate each hydroalcoholic extractive solution component from a qualitative and quantitative chemical point of view, as well as investigate its antioxidant activity.
Numerous studies have shown the benefits of phytocomplexes in increasing vitamin C bioavailability due to the presence of other essential components, such as bioflavonoids, in the structure of the extract. It has been demonstrated that these ingredients act as antioxidants, thus considerably reducing vitamin C oxidation [46,47,48].

2. Materials and Methods

2.1. Materials

Fresh fruits, including Ribes nigri (blackcurrants), Rubi idaei (raspberries), Rubi fruticosi (blackberries) and Fragariae moschatae (strawberries), were collected from spontaneous flora in the forest zone of Arges County, Romania. All reagents and reference substances used were chromatographically pure (manufacturer: Merck, Fluka). Ultra-pure water obtained by ultra-purification with a Millipore system was used.
All reagents used (chloroform, Na2HPO4, KH2PO4, nitrotetrazolium blue (NBT), methyl blue, tetramethylene diamine (TMED) and 0.05 M phosphate buffer, pH 7.8) for the determination of the antioxidant action of the phytocomplex by the superoxide dismutase method were of analytical reagent grade (from Merck).

2.2. Methods

Preparation of the phytocomplex
The extracts of Ribes nigri fructus, Rubi idaei fructus, Rubi fruticosi fructus and Fragariae moschatae fructus were obtained according to a working protocol previously developed in the literature [36,49,50].
10 g of vegetable product was brought into contact with 100 mL of a 70% hydroalcoholic mixture (1/10 ratio).
10 g of vegetable product and 100 mL of 70% hydroalcohol (1/10 ratio) were mixed in a glass vial, wrapped in paper, covered and left to soak for 7 days in a cool place (15–25 °C) protected from direct sunlight. The mixture was shaken 3–4 times/day.
After 7 days, the hydroalcoholic extracts were filtered and the hydroalcoholic extractive solutions were placed into hermetically sealed vessels and kept in the refrigerator. Similarly, 10 g of partially solubilized vegetable products were subjected to a new solubilization process with a 70% hydroalcoholic mixture for another 4 days in similar conditions. After that, the hydroalcoholic extracts were filtered and mixed with the previously obtained hydroalcoholic solutions. Finally, hydroalcoholic extractive solutions at a concentration of 10% in 70% ethyl alcohol were obtained. In these conditions, the phytocomplex consisted of equal parts of the 10% hydroalcoholic solutions obtained from Ribes nigri, Rubi idaei, Rubi fruticosi and Fragariae moschatae fructus.
Qualitative chemical analysis of hydroalcoholic extractive solutions
The obtained hydroalcoholic solutions were chemically analyzed, both qualitatively and quantitatively. The identification of compounds with antioxidant action (flavones, anthocyanins, proanthocyanins and tannins) was performed using specific reactions [51,52,53,54,55].
Varian-ProStar equipment with a quaternary pump and UV-Vis detector with a diode array was used to perform the UV-Vis determinations. A C18 type stationary phase column (Discovery C18 250 × 4.6 mm, 5 µm particles) was used. The mobile phase consisted of two solvents: Solvent A was an aqueous phosphoric acid solution (3 mL of 85% phosphoric acid diluted to 1000 mL with water) and Solvent B was acetonitrile.
The composition of the mobile phase was determined with a linear gradient from 0 to 100%. The measured time was from 0 min to 70 min (0, 20, 25, 65 and 70 min). The composition mobile phase was chosen as follows: Solvent A/Solvent B = 90/10; 70/30; 60/40 and 90/10.
The mobile phase flow rate was 1 mL/min, and the detection was performed at 335 nm. The spectra were recorded in the range of 200–400 nm. The injection volume was 20 µL. All determinations were performed at room temperature.
The peaks were determined by comparing the retention times of the analyzed substances with the retention times of the reference substances.
Quantitative analysis of the extractive solutions from the phytocomplex
The determination of anthocyanosides and proanthocyanosides was made using a spectrophotometric method with a Carry 50 UV-Vis spectrophotometer, equipped with a quartz cuvette (optical path = 1.0 cm). Dilutions (1/10 (v/v)) of hydroalcoholic extractive solutions in 95% ethanol were prepared. The quantitative determination of anthocyanosides and proanthocyanosides was made using the method described in our previous research [36].
Quantitative determination of ascorbic acid
Ascorbic acid was determined using the HPLC method. Considering its antioxidant role, as well as the fact that it is unstable, the samples were analyzed immediately after preparation and also after 60 days.
The HPLC methods used in the literature [56,57] propose determination by reversed-phase liquid chromatography. The determinations were performed with a Varian-ProStar chromatograph with a quaternary pump and a PDA (photo-diode assay) detector. The chromatographic system was previously described in another study [36], together with the accuracy, precision (repeatability and reproducibility) and linearity of the method. The HPLC method was validated and previously published [36].
The sample solutions were obtained by diluting 0.10 mL of antioxidant phytocomplex to 25.0 mL with 0.1% H3PO4 in water with addition of 1% CH3OH.
Accuracy and repeatability were verified by injecting standard solutions in triplicate on the same day, and reproducibility was verified by repeating the determinations for three consecutive days.
The specificity was demonstrated by the chromatogram obtained by analyzing a prepared standard matrix, using the standards of flavones and polyphenolic derivatives of ODP (ortho-dihydroxy phenols), which usually appear with ascorbic acid in vegetable hydroalcoholic extractive solutions.
The linearity and concentration range were established by applying the regression analysis of the experimental data obtained by analyzing three sets of standard solutions prepared as described in our previous study [36]. The results are presented in Figures S1 and S2 in the Supplementary Material.
Analysis of the antioxidant action of the phytocomplex
Determination of antioxidant action by chemiluminescence [36]
To determine the antioxidant activity, a Turner Designs TD 20/20 chemiluminometer was used. The chemiluminometer was coupled with a computer equipped with 1.5 mL encapsulated glass cuvettes, a luminol chemiluminescence generating system, and hydrogen peroxide in TRIS-HCl buffer at pH = 8.6, with a 1 mL volume.
All measurements were made in triplicate.
Determination of the antioxidant action of the phytocomplex by electrochemical methods [36]
The cyclic voltammetry (CV), linear potential sweep voltammetry (LSV) and square wave voltammetry (SWV) techniques were used. The inert electrode was 0.12 mol L−1 H2SO4 in an alcoholic solution (methanol). The reagents (methyl alcohol, sulfuric acid, Trolox (6-hydroxy-2,5,7,8 -tetramethyl chroman-2-carboxylic acid) and 2,2′-diphenyl-1-picrylhydrazyl (DPPH)) were of analytical purity (from Sigma). Measurements were made at a temperature of 25 °C under an inert atmosphere, after a de-aeration of the used solutions.
For stock alcoholic solutions, 10−3 mol L−1 Trolox, freshly prepared and maintained overnight at 4 °C, and 3 × 10−3 mol L−1 DPPH were used.
UV-Vis spectrometric determinations were performed using a JASCO V-530 spectrometer.
Determination of the antioxidant action of the phytocomplex using the superoxide dismutase (SOD) method [36]
The hydroalcoholic extractive solutions from Ribes nigri fructus, Rubi idaei fructus, Fragariae moschatae fructus and Rubi fruticosi fructus, and the phytocomplex solution were tested for antioxidant action. The presence of oxygen in the reaction mixture was ensured by purging air for 20 min. The sample and standard were placed under a 17 W fluorescent lamp for 15 min. A wavelength of 560 nm was used. The quantities used are listed in Table 1.
An SOD unit was expressed as the number of enzymes (unit/mg protein) that reduced 50% of formed formazan [58,59,60].

