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Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results and discussion
  6. Acknowledgements
  7. References

Antioxidant compounds protect plants against oxidative stress caused by environmental conditions. Different light qualities, such as UV-A radiation and blue light, have shown positive effects on the production of phenols in plants. Kalanchoe pinnata (Lamarck) Persoon (Crassulaceae) is used for treating wounds and inflammations. Some of these beneficial effects are attributed to the antioxidant activity of plant components. We investigated the effects of blue light and UV-A radiation supplementation on the total phenol content, antioxidant activity and chromatographic profile of aqueous extracts from leaves of K. pinnata. Monoclonal plants were grown under white light, white plus blue light and white plus UV-A radiation. Supplemental blue light improved the antioxidant activity and changed the phenolic profile of the extracts. Analysis by HPLC of supplemental blue-light plant extracts revealed a higher proportion of the major flavonoid quercetin 3-O-α-l-arabinopyranosyl (1[RIGHTWARDS ARROW]2) α-l-rhamnopyranoside, as well as the presence of a wide variety of other phenolic substances. These findings may explain the higher antioxidant activity observed for this extract. Blue light is proposed as a supplemental light source in the cultivation of K. pinnata, to improve its antioxidant activity.


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results and discussion
  6. Acknowledgements
  7. References

Reactive oxygen species (ROS), including free radicals, are part of the normal human metabolism and act against infectious agents and in several cell-signaling pathways. On the other hand, in high concentrations, these ROS and free radicals can cause oxidative stress, damaging lipids and DNA [1]. For this reason, they have been linked to many diseases, including inflammatory disorders, heart disease, stroke, atherosclerosis, diabetes and cancer [2-4]. To resist the effects of these ROS and free radicals, the human body produces enzymatic and non-enzymatic antioxidant molecules. Nonetheless, a diet rich in antioxidant compounds is also important [5]. Therefore, interest in natural sources of antioxidant molecules, such as spices and herbs, has increased in recent years [2, 6]. Extracts from these plants have shown benefits in human health, slowing the progress of the disease or alleviating its symptoms [7].

Phenolic compounds, including phenolic acids and flavonoids, have been established as the main contributors to the antioxidant activity of plants, and are more effective than Vitamin C, E and carotenoids in vitro [8-10]. Inclusion of these compounds in the human diet could reduce the risk of developing several diseases [11]. Therefore, several studies have correlated the antioxidant activity of plant extracts with their phenolic content [1, 12, 13]. In plants, phenolic compounds contribute to such ecological functions as defense against insects or the attraction of pollinators. Moreover, they protect plants against oxidative stress that may be caused by normal metabolism and/or environmental conditions [5, 14].

Light is one of the most important environmental factors for plants, as a source of energy and in many other respects. Changes in light quality affect plant development; morphological, anatomical and physiological parameters; and the production of secondary metabolites [15, 16]. UV-A radiation [17-20] and blue light [21, 22] increase the production of phenols, mainly flavonoids, which are UV-absorbing compounds with antioxidant capacity, and act as UV filters [23]. These light qualities induce enzymes involved in phenolic metabolism, such as phenylalanine ammonia-lyase and chalcone synthase [24].

Kalanchoe pinnata is a widespread species of Crassulaceae, and is popularly used to treat several diseases [25-27]. Several pharmacological studies have revealed its antibacterial [28], antidiabetic [29], antihypertensive [30], anticancer [31] and antileishmanial properties [25, 32]. K. pinnata is rich in phenolic compounds, which account for some of these biological activities [25]. The composition of phenolics can vary according to light conditions [33-36].

Some of the biological activities and uses of K. pinnata are related to diseases in which ROS and free-radical stress play an important role, such as in arthritis, cardiovascular disease [37], diabetes mellitus [3] and wounds [38]. The widespread use of K. pinnata to treat these ailments can be attributed, at least in part, to its antioxidant activity.

