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Article

Light Can Induce Accumulation of Nutritional Antioxidants in Black Chokeberry Cell Suspension Culture

by
Gergana Krasteva
1,
Tsvetanka Teneva-Angelova
1,
Ilian Badjakov
2,
Ivayla Dincheva
2,
Vasil Georgiev
1,* and
Atanas Pavlov
1,3,*
1
Laboratory of Cell Biosystems, Institute of Microbiology, Bulgarian Academy of Sciences, 4000 Plovdiv, Bulgaria
2
AgroBioInstitute, Agricultural Academy, 1164 Sofia, Bulgaria
3
Department of Analytical Chemistry and Physical Chemistry, Technological Faculty, University of Food Technologies, 4002 Plovdiv, Bulgaria
*
Authors to whom correspondence should be addressed.
Appl. Sci. 2023, 13(20), 11557; https://doi.org/10.3390/app132011557
Submission received: 27 September 2023 / Revised: 20 October 2023 / Accepted: 21 October 2023 / Published: 22 October 2023
(This article belongs to the Special Issue Food Processing and Preservation Technologies: Advances and Applications)
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Abstract

:

Featured Application

This work outlines the potential application of Aronia melanocarpa plant cell suspension in cellular agriculture as a prospective sustainable source of food additives with antioxidant properties.

Abstract

Cultivation of plant cells under controlled conditions is a highly effective and fast developing technology that recently was considered as a branch of cellular agriculture. Cellular agriculture is a multidisciplinary approach for sustainable and renewable production of agricultural goods and raw materials at cellular, rather than organismal, level. However, in contrast to the rapid advance in development of cultured meat and precision fermentation, the production of nutritional supplements from plant cells is still in its infancy. One of the limiting factors, striating commercialization of plant cells for food production, is the low yields of target bioactive metabolites. In this work, the changes in phenolics, anthocyanins and exopolysaccharides accumulation during cultivation of Aronia melanocarpa (Michx.) Elliott cell suspension cultures in darknessor under illumination (16 light and 8 dark) were investigated. The data showed that the highest contents of total phenolics (8.17 ± 0.39 mg GAE/g DW), total anthocyanins (0.011 ± 0.001 mg cyanidin-3-glucoside equivalents/g DW) and antioxidant activities (DPPH—21.36 ± 0.29 µM TE/g DW; TEAC—10.08 ± 0.07 µM TE/g DW; FRAP—34.85 ± 1.47 µM TE/g DW; and CUPRAC—126.74 ± 9.15 µM TE/g DW) were achieved when the cells were grown under illumination (16 light and 8 dark). In contrast, when the culture was grown indarkness, the highest amounts of accumulated dry biomass (8.68 ± 0.35 g/L) and exopolysaccharides production (2.10 ± 0.07 g/L) were reached. The results demonstrated that light can be used as an affordable and highly effective factor to control the production of valuable antioxidants by black chokeberry cell suspension culture.