3. Results

Qualitative chemical composition of hydroalcoholic extractive solutions
In the hydroalcoholic extractive solutions obtained from Ribes nigri fructus, Rubi idaei fructus, Rubi fruticosi fructus and Fragariae moschatae fructus was detected the presence ofanthocyanins, proanthocyanins and tannin, and flavone only in Ribes nigri fructus and Rubi fruticosi fructus (Table 2).
The UV-Vis absorption spectra for the hydroalcoholic extractive solutions of Ribes nigri fructus, Rubi idaei fructus, Rubi fruticosi fructus and Fragariae moschatae fructus are shown in Figures S3–S5 in the Supplementary Materials. The obtained UV-Vis absorption spectra for the hydroalcoholic extractive solutions of Ribes nigri fructus, Rubi idaei fructus, Rubi fruticosi fructus and Fragariae moschatae fructus demonstrated that compounds with antioxidant action from the flavonoid class, such as naringin and naringenin, were present in all the extractive solutions (Figures S6–S9 in the Supplementary Materials) [61,62].
Quantitative chemical composition of hydroalcoholic extractive solutions
The results of the quantitative chemical determinations are presented in Table 3.
In the UV-Vis spectra of all the hydroalcoholic extracts, compounds with a maximum absorption between 270–290 nm were detected, which corresponds with the UV-Vis spectra of naringenin and naringin compounds; this indicates that this type of derivative is also present in the studied solutions.
It is also well known in the literature that flavanols have two major UV-Vis absorption bands at 250–285 nm and 320–385 nm [63].
Determination of vitamin C content
The calculation of the ascorbic acid concentration in the phytocomplex was performed via the external calibration method, using a regression curve linearity analysis for 10 analyses.
The ascorbic acid in the sample was determined using the retention time and the HPLC spectrum of the eluate (ascorbic acid in aqueous solution presents an absorption band at 243 nm). The obtained content of ascorbic acid was 14.14 ± 1.10 mg/mL.
During the analysis, a low stability of ascorbic acid in the solution was observed. Because of this, the determination of ascorbic acid in the phytocomplex was also performed after 60 days.
The two freshly prepared phytocomplex samples were kept for 2 months in parallel (i) at room temperature and (ii) in the refrigerator (4–8 °C). The ascorbic acid content was determined every two weeks by the HPLC method. During this time, the ascorbic acid content of the antioxidant phytocomplex kept in the refrigerator decreased by about 5.5%. The results are presented in Table 4.
The variation of the ascorbic acid content of the antioxidant phytocomplex is shown in Figure S10 in the Supplementary Materials.
Evaluation of the antioxidant action of the phytocomplex by chemiluminescence
In Table 5, the values of the chemiluminescent signal and antioxidant activity of the hydroalcoholic solutions are shown.
The variation in the antioxidant activity of the studied solutions is presented in Figure 1.
The antioxidant activity is explained by the presence of active compounds with antioxidant actions (polyphenolic derivatives), including flavones, catechols and ODPs that were present in the extractive solutions from which the mixtures were formed. Additionally, this category of active compounds included the proanthocyanosides and anthocyanosides identified in the extractive solutions [64,65,66].
Determination of the antioxidant capacity of the phytocomplex by the electrochemical method
In Figure 2, Figure 3, Figure 4, Figure 5 and Figure 6, the behaviors of the hydroalcoholic extracts and phytocomplex with concern to the long-lived radical DPPH are shown at the optimal times established for the measurements.
As shown in Figure 2, Figure 3, Figure 4, Figure 5 and Figure 6, the presence of some antioxidant properties was observed in all investigated samples. Considering that all extracts contained polyphenolic derivatives, including flavones, catechols, ODPs, proanthocyanosides and anthocyanosides, the antioxidant effect was demonstrated [26].
Using this method, the total antioxidant capacity of the extracts was determined, and no fractional separation of the components from the mixture was performed.
The peak current intensities corresponding to each sample were determined, and the differences at 0 and 2 min from the peak current intensity belonging to DPPH in the absence of any antioxidant were also determined. The obtained values are shown in Table 6.
The antioxidant capacity of the samples was determined using Equation (1) as follows:
A C sample = V sample × K × I sample − I blank I Trolox − I blank
where ACsample is the antioxidant capacity of the sample, V is the sample volume, K is the sample dilution factor, Isample is the intensity of the anodic peak current of DPPH in the presence of the sample, Iblank is the intensity of the peak current of DPPH in the absence of any antioxidant and ITrolox is the peak current intensity for DPPH in the presence of Trolox. The obtained values are presented in Figure 7.
The obtained data were verified by using UV-Vis spectrometry to determine the observable changes in the absorption maxima of DPPH at 519 nm. However, the spectrometric determinations for Ribes nigri fructus, Rubi idaei fructus, Rubi fruticosi fructus, Fragariae moschatae fructus and the phytocomplex were inconclusive since the extracts had absorption maxima in the 500–520 nm region, which made it difficult to observe possible changes in the maximum of DPPH.
The electrochemical method has the advantage of avoiding the interference encountered in the spectrometric method; therefore, it is widely applicable.
Determination of the antioxidant capacity of the phytocomplex by the superoxide dismutase (SOD) method
From the analysis of the data presented in Table 7, it was observed that all the solutions used at 1/10 dilutions showed antioxidant activity that inhibited the amount of enzyme by more than 50%. Among the studied solutions, the most accentuated antioxidant action was found in the extract solution from Fragariae moschatae fructus (100%). The antioxidant phytocomplex inhibited the activity of the enzyme by 81%.
The antioxidant activity of the phytocomplex was due to the presence of polyphenolic derivatives, represented by flavones, catechols and ODPs.