A previous study used microchemical techniques to detect phenolic compounds in specimens of K. pinnata grown in vitro under supplemental blue light [39]. To determine if cultivation conditions can improve the antioxidant activity and total phenol content, and if these conditions can change the chemical composition of K. pinnata leaf extracts, we studied the effect of supplemental blue light and UV-A radiation on these parameters, since the literature has described the general positive effect of these qualities of radiation in these aspects.

Materials and Methods

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results and discussion
  6. Acknowledgements
  7. References

Plant materials and growth conditions

To obtain monoclonal plants, leaf borders of a matrix plant of K. pinnata (Lamarck) Persoon (RB292.697) were cut into 1 cm2 pieces and surface-sterilized under aseptic conditions. Sterilized explants were transferred to autoclaved glass flasks containing MS medium [40] without growth regulators and supplemented with 0.6 μm myo-inositol, 2.43 μm pyridoxine, 4.1 μm nicotinic acid, 1.48 μm thiamine and 30 g L−1 sucrose. The medium was gelled with agar (8 g L−1) at pH 5.7 adjusted before autoclaving. In vitro plants were cultured under 12 μmol m−2 s−1 of photosynthetically active radiation (PAR), supplied by white fluorescent lamps (20 W – F20T12; – Sylvania, São Paulo, SP, Brazil). The growth-room temperature and photoperiod was 24 ± 2°C and 16 h respectively.

These monoclonal same-age specimens were planted in individual plastic pots (18 cm diameter and 15 cm high) filled with commercial soil mixture (Nutriplan®). The pots were placed in a greenhouse at 28 ± 3°C air temperature, manual irrigation of 10 mL per week and a light period of 16 h day−1.

Light treatments

Three light treatments were used: white lamps (W), white lamps plus blue lamps (W + B) and white lamps plus UV-A lamps (W + UV) (Table 1). Thirty plants were cultivated under each light treatment. Ten daylight fluorescent lamps (59 W; Golden, São Paulo, SP, Brazil) were used as the white-light source in all treatments; five 15 W blue-light fluorescent lamps (LC Light, Rio de Janeiro, RJ, Brazil) were used in the W + B treatment; and five 25 W UV-A-radiation fluorescent lamps (LC Light) were used in the W + UV treatment. The spectral distribution of the lamp radiation (Fig. 1) was measured with a QM1 spectrofluorometer (PTI Inc., Lawrenceville, NY, USA). The level of photosynthetically active radiation (PAR) was measured with a PAR sensor coupled to an FMS2 Hansatech fluorometer (Hansatech Instruments Ltd., King's Lynn, UK). The total PAR intensity was the same for all treatments, ranging from 100 to 400 μmol m² s−1, depending on the position of plant with respect to the light source. The W + B treatment enhanced the blue light by 4–12 μmol m² s−1 (depending on the plant position) compared with W. The W + UV treatment had a fluency of 6–25 W m² of UV-A radiation (depending on the plant position), as measured with a long-wave ultraviolet intensity meter (Black-Ray J-221; Ultra-Violet Products Inc., Pasadena, CA). The pots were rotated three times per week to minimize edge and positional effects on each bench.

Table 1. Specifications of light treatments
 Light information
TreatmentWhite lampsBlue lampsUV-A lampsTotal PAR (μmol m−2 s−1)Blue supplementation (μmol m−2 s−1)UV-A supplementation (W m−2)
  1. W—control treatment; W + B—supplemental blue light treatment; W + UV—supplemental UV-A treatment. *Depending on the plant position with respect to the light source.

W+100–400*
W + B++100–400*4–12*
W + UV++100–400*6–25*
image

Figure 1. The radiation spectrum of each light source. (A)—white lamp; (B)—blue lamp; (C)—UV-A lamp.