1. Introduction

Aronia melanocarpa [Michx.] Elliot (black chokeberry) is a plant native to East Canada and North America that belongs to the Rosaceae family [1]. The plant is popular, with fruits that are among the richest source of antioxidants and colorants among all berries [2,3,4,5,6,7]. Anthocyanins, procyanidins, flavonoids and phenolics, found in fruits, have severalbeneficial health properties such as antioxidant, anti-inflammatory, anti-cancerogenic, antidiabetic, cardioprotective and hepatoprotective effects [1,8,9,10,11,12,13]. Aronia fruits also contain high amounts of dietary fibers and polysaccharides with interesting medicinal properties [1,4,14,15]. Recent research showed that consumption of Aronia juice has antiatherogenic and cardioprotective effects, and improves significantly the lipid profile in rats [16,17]. Because of the observed protective effects on aged rat hearts and moderation of age-related changes in the aortic wall structure in rat models, consumption of Aronia-based functional beverages was recommended as a health prophylactic [18,19,20] measure. Consumption of black chokeberry juice was also found to have a positive effect on connective tissue and thymus aging in rats [21]. By using rat models, it was also found to have antidepressant, neuroprotective and gastro-protective actions [22,23,24]. It was proposed that the observed health-beneficial properties of Aronia juices were contributed by the high content of polyphenols with strong antioxidant activities [2,25]. Recent studies demonstrated that quercetin and epicatechin were the strongest antioxidants in black chokeberry juices, whereas the proanthocyanidins and anthocyanins were responsible for the antioxidant activity in berries [10]. It was also demonstrated that Aronia anthocyanins, because of their strong antioxidant effects, have cytoprotective effects and could ameliorate the acute renal failure in mice model [26]. Because of alltheabovementioned health promoting properties, black chokeberry is classified as a “super food” and as such it was considered as a promising plant for the creation of a biotechnological process for sustainable production of valuable metabolites.
The establishment of callus and shoot cultures from A. melanocarpa has been reported previously and their potential to produce phenolic acids has been demonstrated [27]. It was demonstrated that the production of phenolic acids depends on both concentration of used growth regulators and the cytokinin/auxin ratio [28]. However, in any studied combination, the callus culture produced a lower amount of phenolics when compared to shoots. Liquid cultures of black chokeberry shoots have been established and successfully used for biotransformationof hydroquinone to arbutin [29]. Aronia shoots, including shoot cultures from A. melanocarpa, A. arbutifolia and A. × prunifolia, have been initiated and adapted to liquid cultivation [30]. The effects of precursor feeding, light conditions (UV-A, monochromatic and multispectral lights) and cultivation conditions (solid, agitated and temporary immersion system cultures) have been studied in a focus on increasing the accumulation of phenolic compounds in Aronia shoots [31,32,33,34,35,36]. However, in available scientific literature, there are no records about characterization of A. melanocarpa, cell suspension culture and evaluation of its potential to produce valuable phenolic antioxidants.
Cultivation of plant cells under controlled conditions has a long history [37]. In the last few years, large scale cultivation of plant in vitro systems has been considered as a prospective branch of cellular agriculture, a technology for sustainable manufacture of agricultural goods by using single cells without involving plants or animals [38,39,40,41,42,43,44]. Recently, plant cells were used for production of cosmetic ingredients, plant-based proteins and even for food products as chocolate and coffee [39,40,41,42,45,46,47]. However, the low productivity and the high cost of produced biomass are among the major limiting factors of this technology. To overcome these limitations, several strategies for the optimization of nutrient medium composition and environmental conditions have to be performed experimentally. Light is a very important factor that could affect the growth and target metabolite production in plant cells and tissue systems [33,48]. In arecent study, it was demonstrated that illumination with blue light (at a photoregime of 16 h light and 8 h dark) can significantly increase the content of phenolic acids and flavonoids in A. arbutifolia and A. × prunifolia shoot cultures [34]. However, such experiments with A. melanocarpa cell suspension culture have been not reported.
The aim of this study is to evaluate the effect of light on biomass production and secondary metabolites (anthocyanins, phenolic acids and flavonoids) with antioxidant activities, produced by Aronia melanocarpa (Michx.) Elliott cell suspension cultures. This is the first step in the development of viable biotechnology for production of food additives from an Aronia in vitro system by using the cellular agriculture approach.

2. Materials and Methods

2.1. Plant Material

Cell suspension culture of Aronia melanocarpa (Michx.) Elliott was obtained from callus as described elsewhere [49]. Cells were cultivated at 24 °C on orbital shaker (110 rpm) in 1 L flasks with 200 mL Woody Plant medium with 30 g/L sucrose, 0.5 mg/L kinetin and 2.0 mg/L picloram. The suspension was transferred to fresh medium every 7 days for more than 6 months before the experiments. For the experiments, 7 days’ old suspension was used for inoculum. The cultivation was performed at the same conditions, but half of the flasks (n = 3) were cultivated in darkness, whereas the other half (n = 3) were cultivated under illumination (8803 lumen, Mars Hydro LED Grow Light, Mars-Hydro, Shenzhen, Guangdong, China) at photoperiod 16 h light and 8 h dark. Cells were harvested after 7 days and the growth was evaluated on the basis of Accumulated Dry Biomass (ADB, g/L) and Growth Index (GI) [50].