4. Discussion

All the hydroalcoholic extractive solutions from Ribes nigri fructus, Rubi idaei fructus, Rubi fruticosi fructus and Fragariae moschatae fructus contained anthocyanins, proanthocyanins and tannins. Flavones were identified only in Ribes nigri fructus and Rubi fruticosi fructus. In the final mixture, obtained by mixing equal proportions of the extractive solutions of Ribes nigri fructus, Rubi idaei fructus, Rubi fruticosi fructus and Fragariae moschatae fructus, the same chemical constituents were identified. The obtained results agree with the data reported in the literature [42,56,67,68,69,70]. The variation of the total anthocyanin and proanthocyanin content is due to environmental factors.
Several authors have reported a total polyphenol amount of 267.5 mg/100 g [67], 720.0 mg/100 g [56] and 799.0 mg/100 g [70] for Ribes nigri; 320.0 mg/100 g [67], 260.0 mg/100 g [56] and 264.0 mg/100 g [70] for Rubi idaei; and 507.5 mg/100 g [67], 320.0 mg/100 g [56] and 460.0 mg/100 g [70] for Rubi fruticose. The total anthocyanins reported by the same authors are as follows: 235.0 mg/100 g [67], 260.0 mg/100 g [56] and 229.0 mg/100 g [70] for Ribes nigri; 37.5 mg/100 g [67], 40.0 mg/100 g [56] and 141.0 mg/100 g [70] for Rubi idaei; and 115.0 mg/100 g [67], 80.0 mg/100 g [56] and 141.0 mg/100 g [70] for Rubi fruticose. The ascorbic acid content was between 6.0 and 20.4 mg/100 g for Rubi fruticose, 18.3 and 30.17 mg/100 g for Rubi idaei, and 44.5 and 167.8 mg/100 g for Ribes nigri [56,67,70]. Light, temperature, soil conditions, rainfall and agronomic practices are factors that may contribute to the differences observed in total polyphenol, anthocyanin and ascorbic acid contents between various studies.
From a chemical quality point of view, the phytocomplex had a content of 0.105 mg of anthocyanosides/100 mL solution and proanthocyanins of 0.041 LA ‰/100 mL solution. In the literature, a strong correlation between phenol and anthocyanin content and antioxidant activity has been observed [71]. Our results confirmed that antioxidant activity is related to the total anthocyanins. Wu et al. determined the total content of anthocyanins in redcurrant compared to blackcurrant. The authors concluded that the blackcurrant, which had a higher concentration of anthocyanins, possessed the highest antioxidant activity [72].
The same range of chemical constituents was identified in the final preparation product, obtained by mixing the extractive solutions of Ribes nigri fructus, Rubi idaei fructus, Rubi fruticosi fructus and Fragariae moschatae fructus in equal proportions. Based on the spectrophotometric results of studies performed on the hydroalcoholic extractive solutions from the extracts of Romanian spontaneous flora berries, it can be concluded that they contain bioactive compounds with antioxidant action. This is confirmed by the presence of active molecules with antioxidant properties (polyphenolic derivatives), including flavones, anthocyanins, proanthocyanins and tannin.
The results also confirmed the presence of ascorbic acid after 60 days (at room temperature) in the antioxidant phytocomplex. Ascorbic acid also acts as an antioxidant compound by preventing free radicals from inducing DNA damage [73].
In the present study, the obtained values of antioxidant activity (AA%) proved that the hydroalcoholic extractive solution of Rubi idaei fructus had the most pronounced antioxidant activity (AA% = 83.80), followed by the hydroalcoholic extractive solution of Ribes nigri fructus (AA% = 79.50), Rubi fructicosi fructus (AA% = 51.04) and that of Fragariae moschatae fructus (AA% = 40.54). The phytocomplex had a lower antioxidant activity (AA% = 68.54) compared with that of the hydroalcoholic extractive solutions of Rubi idaei fructus and Ribes nigri fructus, and a more pronounced activity compared to that of the extractive solutions of Rubi fructicosi fructus and Fragariae moschatae fructus.
Angela and Meireles reported a high antioxidant potential for ethanol samples due to the effect of ethanol in increasing the extraction of bioactive compounds, mostly anthocyanins [74]. The same results were obtained by Peschel et al. with ethanol extract of blackcurrant residue, which had a higher antioxidant potential (20.08%) compared to aqueous extract residue (9.95%) [75].
Our results showed that the berry fruits (dark or red colored fruits) are an abundant source of active substances, and their mixed product showed a significant antioxidant potential due to the synergetic effect of the individual compounds. Subsequently, important steps to improve the final antioxidant properties include the extraction method, processing and storage of the extracts.

5. Conclusions

Based on the studies performed on the hydroalcoholic extractive solutions from Ribes nigri fructus, Rubi idaei fructus, Rubi fruticosi fructus and Fragariae moschatae fructus using specific analytical methods, it can be concluded that bioactive compounds with antioxidant action, such as anthocyanins, proanthocyanins, tannins and vitamin C, are present in their composition. By the determination of ascorbic acid dosage using the HPLC method, it was found that the antioxidant phytocomplex contained 14.14 mg/mL of ascorbic acid.
The methods used to determine the antioxidant activity of the prepared phytocomplex indicated a significant antioxidant activity of both the extractive plant solutions that were part of the phytocomplex and the mixture obtained in the form of a phytocomplex-type nutritional supplement. The higher antioxidant activity of the phytocomplex is due to the presence of several compounds, such as anthocyanins, proanthocyanins, tannins and flavones, with antioxidant actions.