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Sample extraction, leaf fresh weight, leaf-cake dry weight and lyophilized-extracts weight

The plants were grown under the light treatments for 60 days, since this is the cultivation time generally used in studies of light effects in higher plants [16, 20]. The plants from each treatment were divided into three groups of 10 plants each. On the same day immediately after harvesting, all the leaves of these plants were collected and weighed using a Voyager digital balance (leaf fresh weight). The leaves were extracted with water at 50°C [33]. After the extractions, the remaining leaf material was dried and weighed (leaf-cake dry weight). The extracts were lyophilized (Heto Holten A/S model Drywinner 3), weighed (lyophilized-extracts weight) and used for the chemical analyses.

Determination of total phenolic content

The total phenolic content in aqueous extracts from the leaves of K. pinnata grown under each light treatment (W, W + B and W + UV) was estimated using the Folin-Ciocalteu method [41].

The Folin-Ciocalteu reagent consists of a mixture of phosphomolybdic and phosphotungstic acids, which is reduced in the presence of reducing agents, such as phenolic compounds. This reduction is accompanied by a color change that can be rapidly measured using spectrophotometry [13].

Sample stock solutions (1 mg mL−1) were diluted to final concentrations of 750, 500, 250, 125 and 50 μg mL−1 in distilled water, and total phenolic content was estimated according to the method of Andrade et al. [42], with slight modifications. After adding Folin-Ciocalteau reagent (2n, 10% v/v in distilled water; Sigma-Aldrich®) to samples, 1 mL sodium carbonate—Na2CO3 (7.5% v/v in distilled water; Sigma-Aldrich®) was added and the mixture was allowed to react at room temperature and in the dark. After 60 min, the absorbance was measured using a spectrophotometer (Libra S22; Biochrom Ltd., Cambridge, UK) at 760 nm. The absorbance values were plotted against a standard curve obtained from gallic acid (y = 0.0132x + 0.0324; R² = 0.9987), and the results are given in gallic-acid equivalents.

Antioxidant activity assay

Aqueous leaf extracts (from light treatments W, W + B and W + UV) and the previously isolated major flavonoid in K. pinnata (quercetin 3-O-α-l-arabinopyranosyl (1[RIGHTWARDS ARROW]2) α-l-rhamnopyranoside) [43] antioxidant activities were measured using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) free-radical method [44]. This method is based on the reduction of this radical in the presence of an antioxidant molecule. This reaction is accompanied by a violet-to-yellow color change in the radical solution. The changes in sample color are proportional to the concentration of free-radical scavengers. This is a rapid and easy method to evaluate the antioxidant activity of a sample [44, 45].

Sample stock solutions (1 mg mL−1) were diluted to final concentrations of 250, 125, 50, 25, 10, 5, 2.5 and 0.5 μg mL−1 in methanol, and antioxidant activity was measured according to the method of Mensor et al. [45], with slight modifications. One milliliter of DPPH solution (0.1 mm in methanol; Sigma-Aldrich®) was added to the samples and the mixture was allowed to react at room temperature for 60 min in the dark. The absorbencies of blank (1 mL methanol and 2.5 mL samples) and negative control (2.5 mL methanol and 1 mL DPPH solution) were also measured at 518 nm, using a spectrophotometer (Libra S22; Biochrom Ltd).

The percentage of antioxidant activity (AA%) and the EC50 (concentration of the extract sufficient to obtain 50% of the total antioxidant activity) values were calculated. The EC50 was calculated with the Microsoft Excel® program. The following formula was used to calculate the AA%: AA% = 100 − {[(ABSsample − ABSblank) × 100]/ABSnegative control}

Thin-layer chromatography

Thin-layer chromatography (TLC) was carried out on silica gel 60 (F254, 0.25 mm; Merck) using a mobile phase consisting of n-butanol/acetic acid/water (BAW 8:1:1;v/v/v). Samples (100 μL) of two flavonoids previously isolated from K. pinnata leaves [32, 43] [quercetin 3-O-α-l-arabinopyranosyl (1[RIGHTWARDS ARROW]2) α-l-rhamnopyranoside and quercetin 3-O-α-rhamnopyranoside; quercitrin], both at 1 mg mL−1, were analyzed by TLC in comparison with the sample extracts (W, W + B and W + UV—100 μL at 20 mg mL−1). After development, plates were dried and the spots observed under UV light lamp (254 and 365 nm). The antioxidant spots were detected by a DPPH (Sigma-Aldrich®) methanol solution (2.54 mm) [46].