2.2. Exopolysaccharide Content

Accumulation of exopolysaccharides in culture media was determined by weight as described previously [51] with slight modifications. In brief, polysaccharides were precipitated with ethanol (4 volumes) for 12 h at 4 °C. After vacuum filtration, the residues were air-dried at 50 °C to reach constant weight.

2.3. Total Anthocyanins Assay

Anthocyanins content in cell biomass was measured spectrophotometrically by using pH differential method [52]. Briefly, Aronia cells were air-dried at 50 °C for 12 h. Approximately 0.5 g dry biomass was extracted under sonication (30 min, 120 W) with 50 mL of water/2% HCl in methanol solution (1:1 v/v) [53]. After centrifugation (5000 rpm, 15 min) and filtration of supernatant (0.45 µm PTFE filter), two equal portions of anthocyanins containing extract were diluted with 0.025 M potassium chloride buffer (pH 1.0) and 0.4 M sodium acetate buffer (pH 4.5) by using the same dilution factor. The solutions were incubated for 15 min at room temperature and the absorbance at 520 nm and 700 nm was measured by spectrophotometer (Biochrom WPA Biowave DNA). Cyanidin-3-glucoside (having MW = 449.2, and ε = 26,900) was used to calculate the total anthocyanins content. The results were expressed as mg cyanidin-3-glucoside equivalents per g dry weight (DW) as described elsewhere [52].

2.4. Phenolic Extraction

Air-driedcell biomass was extracted three times with 70% methanol (1:10 w/v) in ultrasonic bath (40 kHz, 80 W) for 30 min. The methanol was removed from the combined fraction by evaporation at 40 °C under vacuum, and the volume was adjusted to 10 mL by using distilled water. Solid phase extraction (C18 Strata, Phenomenex, Danaher Corporation, Washington, DC, USA) was used to separate phenolic fraction [54]. Briefly, the column was conditioned by passing 10 mL of methanol, followed by 10 mL of distilled water [54]. A quantity of 5 mL of sample was loaded and then the polar fraction was removed with 10 mL water. The phenolics were eluted with 2 mL methanol and used for HPLC analyses and antioxidant assays.

2.5. HPLC Quantification

High-performance liquid chromatography (HPLC) was used to analyze the content of phenolic and flavonoids accumulated in Aronia cell biomass. The used HPLC system was Waters 1525 Binary Pump and Waters 2484 Detector. Separation was performed on Supelco Discovery HS C18 column (5 μm, 25 cm × 4.6 mm). The elution solvents were: Solvent A (1% acetic acid) and Solvent B (methanol). Gradient elution at 1.0 mL/min flow rate was used as described previously [54]. For compounds quantification, standard calibration curves (gallic acid, protocatechuic acid, chlorogenic acid, caffeic acid, ferulic acid, vanillic acid, syringic acid, p-coumaric acid, salicylic acid, rosmarinic acid, catechin, epicatechin, hesperidin, rutin, quercetin and kaempferol) were used.

2.6. Total Phenolic Content

The content of total phenolics was analyzed by using the Folin–Ciocalteu assay as reported previously [54]. In brief, the 20 μL of phenolic fraction from Aronia cells biomass extract were mixed with 180 μL Folin–Ciocalteu reagent in 96-well plate. After mixing for 2 min, 100 μL sodium carbonate (7.5%) was added and the plate was incubated at 37 °C. After 8 min, the absorbance at λ = 750 nm was recorded against blank [54]. Total phenolic content was presented as gallic acid equivalents per gram dry biomass.

2.7. Antioxidant Activity Assays

Antioxidant activity of phenolic fractions from Aronia cells biomass extracts was analyzed by using four methods: scavenging of DPPH radical, scavenging of ABTS radical by using trolox equivalent antioxidant capacity (TEAC) assay, ferric reducing antioxidant power (FRAP) and cupric ion reducing antioxidant capacity (CUPRAC), as described elsewhere [54]:

2.7.1. DPPH

Twenty microliters of phenolic fractions from Aronia cells biomass extracts were mixed with 280 μL DPPH (1,1-Diphenyl-2-picrylhydrazyl) radical. Blank sample without extract was prepared as well. The reaction time was 15 min at 37 °C, then the absorbance at λ = 517 nm was recorded and used to calculate the percentage of DPPH radical inhibition. Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid) was used as standard.