Supplementary Materials

The following supporting information can be downloaded at: https://0-www-mdpi-com.brum.beds.ac.uk/article/10.3390/pr10040646/s1, Figure S1. Calibration curve for ascorbic acid dosage in the antioxidant phytocomplex; Figure S2. Response factor to ascorbic acid determinations; Figure S3. Chromatogram of the hydroalcoholic extractive solution from Ribes nigri fructus; Figure S4. Chromatogram of the hydroalcoholic extractive solution from Rubi fruticosi fructus; Figure S5. Chromatogram of the hydroalcoholic extractive solution from Fragariae moschatae fructus; Figure S6. UV-Vis spectrum of the hydroalcoholic extractive solution from Ribes nigri fructus; Figure S7. UV-Vis spectrum of the hydroalcoholic extractive solution from Rubi fruticosi fructus; Figure S8. UV-Vis spectrum of the hydroalcoholic extractive solution from Rubi idaei fructus; Figure S9. UV-Vis spectrum of the hydroalcoholic extractive solution from Fragariae moschatae fructus; Figure S10. Chromatogram with time variation of ascorbic acid concentration in the antioxidant phytocomplex.

Author Contributions

Conceptualization, E.M. and M.M.; methodology, A.A.S., A.C.I. and O.K.; formal analysis, E.M. and A.M.D.; investigation, M.M., A.A.S., O.K. and A.C.I.; data curation, M.M., E.A.O. and A.M.M.; writing—original draft preparation, M.M., A.M.M. and E.A.O.; writing—review and editing, M.M., A.M.M. and E.A.O. All authors have read and agreed to the published version of the manuscript.