HPLC-DAD analysis

The extracts (W, W + B and W + UV) were analyzed using a HPLC Shimadzu SPD-M10A VP instrument with a diode array detector (DAD), using 200–400 nm as the wavelength range. An RP-18 reverse-phase column (5 μm, 250 mm × 4 mm, LichroCART®/Lichrospher®100; Merck) was used. The eluents were (1): H2O adjusted to pH 3.2 with phosphoric acid, and (2): CH3CN. The following solvent gradient (v/v) was applied: from H2O (pH 3.2)—CH3CN (10:0) to H2O (pH 3.2)—CH3CN (8:2) within 10 min; from H2O (pH 3.2)—CH3CN (8:2) to H2O (pH 3.2)—CH3CN (7.8:2.2) within 10 min; from H2O (pH 3.2)—CH3CN (7.8:2.2) to H2O (pH 3.2)—CH3CN (7.5:2.5) within 15 min; from H2O (pH 3.2)—CH3CN (7.5:2.5) to H2O (pH 3.2)—CH3CN (7:3) within 5 min; from H2O (pH 3.2)—CH3CN (7:3) to H2O (pH 3.2)—CH3CN (0:10); 45 min as total time of analysis.

Flow elution was 1 mL min−1, and 20 μL of each sample was injected. HPLC-DAD analysis was performed on each sample (W, W + B, and W + UV extracts) after dilution of 10 mg of the extract in 1 mL of deionized water. The previously isolated flavonoids: quercitrin (quercetin 3-O-α-rhamnopyranoside) and the major flavonoid in K. pinnata (quercetin 3-O-α-l-arabinopyranosyl (1[RIGHTWARDS ARROW]2) α-l-rhamnopyranoside) [32, 43] were used as controls in the extracts analyzed (dilution of 1 mg of these flavonoids in 1 mL of deionized water). Their retention time and ultraviolet spectra were checked in the chromatograms obtained for the extracts. The relative areas of the peaks in chromatograms of the extracts from leaves under the three different light treatments were compared.

Statistical analysis

Plants grown in each treatment (W, W + B and W + UV) were compared statistically, analyzing the following variables: leaf fresh weight (LFW), leaf-cake dry weight (LCDW), weight of lyophilized extracts (WLE), total phenolic content and antioxidant activity. The results are given as mean ± standard deviation. The statistical analysis was conducted with GraphPad Instat 3.0 for Windows®. The data were analyzed using a one-way ANOVA with Tukey's post test. Data normality and homogeneity of the variances were tested with Shapiro-Wilks and Levene tests, when necessary. Differences were considered statistically significant when P ≤ 0.05.

Results and discussion

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results and discussion
  6. Acknowledgements
  7. References

Leaf fresh weight, leaf-cake dry weight, and weight of lyophilized extracts

According to Muzitano et al. [33], hot-water extraction (50°C) of fresh leaves from K. pinnata is the best method to assess the content of active compounds.

We found no differences in leaf fresh weight (= 0.6954), relative leaf-cake dry weight (= 0.2424), or the relative weight of the lyophilized extracts (= 0.1238), according to the light-quality treatment (Table 2).

Table 2. Leaf fresh weight, weight of lyophilized extract and leaf-cake dry weight from leaves of Kalanchoe pinnata grown under three light treatments
 WW + BW + UV
  1. Results are given in mean ± standard deviation, = 3, P ≤ 0.05 (ANOVA). There were no statistical differences between the three light treatments in the parameters tested. W—White-light treatment; W + B—White plus blue-light treatment; W + UV—White plus UV-A-radiation treatment.