2.7.2. TEAC

Twenty microliters of phenolic fractions from Aronia cells biomass extracts were mixed with 280 μL ABTS (2,2′azinobis (3)-ethylbenzthiazoline-6-sulfonic acid) radical. Blank sample without extract was prepared as well. The reaction time was 15 min at 37 °C, then the absorbance at λ = 734 nm was recorded and used to calculate the percentage of ABTS radical inhibition. Trolox was used as standard.

2.7.3. CUPRAC

Twenty microliters of phenolic fractions from Aronia cells biomass extracts, 70 μL of Copper (II) chloride (10 mM), 70 μL of neocuproine (7.5 mM), 70 μL of ammonium acetate buffer (pH 7) and 70 μL of water were mixed in 96-well plate. Blank sample without extract was prepared as well. The reaction time was 10 min at 37 °C, then the absorbance at λ = 450 nm was recorded against the blank. Trolox was used as standard.

2.7.4. FRAP

Twenty microliters of phenolic fractions from Aronia cells biomass extracts were mixed with 280 μL FRAP reagent. Blank sample without extract was prepared as well. The reaction time was 10 min at 37 °C, then the absorbance at λ = 593 nm against the blank was recorded. Trolox was used as standard.
In all antioxidant assays, the results were presented as micromoles trolox equivalents per gram dry biomass.

2.8. Statistical Analyses

All data means were calculated for three independent experiments (n = 3). Standard deviations (±SD) were included. All measurements were implemented in 8 technical repeats. ANOVA analysis, followed by Tukey post hoc test, was used to analyze the data. The significant differences were considered at p ≤ 0.01. Correlation analyses were performed by using Pearson method. Normalization of the data was done by using normalization factors = 1. Each group of variables was tested for normality. Metaboanalyst 5.0 (https://www.metaboanalyst.ca/, accessed on 23 August 2023) and MiniTab 17 (Minitab INC, State College, PA, USA) were used to process the data.

3. Results

The effect of light on biomass and valuable antioxidants accumulation in Aronia melanocarpa cell suspension cultures was investigated in shaking flask cultures. In preliminary experiments (unpublished data), it was found that the culture reached maximal growth at day 7 from the beginning of cultivation, so the cells were grown for 7 days before harvest.

3.1. Evaluation of Cell Culture Growth

Aronia melanocarpa cells showed fast growth in liquid media and accumulated 7.11 ± 0.06 g/L and 8.68 ± 0.06 g/L dry biomass, when cultured under illumination (16 light and 8 dark)and in darkness, respectively (Figure 1A). The corresponding growth indexes are 1.62 ± 0.01 and 2.43 ± 0.06 (Figure 1B).
The data showed that light has a significant negative effect on biomass accumulation (p ≤ 0.01; n = 3). The microscopy study of cell morphology showed that when grown under illumination (16 light and 8 dark), the Aronia cells are an increased size, elongated and with big vacuoles, which is a marker for partial differentiation (Figure 1C). In contrast, when cultured under darkness, the cells are smaller with spherical shape and several small vacuoles, which is a typical profile of plant stem cells (Figure 1D) [55,56,57].

3.2. Anthocyanins Production and Secretion of Exopolysaccharides

Lightstimulates the production of anthocyanins in A. melanocarpa cells, whereas these pigments were completely absent in cells growing in darkness (Figure 2A). Interestingly, when grown in darkness, the cells produced a significantly higher amount of extra cellular polysaccharides (2.10 ± 0.07 g/L) when compared to cells grown under illumination (16 light and 8 dark) (1.30 ± 0.07 g/L) (Figure 2B).