Funding

This research received no external funding. The APC was funded by the “Carol Davila” University of Medicine and Pharmacy.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Forman, H.J.; Zhang, H. Targeting oxidative stress in disease: Promise and limitations of antioxidant therapy. Nat. Rev. Drug Discov. 2021, 20, 689–709. [Google Scholar] [CrossRef]
  2. Pizzino, G.; Irrera, N.; Cucinotta, M.; Pallio, G.; Mannino, F.; Arcoraci, V.; Squadrito, F.; Altavilla, D.; Bitto, A. Oxidative Stress: Harms and Benefits for Human Health. Oxid. Med. Cell. Longev. 2017, 2017, 8416763. [Google Scholar] [CrossRef]
  3. Lobo, V.; Patil, A.; Phatak, A.; Chandra, N. Free radicals, antioxidants and functional foods: Impact on human health. Pharmacogn. Rev. 2010, 4, 118–126. [Google Scholar] [CrossRef] [Green Version]
  4. Pham-Huy, L.A.; He, H.; Pham-Huy, C. Free radicals, antioxidants in disease and health. Int. J. Biomed. Sci. IJBS 2008, 4, 89–96. [Google Scholar] [PubMed]
  5. Di Meo, S.; Venditti, P. Evolution of the Knowledge of Free Radicals and Other Oxidants. Oxid. Med. Cell. Longev. 2020, 2020, 9829176. [Google Scholar] [CrossRef] [PubMed]
  6. Khan, F.; Garg, V.K.; Singh, A.K.; Tinku, T. Role of free radicals and certain antioxidants in the management of huntington’s disease: A review. J. Anal. Pharm. Res. 2018, 7, 386–392. [Google Scholar] [CrossRef] [Green Version]
  7. Balaci, T.; Velescu, B.; Karampelas, O.; Musuc, A.M.; Nitulescu, G.M.; Ozon, E.A.; Nitulescu, G.; Gird, C.E.; Fita, C.; Lupuliasa, D. Physico-Chemical and Pharmaco-Technical Characterization of Inclusion Complexes Formed by Rutoside with beta-Cyclodextrin and Hydroxypropyl-beta-Cyclodextrin Used to Develop Solid Dosage Forms. Processes 2021, 9, 26. [Google Scholar] [CrossRef]
  8. Nicoară, A.C.; Cazacincu, R.G.; Lupuleasa, D.; Miron, D.S.; Rădulescu, F.S. Formulation and in-vitro release testing of rectal suppositories containing nimesulide. Farmacia 2015, 63, 111–117. [Google Scholar]
  9. Pizzino, G.; Bitto, A.; Interdonato, M. Oxidative stress and DNA repair and detoxification gene expression in adolescents exposed to heavy metals living in the Milazzo-Valle del Mela area (Sicily, Italy). Redox Biol. 2014, 2, 686–693. [Google Scholar] [CrossRef] [Green Version]
  10. Rajendran, P.; Nandakumar, N.; Rengarajan, T.; Palaniswami, R.; Gnanadhas, E.N.; Lakshminarasaiah, U.; Gopas, J.; Nishigaki, I. Antioxidants and human diseases. Clin. Chim. Acta 2014, 436, 332–347. [Google Scholar] [CrossRef]
  11. Halliwell, B. Biochemistry of oxidative stress. Biochem. Soc. Trans. 2007, 35, 1147–1150. [Google Scholar] [CrossRef] [PubMed]
  12. Zorov, D.B.; Juhaszova, M.; Sollott, S.J. Mitochondrial Reactive Oxygen Species (ROS) and ROS-Induced ROS Release. Physiol. Rev. 2014, 94, 909–950. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  13. Adam-Vizi, V.; Chinopoulos, C. Bioenergetics and the formation of mitochondrial reactive oxygen species. Trends Pharmacol. Sci. 2006, 27, 639–645. [Google Scholar] [CrossRef] [PubMed]
  14. Bae, Y.S.; Oh, H.; Rhee, S.G.; Yoo, Y.D. Regulation of reactive oxygen species generation in cell signaling. Mol. Cells 2011, 32, 491–509. [Google Scholar] [CrossRef] [Green Version]
  15. Dikalov, S. Cross talk between mitochondria and NADPH oxidases. Free Radic. Biol. Med. 2011, 51, 1289–1301. [Google Scholar] [CrossRef] [Green Version]
  16. Kurutas, E.B. The importance of antioxidants which play the role in cellular response against oxidative/nitrosative stress: Current state. Nutr. J. 2016, 15, 71. [Google Scholar] [CrossRef] [Green Version]
  17. Valko, M.; Leibfritz, D.; Moncol, J.; Cronin, M.T.D.; Mazur, M.; Telser, J. Free radicals and antioxidants in normal physiological functions and human disease. Int. J. Biochem. Cell Biol. 2007, 39, 44–84. [Google Scholar] [CrossRef]
  18. Năstăsescu, V.; Mititelu, M.; Goumenou, M.; Docea, A.O.; Renieri, E.; Udeanu, D.I.; Oprea, E.; Arsene, A.L.; Dinu-Pîrvu, C.E.; Ghica, M. Heavy metal and pesticide levels in dairy products: Evaluation of human health risk. Food Chem. Toxicol. 2020, 146, 111844. [Google Scholar] [CrossRef]
  19. Singh, A.; Kukreti, R.; Saso, L.; Kukreti, S. Oxidative Stress: A Key Modulator in Neurodegenerative Diseases. Molecules 2019, 24, 1583. [Google Scholar] [CrossRef] [Green Version]
  20. Tan, B.L.; Norhaizan, M.E.; Liew, W.-P.-P.; Rahman, H.S. Antioxidant and Oxidative Stress: A Mutual Interplay in Age-Related Diseases. Front. Pharmacol. 2018, 9, 1162. [Google Scholar] [CrossRef] [Green Version]
  21. Akash, M.S.H.; Rehman, K.; Chen, S. Spice plant Allium cepa: Dietary supplement for treatment of type 2 diabetes mellitus. Nutrition 2014, 30, 1128–1137. [Google Scholar] [CrossRef] [PubMed]
  22. Aoki, A.; Inoue, M.; Nguyen, E.; Obata, R.; Kadonosono, K.; Shinkai, S.; Hashimoto, H.; Sasaki, S.; Yanagi, Y. Dietary n-3 Fatty Acid, α-Tocopherol, Zinc, vitamin D, vitamin C and β-carotene are Associated with Age-Related Macular Degeneration in Japan. Sci. Rep. 2016, 6, 20723. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  23. Hovanet, M.V.; Ancuceanu, R.; Dinu, M.; Oprea, E.; Budura, E.A.; Negreṣ, S.; Velescu, B.; Duṭu, L.; Anghel, I.; Ancu, I.; et al. Toxicity and anti-inflammatory activity of Ziziphus jujuba Mill. leaves. Farmacia 2016, 64, 802–808. [Google Scholar]
  24. Hovanet, M.V.; Dociu, N.; Dinu, M.; Ancuceanu, R.; Moroṣan, E.; Oprea, E. A Comparative Physico-chemical Analysis of Acer platanoides and Acer pseudoplatanus Seed Oils. Rev. Chim. 2015, 66, 987–991. [Google Scholar]
  25. Manolescu, B.N.; Oprea, E.; Mititelu, M.; Ruta, L.L.; Farcasanu, I.C. Dietary Anthocyanins and Stroke: A Review of Pharmacokinetic and Pharmacodynamic Studies. Nutrients 2019, 11, 1479. [Google Scholar] [CrossRef] [Green Version]
  26. Popescu, M.L.; Costea, T.; Gird, C.E.; Fierascu, I.; Balaci, T.D.; Fierascu, R.C. Antioxidant activity of romanian Agaricus Blazei Murrill. and Agaricus Bisporus J.E. Lange mushrooms. Farmacia 2017, 65, 329–335. [Google Scholar]
  27. Maulik, N.; Yoshida, T.; Das, D.K. Oxidative Stress Developed During the Reperfusion of Ischemic Myocardium Induces Apoptosis. Free Radic. Biol. Med. 1998, 24, 869–875. [Google Scholar] [CrossRef]
  28. Ioniță, A.C.; Ghica, M.; Moroşan, E.; Nicolescu, F.; Mititelu, M. In Vitro effects of some synthesized aminoacetanilide N’-Substituted on human leukocytes separated from peripheral blood. Farmacia 2019, 67, 684–690. [Google Scholar] [CrossRef]
  29. Ciolan, D.F.; Mînea, A.; Andrieș, A.; Nicoară, A.C.; Rădulescu, F.S.; Miron, D.S. The influence of compendial cells design and experimental setup on the in-vitro similarity of ketoconazole topical creams. Farmacia 2015, 63, 865–871. [Google Scholar]
  30. Oskarsson, H.J.; Coppey, L.; Weiss, R.M.; Li, W.-G. Antioxidants attenuate myocyte apoptosis in the remote non-infarcted myocardium following large myocardial infarction. Cardiovasc. Res. 2000, 45, 679–687. [Google Scholar] [CrossRef] [Green Version]
  31. Teringova, E.; Tousek, P. Apoptosis in ischemic heart disease. J. Transl. Med. 2017, 15, 87. [Google Scholar] [CrossRef] [Green Version]
  32. Sun, L.; Fan, H.; Yang, L.; Shi, L.; Liu, Y. Tyrosol Prevents Ischemia/Reperfusion-Induced Cardiac Injury in H9c2 Cells: Involvement of ROS, Hsp70, JNK and ERK, and Apoptosis. Molecules 2015, 20, 3758–3775. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  33. Wang, W.; Kang, P.M. Oxidative Stress and Antioxidant Treatments in Cardiovascular Diseases. Antioxidants 2020, 9, 1292. [Google Scholar] [CrossRef] [PubMed]