LFW (g)258.02 ± 13.43244.72 ± 53.45242.39 ± 36.66
WLE (g/g LFW)0.026 ± 0.0030.025 ± 0.0010.022 ± 0.001
LCDW (g/g LFW)0.060 ± 0.0010.062 ± 0.0080.054 ± 0.003

The lack of changes in these parameters indicates that neither supplemental blue light nor UV-A radiation affected the biomass of the leaves or the yield of the extract.

Total phenolic content

All the extracts tested contained phenolic compounds (gallic-acid equivalents): W = 59.02 ± 3.24; W + B = 58.07 ± 2.27; W + UV = 57.40 ± 4.04. After 60 days, we found no significant differences in the total phenolic contents of the extracts from leaves under the three different light treatments (= 0.8354, = 3).

In contrast to some species, including Vigna sinensis, Phaseolus vulgaris, Capsicum annuum and Amaranthus tricolor, in which blue light and UV-A radiation improve the content of phenolic compounds [18, 21, 47], these light qualities did not affect the total phenolic content of K. pinnata in this study. Compared to Ocimum basilicum, for example, blue light did not affect the total phenolic content after 7 days of treatment, although a decrease was observed after 14 days [48].

The effects of light quality on morphological and phytochemical production in plants are often diverse. The mixed results reported in different studies [49] seem to be species-dependent, with different results for different species cultivated in the same conditions [50, 51]. Furthermore, studies on the effects of light qualities on plants use different cultivation conditions, including in vitro cultures or growth chambers, different light sources (LEDs, emitting fluorescent tubes), different photoperiods and ages of leaves [16, 20, 49, 52]. All of these cultivation conditions could affect the plant phenolic production, varying the results.

Antioxidant activity

Antioxidant activity was detected in all the extracts. Supplementation with blue light improved the antioxidant activity compared with the other light treatments (= 0.0185). Similar results were found for barley, where supplemental blue light also improved the antioxidant activity of its plant extracts [14]. The W + UV treatment did not induce a significant difference in antioxidant activity compared with the white-light treatment (Fig. 2).

image

Figure 2. Antioxidant activity expressed in EC50 values (μg mL−1) of the extracts from Kalanchoe pinnata leaves grown under three different light treatments. W treatment—white column; W + B treatment—gray column; W + UV treatment—black column. Results given in mean ± standard deviation; = 3, P ≤ 0.05 (ANOVA). *Significant differences between the treatments.

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Some studies have reported a positive correlation between the increase in phenolic content and antioxidant activity [1, 13]. However, in our study, as well as for a Sedum species [12], another member of Crassulaceae and for other species [13], no such correlation was found.

Although correlations between the total phenolic content and antioxidant activity of a sample are commonly reported, these interpretations do not include a qualitative analysis of the mixture [1]. Differences in phenolic profile (type of phenolics present, and the relative amounts or proportions) could explain the changes in antioxidant activity found in this study.

The antioxidant activity of a phenolic compound depends on its proton-donating capacity, which is related to its structure. The efficiency of a phenolic compound as a free-radical scavenger can change according to the number of hydroxyl groups, their position on the molecule, the presence of glycosylation and the proximity of carboxylate and hydroxyl groups on the phenol ring [53, 54].

The antioxidant activity analysis of K. pinnata major flavonoid showed extremely low EC50 value (1.41 μg mL−1), highlighting its high antioxidant potential. Some flavonoid structural features are important for antioxidant and free-radical scavenging activities, including the following: an o-diphenolic group (in ring B), a 2–3 double bond conjugated with a 4-keto function, and hydroxyl groups in positions 3 and 5 [10, 55]. These structural features are present in the major flavonoid in K. pinnata, as well as in quercitrin (Fig. 3—bold regions), another flavonoid found in the species, which also shows high antioxidant activity [25, 32, 53].

image

Figure 3. Structures of Kalanchoe pinnata flavonoids. (A) quercetin 3-O-α-l-arabinopyranosyl (1[RIGHTWARDS ARROW]2) α-l-rhamnopyranoside, the major flavonoid; and (B) quercetin 3-O-α-rhamnopyranoside, quercitrin. Bold regions indicate important structural features for antioxidant and free-radical scavenging activities of flavonoids.