3.3. HPLC Analyses

Phenolics and flavonoids compounds, found in A. melanocarpa cells, were analyzed by using HPLC. The data are presented in Table 1.
A hierarchically clustered heatmap was used for better visualization of the differences in accumulated phenolic compounds found in cells grown in different regimes (Figure 3). Clustering of the data clearly showed the existence of significant differences between the two groups, where the cells cultivated under light cycles showed higher phenolic content.
Identification of the most significant compounds, responsible for the observed variations in the data, was performed by principal component analysis (PCA) (Figure 4).
The analyses showed that principal component 1 (PC1) explains 84.3% of the variations (eigenvalue = 9.2707), and the principal component 2 (PC2) explains 12.6% of the data variations (eigenvalue = 1.3821). PC1 ispositively associated with (+)-Catechin, Chlorogenic acid, Rutin and Hesperidin, whereas the PC2 showed positive associations with Caffeic acid and (−)-Epicatechin.

3.4. Total Phenolic Content and Antioxidant Activities Assay

The total phenolic contents and the antioxidant activities of Aronia cell biomasses obtained in different light regimes (light cycles and darkness) are presented in Table 2.
When cultured under illumination (16 h light and 8 h dark), the cell suspension culture accumulates a significantly higher amount of total phenolics when compared to the dark growing culture (8.17 ± 0.04 mg GAE/g DW, compared to 1.58 ± 0.03 mg GAE/g DW, respectively). In all used assays, the cells cultivated under light cycles showed significantly higher antioxidant activity (Table 2). Correlation analyses showed that total phenolic content has strong correlations with all used methods for antioxidant assay (Table 3).

4. Discussion

The challenges faced by mankind during the last decade are the drivers for the search for and development of innovative technologies for sustainable production of foods and plant-derived goods. Cultivation of plant cells is a promising branch of cellular agriculture with amazing potential in the large scale production of foods and technical goods [39,41,45]. The black chokeberry (Aronia melanocarpa) is a rich source of pigments (anthocyanins), polysaccharides and valuable antioxidants (flavonoids, phenolic acids and flavan-3-ol) [3,6,11,12,13,14,15,58,59]. Because many of these secondary metabolites have proven health benefits, the Aronia was classified as a “superfood” [7,11]. Early reports demonstrated the potential of Aronia melanocarpa shoot and callus cultures for in vitro production of valuable compounds [27,28]. The effects of varying the concentration of used growth regulators [28] and different cultivation conditions, including light [33], on phenolics production were investigated as well, and the potential of Aronia in vitro cultures for production of therapeutic, health-promoting and cosmetic ingredients was suggested. In this work, we demonstrated that the Aronia melanocarpa cell suspension culture accumulates a significant amount of biomass and has great potential for the development of biotechnology for sustainable production of food supplements. When cultivated in darkness, Aronia cells accumulated 8.68 ± 0.06 g/L dry biomass, reaching a growth index of 2.43 ± 0.06 for seven days. This is very high productivity, compared to A. melanocarpa shoot cultures, which reach a growth index of 3.78 ± 0.07 after four weeks’ cultivation in darkness [34].Data showed that light can induce changes in cell morphology, initiate the biosynthesis of anthocyanins and significantly increase the content of phenolic compounds (Figure 1 and Figure 2, Table 1). When cultivated under illumination (photoperiod 16 h light and 8 h dark), Aronia cell suspension accumulates 1812.79 ± 46.19 µg/g DW (or 181.28 mg/100 g DW) chlorogenic acid, which is much higher than the reported amount in A. melanocarpa shoots (127.77 ± 9.41 mg/100 g DW) cultivated under illumination with blue light (photoperiod 16 h light and 8 h dark) for four weeks [31]. The effect of light cycles on phenolic compounds was dramatic, as was demonstrated by heatmap clustering and PCA. It has been reported previously that light can increase the level of secondary metabolites in plant cell suspensions [60]. The positive effect of light on anthocyanins production has been well studied in grape cell suspension cultures [48]. However, this effect has not been studied in A. melanocarpa in vitro cultures. Here, we demonstrated that light cycles can initiate pigment production in A. melanocarpa cells and the content of total anthocyanins-rich 0.011 ± 0.001 mg cyanidin-3-glucoside equivalents/g DW. It is worth noting that cultivation under light cycles leads to a significant decrease in the levels of exopolysaccharides secreted by Aronia cells, from 2.10 ± 0.07 g/L to 1.30 ± 0.07 g/L. Such an effect has not been reported previously. The increased phenolic content in Aronia cells cultivated under illumination correlates with the observed significant increase in antioxidant activities (Table 1, Table 2 and Table 3). The existence of a positive correlation between total phenolic content and antioxidant activity is well known and was reported previously [9,61]. Here, we confirm this in A. melanocarpa cell suspension as well. It is worth noting that, when cultured in darkness, the cell accumulated a higher amount of biomass and secreted more extracellular polysaccharides. This is probably due to the stress that light exerts in cultivated A. melanocarpa cells, and it is well known that secondary metabolites are produced in response to stress, and this could affect the production of primary metabolites and accumulation of biomass [48,62]. However, such a suggestion has to be confirmed on a molecular level and more experiments are needed.