  34. Elahi, M.M.; Kong, Y.X.; Matata, B.M. Oxidative Stress as a Mediator of Cardiovascular Disease. Oxid. Med. Cell. Longev. 2009, 2, 259–269. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  35. Lushchak, V.I. Glutathione Homeostasis and Functions: Potential Targets for Medical Interventions. J. Amino Acids 2012, 2012, 736837. [Google Scholar] [CrossRef] [Green Version]
  36. Moroșan, E.; Mititelu, M.; Drăgănescu, D.; Lupuliasa, D.; Ozon, E.A.; Karampelas, O.; Gîrd, C.E.; Aramă, C.; Hovaneț, M.V.; Musuc, A.M.; et al. Investigation into the Antioxidant Activity of Standardized Plant Extracts with Pharmaceutical Potential. Appl. Sci. 2021, 11, 8685. [Google Scholar] [CrossRef]
  37. Puupponen-Pimiä, R.; Nohynek, L.; Alakomi, H.-L.; Oksman-Caldentey, K.-M. Bioactive berry compounds?novel tools against human pathogens. Appl. Microbiol. Biotechnol. 2005, 67, 8–18. [Google Scholar] [CrossRef]
  38. Cieślik, E.; Gręda, A.; Adamus, W. Contents of polyphenols in fruit and vegetables. Food Chem. 2006, 94, 135–142. [Google Scholar] [CrossRef]
  39. Szajdek, A.; Borowska, E.J. Bioactive Compounds and Health-Promoting Properties of Berry Fruits: A Review. Plant Foods Hum. Nutr. 2008, 63, 147–156. [Google Scholar] [CrossRef]
  40. Benvenuti, S.; Pellati, F.; Melegari, M.; Bertelli, D. Polyphenols, anthocyanins, ascorbic acid, and radical scavenging activity of Rubus, Ribes, and Aronia. J. Food Sci. 2004, 69, FCT164–FCT169. [Google Scholar] [CrossRef]
  41. Häkkinen, S.H.; Törrönen, A.R. Content of flavonols and selected phenolic acids in strawberries and Vaccinium species: Influence of cultivar, cultivation site and technique. Food Res. Int. 2000, 33, 517–524. [Google Scholar] [CrossRef]
  42. Wang, S.Y.; Lin, H.S. Antioxidant Activity in Fruits and Leaves of Blackberry, Raspberry and Strawberry Varies with Cultivar and Developmental Stage. J. Agric. Food Chem. 2000, 48, 140–146. [Google Scholar] [CrossRef] [PubMed]
  43. Connor, A.M.; Luby, J.J.; Hancock, J.F.; Berkheimer, S.; Hanson, E.J. Changes in Fruit Antioxidant Activity among Blueberry Cultivars during Cold-Temperature Storage. J. Agric. Food Chem. 2002, 50, 893–898. [Google Scholar] [CrossRef] [PubMed]
  44. Hakala, M.; Lapveteläinen, A.; Huopalahti, R.; Kallio, H.; Tahvonen, R. Effects of varieties and cultivation conditions on the composition of strawberries. J. Food Compos. Anal. 2003, 16, 67–80. [Google Scholar] [CrossRef]
  45. Skupień, K.; Oszmiański, J. Comparison of six cultivars of strawberries (Fragaria × ananassa Duch.) grown in northwest Poland. Eur. Food Res. Technol. 2004, 219, 66–70. [Google Scholar] [CrossRef]
  46. Carr, A.C.; Frei, B. Toward a new recommended dietary allowance for vitamin C based on antioxidant and health effects in humans. Am. J. Clin. Nutr. 1999, 69, 1086–1107. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  47. Englard, S.; Seifter, S. The biochemical functions of ascorbic acid. Annu. Rev. Nutr. 1986, 6, 365–406. [Google Scholar] [CrossRef]
  48. Pullar, J.M.; Carr, A.C.; Vissers, M.C.M. The Roles of Vitamin C in Skin Health. Nutrients 2017, 9, 866. [Google Scholar] [CrossRef] [Green Version]
  49. Li, H.-B.; Jiang, Y.; Wong, C.-C.; Cheng, K.-W.; Chen, F. Evaluation of two methods for the extraction of antioxidants from medicinal plants. Anal. Bioanal. Chem. 2007, 388, 483–488. [Google Scholar] [CrossRef]
  50. Association of Oficial Agricultural Chemists (AOAC). Official Methods of Analysis, 16th ed.; AOAC International: Washington, DC, USA, 1997. [Google Scholar]
  51. Maity, S.; Chatterjee, S.; Variyar, P.S.; Sharma, A.; Adhikari, S.; Mazumder, S. Evaluation of Antioxidant Activity and Characterization of Phenolic Constituents of Phyllanthus amarus Root. J. Agric. Food Chem. 2013, 61, 3443–3450. [Google Scholar] [CrossRef]
  52. Chai, W.-M.; Shi, Y.; Feng, H.-L.; Qiu, L.; Zhou, H.-C.; Deng, Z.-W.; Yan, C.-L.; Chen, Q.-X. NMR, HPLC-ESI-MS, and MALDI-TOF MS Analysis of Condensed Tannins from Delonix regia (Bojer ex Hook.) Raf. and Their Bioactivities. J. Agric. Food Chem. 2012, 60, 5013–5022. [Google Scholar] [CrossRef] [PubMed]
  53. López, A.; Javier, G.-A.; Fenoll, J.; Hellín, P.; Flores, P. Chemical composition and antioxidant capacity of lettuce: Comparative study of regular-sized (Romaine) and baby-sized (Little Gem and Mini Romaine) types. J. Food Compos. Anal. 2014, 33, 39–48. [Google Scholar] [CrossRef]
  54. Ahn, C.-B.; Shin, T.-S.; Seo, H.K.; Je, J.-Y. Phenolic composition and antioxidant effect of aqueous extract of Arisaema cum Bile, the Oriental Herb Medicine, in human fibroblast cells. Immunopharmacol. Immunotoxicol. 2012, 34, 661–666. [Google Scholar] [CrossRef] [PubMed]
  55. Sunmonu, T.O.; Afolayan, A.J. Evaluation of Polyphenolic Content and Antioxidant Activity of Artemisia afra Jacq. Ex Willd. Aqueous Extract. Pak. J. Nutr. 2012, 11, 618–623. [Google Scholar] [CrossRef] [Green Version]
  56. Rodriguez, M.A.R.; Oderiz, M.L.V.; Lopez-Hernandez, J.; Lozano, J.S. Determination of Vitamin C and Organic Acids in Various Fruits by HPLC. J. Chromatogr. Sci. 1992, 30, 433–437. [Google Scholar] [CrossRef]
  57. Robitaille, L.; Hoffer, L.J. A simple method for plasma total vitamin C analysis suitable for routine clinical laboratory use. Nutr. J. 2015, 15, 40. [Google Scholar] [CrossRef] [Green Version]
  58. Luo, S.; Jiang, X.; Jia, L.; Tan, C.; Li, M.; Yang, Q.; Du, Y.; Ding, C. In Vivo and In Vitro Antioxidant Activities of Methanol Extracts from Olive Leaves on Caenorhabditis elegans. Molecules 2019, 24, 704. [Google Scholar] [CrossRef] [Green Version]
  59. Kim, S.J.; Han, D.; Moon, K.D.; Rhee, J.S. Measurement of Superoxide Dismutase-like Activity of Natural Antioxidants. Biosci. Biotechnol. Biochem. 1995, 59, 822–826. [Google Scholar] [CrossRef] [Green Version]
  60. Deepika, S.; Rajagopal, S.V. Evaluation of in vitro antioxidant activity of flowers of Blepharis Molluginifolia. IJPSR 2014, 5, 2225–2229. [Google Scholar] [CrossRef]
  61. Choi, J.M.; Hahm, E.; Park, K.; Jeong, D.; Rho, W.-Y.; Kim, J.; Jeong, D.H.; Lee, Y.-S.; Jhang, S.H.; Chung, H.J.; et al. SERS-Based Flavonoid Detection Using Ethylenediamine-β-Cyclodextrin as a Capturing Ligand. Nanomaterials 2017, 7, 8. [Google Scholar] [CrossRef]
  62. Fan, R.; Huang, X.; Wang, Y.; Chen, X.; Ren, P.; Ji, H.; Xie, Y.; Zhang, Y.; Huang, W.; Qiu, X.; et al. Ethnopharmacokinetic- and Activity-Guided Isolation of a New Antidepressive Compound from Fructus Aurantii Found in the Traditional Chinese Medicine Chaihu-Shugan-San: A New Approach and Its Application. Evid.-Based Complement. Altern. Med. 2012, 2012, 607584. [Google Scholar] [CrossRef] [PubMed]
  63. Kumar, S.; Pandey, A.K. Chemistry and Biological Activities of Flavonoids: An Overview. Sci. World J. 2013, 2013, 162750. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  64. Paduraru, D.N.; Coman, F.; Ozon, E.A.; Gherghiceanu, F.; Andronic, O.; Ion, D.; Stanescu, M.; Bolocan, A. The use of nutritional supplement in romanian patients–attitudes and beliefs. Farmacia 2019, 67, 1060–1065. [Google Scholar] [CrossRef]
  65. Melendez-Martinez, A.J.; Vicario, I.M.; Heredia, F.J. Provitamin A carotenoids and ascorbic acid contents of the different types of orange juices marketed in Spain. Food Chem. 2007, 101, 177–184. [Google Scholar] [CrossRef]
  66. Yao, X.; Wang, Y.T.; Chen, G. Simultaneous determination of am aminothiols, ascorbic acid and uric acid in biological samples by capillary electrophoresis with electrochemical detection. Biomed. Chromatogr. 2007, 21, 520–526. [Google Scholar] [CrossRef]
  67. Costantino, L.; Albasini, A.; Rastelli, G.; Benvenuti, S. Activity of polyphenolic crude extracts as scavengers of superoxide radicals and inhibitors of xanthine oxidase. Planta Med. 1992, 58, 342–344. [Google Scholar] [CrossRef]