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Both glycosyl flavonoids have the flavonol quercetin as an aglycon skeleton. Quercetin has all the above features, and for this reason is a more effective antioxidant than other flavonols [10]. This could explain the high antioxidant activity of the major flavonoid in K. pinnata, as shown in this study, and quercitrin, as reported in literature data [10, 55].

Thin-layer chromatography

Bands with the DPPH scavenging activity on TLC plates are observed as whitish-yellow spots on a purple background [46]. TLC of the extracts revealed by DPPH solution showed a high profile of antioxidant constituents in all extracts analyzed, with some differences among aqueous extracts of leaves from plants grown under the three different light conditions (Fig. 4). All leaf extracts shared two bands (Rf 0.32 and 0.37) (Fig. 4a,b). The W + B leaf extract showed a compound with radical scavenger activity, which was observed only in this treatment (Rf 0.25) (Fig. 4c), and also showed more intense yellow staining immediately after being developed, which indicates its higher antioxidant profile. This result agrees with the results observed in the spectrophotometric DPPH assay. The major flavonoid (quercetin 3-O-α-l-arabinopyranosyl (1[RIGHTWARDS ARROW]2) α- l-rhamnopyranoside—Rf 0.50) (Fig. 4d) and quercitrin (quercetin 3-O-α-rhamnopyranoside—Rf 0.85) (Fig. 4e) were observed in all treatments.

image

Figure 4. Thin-layer chromatography (TLC) of extracts from Kalanchoe pinnata (silica gel 60 F254; BAW 8/1/1; DPPH solution reagent). 1—W treatment; 2—W + UV treatment; 3—W + B treatment; 4—major flavonoid; 5—quercitrin; a—Rf 0.32; b—Rf 0.37; c—Rf 0.25; d—Rf 0.50; e—Rf 0.85.

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Five flavonoids were described for K. pinnata leaves [25]. The main quercetin 3-O-α-l-arabinopyranosyl (1[RIGHTWARDS ARROW]2) α-l-rhamnopyranoside (major flavonoid) and quercetin 3-O-α-rhamnopyranoside (quercitrin) were found in extracts of all treatments tested. The additional flavonoids kaempferol 3-O-α-l-arabinopyranosyl (1[RIGHTWARDS ARROW]2)-α-l-rhamnopyranoside (kapinnatoside), kaempferol 3-O-α-rhamnopyranoside and 4′,5-dihydroxy-3′,8-dimethoxyflavone 7-O-β-d-glucopyranoside are present in very low concentrations in the aqueous extract [43]. Hence, they are very difficult to detect by the TLC technique.

HPLC-DAD analysis

HPLC-DAD provided information about the chromatographic profiles and composition of the extracts. Both the major flavonoid (retention time—tR = 30.68 min) and quercitrin (tR = 31.95 min) (Fig. 5) were detected in all extracts by this technique.

image

Figure 5. HPLC-DAD chromatograms (254 nm) and UV spectra of previously isolated flavonoids from Kalanchoe pinnata. (A) quercetin 3-O-α-l-arabinopyranosyl (1[RIGHTWARDS ARROW]2) α-l-rhamnopyranoside (major flavonoid) and (B) quercitrin.

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The major flavonoid and quercitrin belong to the subclass of flavonols. The UV spectra of flavonols exhibit two major absorption peaks in the region 240–400 nm, commonly referred to as Band I (300–380 nm) and Band II (240–280 nm). Band I is associated with the absorption of the B-ring cinnamoyl system of flavonoids, and Band II with absorption involving the A-ring benzoyl system. The position of Band I provides information about the flavonoid skeleton. Flavonols with a substituted 3-hydroxyl group (methylated or glycosylated) exhibit the band I in the region 328–357 nm, while this band is shifted to 352–385 nm in flavonols without substitution in this position [56]. This pattern is consistent with the UV spectra obtained for both flavonols analyzed.