5. Conclusions

Light can be used as an effective and cost-efficient factor for increasing the production of pigments and valuable secondary metabolites with antioxidant activities by Aronia melanocarpa cell suspension cultures. All of the reported results support our hypotheses that A. melanocarpa cell suspension can be used as a prospective in vitro system for sustainable production of food additives by using the cellular agriculture approach. The limitation of this study is the usage of only one source of light and lack of experiments for optimization of the photoperiod. However, the presented experiments provide the fundamentals for future detailed investigations on the molecular mechanisms lying behind the activation of metabolite pathways in Aronia cell suspension cultures by light, and exploring its full potential as a production system for nutritional antioxidants.

Author Contributions

Conceptualization, V.G.; methodology, V.G., I.D. and I.B.; validation, G.K. and T.T.-A.; investigation, G.K., I.D. and I.B.; data curation, V.G. and G.K.; writing—original draft preparation, V.G.; writing—review and editing, A.P. and T.T.-A.; supervision, A.P.; project administration, V.G.; funding acquisition, V.G. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the Bulgarian National Science Fund, Bulgarian Ministry of Education and Science, under the contract KP-06-H66/14 (BG-175467353-2022-04-0214-C01).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The data presented in this study are available on request from the corresponding author.

Conflicts of Interest

The authors declare no conflict of interest.