  68. Rotundo, A.; Bounous, G.; Benvenuti, S.; Vampa, G.; Melegari, M.; Soragni, F. Quality and yield of ribes and rubus cultivars grown in Southern Italy hilly locations. Phytother. Res. 1998, 12, S135–S137. [Google Scholar] [CrossRef]
  69. Ancos, B.; Gonzáles, E.M.; Cano, M.P. Ellagic acid, vitamin C, and total phenolic contents and radical scavenging capacity affected by freezing and frozen storage in raspberry fruit. J. Agric. Food Chem. 2000, 48, 4565–4570. [Google Scholar] [CrossRef] [Green Version]
  70. Moyer, R.A.; Hummer, K.E.; Finn, C.E.; Frei, B.; Wrolstad, R.E. Anthocyanins, phenolics, and antioxidant capacity in diverse small fruits: Vaccinium, Rubus, and Ribes. J. Agric. Food Chem. 2002, 50, 519–525. [Google Scholar] [CrossRef]
  71. Kalt, W.; Forney, C.F.; Martin, A.; Prior, R.L. Antioxidant capacity, vitamin C, phenolics, and anthocyanins after fresh storage of small fruits. J. Agric. Food Chem. 1999, 47, 4638–4644. [Google Scholar] [CrossRef]
  72. Wu, X.; Beecher, G.R.; Holden, J.M.; Haytowitz, D.B.; Gebhardt, S.E.; Prior, R.L. Concentrations of Anthocyanins in Common Foods in the United States and Estimation of Normal Consumption. J. Agric. Food Chem. 2006, 54, 4069–4075. [Google Scholar] [CrossRef] [PubMed]
  73. Padayatty, S.J.; Katz, A.; Wang, Y.; Eck, P.; Kwon, O.; Lee, J.H.; Chen, S.; Corpe, C.; Dutta, A.; Dutta, S.K.; et al. Vitamin C as an antioxidant: Evaluation of its role in disease prevention. J. Am. Coll. Nutr. 2003, 22, 18–35. [Google Scholar] [CrossRef] [PubMed]
  74. Angela, M.; Meireles, A. Extracting Bioactive Compounds for Food Products: Theory and Applications; CRC Press: London, UK, 2008. [Google Scholar]
  75. Peschel, W.; Dieckmann, W.; Sonnenschein, M.; Plescher, A. High antioxidant potential of pressing residues from evening primrose in comparison to other oilseed cakes and plant antioxidants. Ind. Crops Prod. 2007, 25, 44–54. [Google Scholar] [CrossRef]
Processes 10 00646 g001 550
Figure 1. Variation of antioxidant activity values in the studied solutions as a function of time.
Figure 1. Variation of antioxidant activity values in the studied solutions as a function of time.
Processes 10 00646 g001
Processes 10 00646 g002 550
Figure 2. Overlapping voltammograms corresponding to DPPH and the addition of 10 µL of the hydroalcoholic solution of Ribes nigri fructus at 0 and 2 min, respectively.
Figure 2. Overlapping voltammograms corresponding to DPPH and the addition of 10 µL of the hydroalcoholic solution of Ribes nigri fructus at 0 and 2 min, respectively.
Processes 10 00646 g002
Processes 10 00646 g003 550
Figure 3. Overlapping voltammograms corresponding to DPPH and the addition of 10 µL of the hydroalcoholic solution of Rubi fruticosi fructus at 0 and 2 min, respectively.
Figure 3. Overlapping voltammograms corresponding to DPPH and the addition of 10 µL of the hydroalcoholic solution of Rubi fruticosi fructus at 0 and 2 min, respectively.
Processes 10 00646 g003
Processes 10 00646 g004 550
Figure 4. Overlapping voltammograms corresponding to DPPH and the addition of 10 µL of Rubi idaei fructus hydroalcoholic solution at 0 and 2 min, respectively.
Figure 4. Overlapping voltammograms corresponding to DPPH and the addition of 10 µL of Rubi idaei fructus hydroalcoholic solution at 0 and 2 min, respectively.
Processes 10 00646 g004
Processes 10 00646 g005 550
Figure 5. Overlapping voltammograms corresponding to DPPH and the addition of 10 µL of the hydroalcoholic solution of Fragariae moschatae fructus at 0 and 2 min, respectively.
Figure 5. Overlapping voltammograms corresponding to DPPH and the addition of 10 µL of the hydroalcoholic solution of Fragariae moschatae fructus at 0 and 2 min, respectively.
Processes 10 00646 g005
Processes 10 00646 g006 550
Figure 6. Overlapping voltammograms corresponding to DPPH and the addition of 10 µL of the hydroalcoholic solution of the phytocomplex at 0 and 2 min, respectively.
Figure 6. Overlapping voltammograms corresponding to DPPH and the addition of 10 µL of the hydroalcoholic solution of the phytocomplex at 0 and 2 min, respectively.
Processes 10 00646 g006
Processes 10 00646 g007 550
Figure 7. Graphical representation of the antioxidant capacity of the hydroalcoholic solutions of Ribes nigri fructus, Rubi fruticosi fructus, Rubi idaei fructus, Fragariae moschatae fructus and the phytocomplex.
Figure 7. Graphical representation of the antioxidant capacity of the hydroalcoholic solutions of Ribes nigri fructus, Rubi fruticosi fructus, Rubi idaei fructus, Fragariae moschatae fructus and the phytocomplex.
Processes 10 00646 g007
Table
Table 1. Working technique.
Table 1. Working technique.
SampleDry Residue/g%SOD Like/(U/mL)
Ribes nigri fructus17.51000
Rubi idaei fructus2.21000
Rubi fruticosi fructus2.44120
Fragariae moschatae fructus3.31000
Phytocomplex5120
ReagentsSample/mLBlank/mL
Phosphate buffer 0.05 M pH 7.82.6 mL2.7 mL
TMED 0.78 M0.05 mL0.05 mL
AM 2 × 10−4 M0.05 mL0.05 mL
NBT 3 × 10−3 M0.2 mL0.2 mL
Vegetable extract0.1 mL-
Table
Table 2. The results of the qualitative chemical analysis.
Table 2. The results of the qualitative chemical analysis.
Extractive SolutionFlavoneAnthocyaninsProanthocyaninsTannin
Ribes nigri fructus+++++++
Rubi fruticosi fructus+++++
Rubi idaei fructus−++++
Fragariae moschatae fructus−+++
“−“—negative reaction; “+”—positive reaction; “++”—intensely positive reaction.
Table
Table 3. The chemical composition of the analyzed hydroalcoholic extracts.
Table 3. The chemical composition of the analyzed hydroalcoholic extracts.
Extractive SolutionAnthocyanins/(mg%)Proanthocyanins/(LA‰/100 mL)
Ribes nigri fructus0.1760 ± 0.00310.0150 ± 0.0018
Rubi fruticosi fructus0.1040 ± 0.00280.0720 ± 0.0023
Rubi idaei fructus0.0733 ± 0.00150.0092 ± 0.0009
Fragariae moschatae fructus0.0244 ± 0.0011 0.0100 ± 0.0012
Table
Table 4. Variation of ascorbic acid content in the antioxidant phytocomplex after 60 days.
Table 4. Variation of ascorbic acid content in the antioxidant phytocomplex after 60 days.
SampleNumber of AnalysesAscorbic Acid after 60 Days
(at Room Temperature)		RSD %
(Relative Standard Deviation)		Ascorbic Acid after 60 Days
(In Refrigerator)
mg/mL%mg/mL%
Antioxidant
phytocomplex		1013.6696.601.1813.8595.47
Table
Table 5. Values of the chemiluminescent signal as a function of time and antioxidant activity in the presence of hydroalcoholic solutions.
Table 5. Values of the chemiluminescent signal as a function of time and antioxidant activity in the presence of hydroalcoholic solutions.
SampleIpAA%v (s−1)
Ribes nigri fructus396.279.5079.2
Rubi fruticosi fructus108451.04216.8
Rubi idaei fructus287.783.857.6
Fragariae moschatae fructus127240.54254.4
Phytocomplex282068.54564.0
Ip—chemiluminescent signal intensity in the presence of the studied solutions at t = 5 s; AA%—antioxidant activity; V—chemiluminescent signal speed after the first 5 s.
Table
Table 6. Values of anodic peak currents of DPPH in the presence of vegetable hydroalcoholic solutions and the phytocomplex.
Table 6. Values of anodic peak currents of DPPH in the presence of vegetable hydroalcoholic solutions and the phytocomplex.
SampleIDPPH (mA)Time (min)Intensity
Ribes nigri fructus3.444t = 00.628
3.208t = 20.864
Rubi fruticosi fructus2.261t = 00.317
2.24t = 20.338
Rubi idaei fructus3.445t = 00.620
3.205t = 20.856
Fragariae moschatae fructus2.801t = 00.096
2.743t = 20.154
Phytocomplex2.338t = 00.437
2.064t = 20.711
Table
Table 7. Percentage inhibitions (%) of lipid peroxidation.
Table 7. Percentage inhibitions (%) of lipid peroxidation.
SampleDilutionsDilutionsDilutions
1/101/1001/1000
Ribes nigri fructus754937
Rubi fruticosi fructus914837
Rubi idaei fructus513737
Fragariae moschatae fructus1004943
Antioxidant phytocomplex814311
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Share and Cite