The extracts exhibited a similar profile (Fig. 6), according to analysis of the retention time of the substances and the respective UV spectrum (200–400 nm). Nevertheless, comparison of the percentages of the heights of the peaks of the substances revealed that the diglycosylated flavonoid (major flavonoid) appeared in a slightly higher proportion in the W + B extract (24.7%; but only 22.6% in the W + UV extract and 19.6% in the W extract). The W + B chromatogram also showed more peaks in the region between 10 and 20 min, compared to the other experiments. The retention time and UV spectrum analysis suggest that these substances correspond to phenolic acids, metabolites with some well described biological activities [57], including antimicrobial [58] and antioxidant effects [10, 57]. Analysis by HPLC also showed that the W + B extract contained more peaks of substances detected compared to the W and W + UV extracts.

image

Figure 6. HPLC-DAD chromatograms of extracts of Kalanchoe pinnata cultivated under different light qualities. (A) W + B treatment, (B) W + UV treatment, (C) W treatment. The arrows indicate the UV profiles of some bioactive compounds: flavonoids and phenolic acids.

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The K. pinnata is rich in phenolic compounds, which account for some of the biological activities of the species [25]. The anti-inflammatory and wound-healing potential of this species may result from the antioxidant activity provided by these compounds [38]. Flavonoids and other polyphenols exhibit significant antioxidant activity, reducing the concentration of ROS that can initiate and also perpetuate inflammatory cascades and cause subsequent tissue damage [6, 38, 59].

In addition to their antioxidant activity, flavonoids and phenolic acids are important metabolites of K. pinnata, and can be involved in important biological activities of the species. Quercitrin and the major flavonoid, both detected using chromatography (TLC and HPLC) in all the extracts tested in this study, have in vitro and in vivo antileishmanial activity [25, 32, 43]. Therefore, their increases in K. pinnata extracts caused by supplementation of blue light are significant results, since they are the most important bioactive flavonoids in this species.

Phenolic acids form a diverse group and constitute about one-third of dietary phenols, which may be present in plants in free and bound forms [60]. This group includes the widely distributed hydroxybenzoic acids (λ max between 200 and 290 nm), such as gallic acid, as well as hydroxycinnamic acids (λ max between 270 and 360 nm), which are represented by coumaric, caffeic and ferulic acids, among others [61, 62]. Three phenolic acids were detected in K. pinnata extracts: gallic, caffeic and coumaric acids [62].

Although we did not find a direct correlation between the increase in antioxidant activity caused by supplemental blue light and the total phenolic content, we observed a change in the extract phenolic profile caused by this light-quality supplementation: higher proportions of quercetin 3-O-α-l-arabinopyranosyl (1[RIGHTWARDS ARROW]2) α-l-rhamnopyranoside, the presence of quercitrin, more types of phenolic acids and a more-complex mixture of substances. These changes could explain the higher antioxidant activity of this extract, compared to the control and supplementation with UV-A radiation.

Blue light can be proposed as a supplemental light source in cultivation of K. pinnata. The increase in the production of bioactive compounds, together with the higher antioxidant activity induced by this light-quality supplementation, are important results, improving some medicinal uses of the species, especially those related to its antioxidant activity.

Acknowledgements

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results and discussion
  6. Acknowledgements
  7. References

The authors express their gratitude to Ms. Gabriela de Souza, Ms. Claudia Lage, and Mr. Jorge Fernando Menezes for their contributions. The authors also thank Dr. Janet W. Reid and Mr. David Martin for the revision of the English text. This study was supported by FAPERJ and CAPES.

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results and discussion
  6. Acknowledgements
  7. References
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