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Applsci 13 11557 g001
Figure 1. Accumulated dry biomass (A), growth index (B) and cell morphology of Aronia melanocarpa cell suspension culture (light microscopy, Olympus CX23, 40×, bars are 20 µm), grown under light cycles (C) and in darkness (D) for 7 days. The means with different letters differed significantly at p ≤ 0.01.
Figure 1. Accumulated dry biomass (A), growth index (B) and cell morphology of Aronia melanocarpa cell suspension culture (light microscopy, Olympus CX23, 40×, bars are 20 µm), grown under light cycles (C) and in darkness (D) for 7 days. The means with different letters differed significantly at p ≤ 0.01.
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Figure 2. Total anthocyanins content (A) and extracellular polysaccharides production (B) by Aronia melanocarpa cell suspension culture cultivated under light cycles and in darkness for 7 days. The means with different letters differed significantly at p ≤ 0.01.
Figure 2. Total anthocyanins content (A) and extracellular polysaccharides production (B) by Aronia melanocarpa cell suspension culture cultivated under light cycles and in darkness for 7 days. The means with different letters differed significantly at p ≤ 0.01.
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Figure 3. Hierarchically clustered heatmap of phenolic compounds found in Aronia melanocarpa cell suspension culture cultivated under light cycles (green) and in darkness (red). The Ward method with Euclidean distance was used for data clustering. The samples are organized in columns and the metabolites in rows. The highest (red) and lowest (blue) levels of the compounds are mapped.
Figure 3. Hierarchically clustered heatmap of phenolic compounds found in Aronia melanocarpa cell suspension culture cultivated under light cycles (green) and in darkness (red). The Ward method with Euclidean distance was used for data clustering. The samples are organized in columns and the metabolites in rows. The highest (red) and lowest (blue) levels of the compounds are mapped.
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Figure 4. Score plot presenting samples discrimination by the first two principal components of phenolic compounds found in Aronia melanocarpa cell suspension culture cultivated under light cycles (red) and in darkness (blue).
Figure 4. Score plot presenting samples discrimination by the first two principal components of phenolic compounds found in Aronia melanocarpa cell suspension culture cultivated under light cycles (red) and in darkness (blue).
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Table 1. HPLC analyses of phenolic compounds by Aronia melanocarpa cell suspension culture cultivated under light cycles and in darkness for 7 days.
Table 1. HPLC analyses of phenolic compounds by Aronia melanocarpa cell suspension culture cultivated under light cycles and in darkness for 7 days.
Compound,
µg/g DW		Light CyclesDark
Gallic acidND *32.84 ± 16.56
(+)-Catechin2140.32 ± 527.61 a163.99 ± 8.69 b
Chlorogenic acid1812.79 ± 46.19 a86.96 ± 12.68 b
Vanillic acidND64.83 ± 2.06
Caffeic acid48.26 ± 15.28 a37.42 ± 4.89 a
(−)-Epicatechin1934.60 ± 727.56 a154.22 ± 11.43 b
p-Coumaric acid18.81 ± 4.39 a8.35 ± 3.84 b
Salicylic acid686.39 ± 299.58ND
Rutin18.74 ± 0.61ND
Hesperidin81.09 ± 16.62 a7.03 ± 1.59 b
* ND—Not detected; the means with different letters in rows are significantly different at p ≤ 0.01.
Table 2. Antioxidant activities of extracts from Aronia melanocarpa cell suspension cultures cultivated under light cycles and in darkness.
Table 2. Antioxidant activities of extracts from Aronia melanocarpa cell suspension cultures cultivated under light cycles and in darkness.
Antioxidant Activity *Light CyclesDark
DPPH21.36 ± 0.11 a4.27 ± 0.17 b
TEAC10.08 ± 0.07 a1.41 ± 0.12 b
FRAP34.85 ± 0.46 a4.26 ± 0.08 b
CUPRAC126.74 ± 1.49 a18.44 ± 0.54 b
Total Phenolic **8.17 ± 0.04 a1.58 ± 0.03 b
* µM Trolox Equivalents/g Dry Biomass; ** mg Gallic Acid Equivalents/g Dry Biomass; the means with different letters in rows are significantly different at p ≤ 0.01.
Table 3. Correlations between antioxidant activity and total phenolic content of extracts from Aronia melanocarpa cell suspension culture.
Table 3. Correlations between antioxidant activity and total phenolic content of extracts from Aronia melanocarpa cell suspension culture.
Total Phenolic
r *p-Value
CUPRAC0.999870.000000023681
TEAC0.999910.000000013358
DPPH0.999820.000000045980
FRAP0.999850.000000035341
*—Pearson correlation coefficients.
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Krasteva, G.; Teneva-Angelova, T.; Badjakov, I.; Dincheva, I.; Georgiev, V.; Pavlov, A. Light Can Induce Accumulation of Nutritional Antioxidants in Black Chokeberry Cell Suspension Culture. Appl. Sci. 2023, 13, 11557. https://doi.org/10.3390/app132011557

AMA Style

Krasteva G, Teneva-Angelova T, Badjakov I, Dincheva I, Georgiev V, Pavlov A. Light Can Induce Accumulation of Nutritional Antioxidants in Black Chokeberry Cell Suspension Culture. Applied Sciences. 2023; 13(20):11557. https://doi.org/10.3390/app132011557

Chicago/Turabian Style

Krasteva, Gergana, Tsvetanka Teneva-Angelova, Ilian Badjakov, Ivayla Dincheva, Vasil Georgiev, and Atanas Pavlov. 2023. "Light Can Induce Accumulation of Nutritional Antioxidants in Black Chokeberry Cell Suspension Culture" Applied Sciences 13, no. 20: 11557. https://doi.org/10.3390/app132011557

APA Style

Krasteva, G., Teneva-Angelova, T., Badjakov, I., Dincheva, I., Georgiev, V., & Pavlov, A. (2023). Light Can Induce Accumulation of Nutritional Antioxidants in Black Chokeberry Cell Suspension Culture. Applied Sciences, 13(20), 11557. https://doi.org/10.3390/app132011557

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