MDPI and ACS Style

Moroșan, E.; Secăreanu, A.A.; Musuc, A.M.; Mititelu, M.; Ioniță, A.C.; Ozon, E.A.; Dărăban, A.M.; Karampelas, O. Advances on the Antioxidant Activity of a Phytocomplex Product Containing Berry Extracts from Romanian Spontaneous Flora. Processes 2022, 10, 646. https://0-doi-org.brum.beds.ac.uk/10.3390/pr10040646

AMA Style

Moroșan E, Secăreanu AA, Musuc AM, Mititelu M, Ioniță AC, Ozon EA, Dărăban AM, Karampelas O. Advances on the Antioxidant Activity of a Phytocomplex Product Containing Berry Extracts from Romanian Spontaneous Flora. Processes. 2022; 10(4):646. https://0-doi-org.brum.beds.ac.uk/10.3390/pr10040646

Chicago/Turabian Style

Moroșan, Elena, Ana Andreea Secăreanu, Adina Magdalena Musuc, Magdalena Mititelu, Ana Corina Ioniță, Emma Adriana Ozon, Adriana Maria Dărăban, and Oana Karampelas. 2022. "Advances on the Antioxidant Activity of a Phytocomplex Product Containing Berry Extracts from Romanian Spontaneous Flora" Processes 10, no. 4: 646. https://0-doi-org.brum.beds.ac.uk/10.3390/pr10040646

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop