Indução de calos e produção de compostos fenólicos bioativos em <i>Byrsonima verbascifolia</i> (L.) DC. (Malpighiaceae)

Castro, Ana Hortência Fonsêca; Braga, Karina de Queiroz; Sousa, Fernanda Marinez de; Coimbra, Mairon César; Chagas, Rafael César Russo

doi:10.5935/1806-6690.20160017

Ana Hortência Fonsêca Castro ^** Autor para correspondência

Programa de Pós-Graduação em Ciências Farmacêuticas, Laboratório de Farmacobotânica e Plantas Medicinais, Universidade Federal de São João Del-Rei, Campus Centro-Oeste, Av. Sebastião Gonçalves Coelho, 400, Chanandour, Divinópolis-MG, Brasil, 35.501-296, acastro@ufsj.edu.br

Karina de Queiroz Braga

Laboratório de Farmacobotânica e Plantas Medicinais, Universidade Federal de São João Del-Rei, Campus Centro-Oeste, Av. Sebastião Gonçalves Coelho, 400, Chanandour, Divinópolis-MG, Brasil, 35.501-296, karinaqbraga@yahoo.com.br, fernandamarinez@gmail.com, maironcoimbra@gmail.com

Fernanda Marinez de Sousa

Laboratório de Farmacobotânica e Plantas Medicinais, Universidade Federal de São João Del-Rei, Campus Centro-Oeste, Av. Sebastião Gonçalves Coelho, 400, Chanandour, Divinópolis-MG, Brasil, 35.501-296, karinaqbraga@yahoo.com.br, fernandamarinez@gmail.com, maironcoimbra@gmail.com

Mairon César Coimbra

Laboratório de Farmacobotânica e Plantas Medicinais, Universidade Federal de São João Del-Rei, Campus Centro-Oeste, Av. Sebastião Gonçalves Coelho, 400, Chanandour, Divinópolis-MG, Brasil, 35.501-296, karinaqbraga@yahoo.com.br, fernandamarinez@gmail.com, maironcoimbra@gmail.com

Rafael César Russo Chagas

Programa de Pós-Graduação em Biotecnologia, Universidade Federal de São João Del-Rei, Campus Centro-Oeste, Av. Sebastião Gonçalves Coelho, 400, Chanandour, Divinópolis-MG, Brasil, 35.501-296, rafaelchagas@ufsj.edu.br

* Autor para correspondência

Figuras (2)
Tabelas (3)

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Figure 1
Total phenols (A), flavonoids (B) and tannins (C) contents in B. verbascifolia’s callus. The bar represents the mean􀁲standard deviation of 3 observations. Legend: 1) 4.52 µM 2,4-D; 2) 9.05 µM 2,4-D; 3) 18.10 µM 2,4-D; 4) 4.44 µM BAP; 5) 8.88 µM BAP; 6) 17.75 µM BAP; 7) 4.52 µM 2,4-D + 4.44 µM BAP; 8) 4.52 µM 2,4-D + 8.88 µM BAP; 9) 4.52 µM 2,4-D + 17.75 µM BAP; 10) 9.05 µM 2,4-D + 4.44 µM BAP; 11) 9.05 µM 2,4-D + 8.88 µM BAP; 12) 9.05 µM 2,4-D + 17.75 µM BAP; 13) 18,10 µM 2,4-D + 4.44 µM BAP; 14) 18.10 µM 2,4-D + 8.88 µM BAP; 15) 18.10 µM 2,4-D +17.75 µM BAP; IE = initial explant

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Figure 2
Chromatograms obtained by HPLC-DAD analysis of hydromethanolic extracts by detection at 254 nm. A. Chromatographic profile of the Initial Explant - Peak 1 = benzoic acid derivative, Peaks 2-3 = p-coumaric acid derivative. B. Chromatographic profile of callus from treatment 6 (17.75 µM BAP) in the presence of light - Peaks 1-3 = benzoic acid derivatives, Peaks 4-6 = catechins, Peaks 7-8 = m-coumaric acid derivatives. C. Chromatographic profile of callus from treatment 6 (17.75 µM BAP) in the absence of light - Peaks 1-3 = benzoic acid derivatives, Peaks 4-7 = gallic acid derivatives, Peaks 8-12 = catechins, Peaks 13-15 = m-coumaric acid derivatives. D. Chromatographic profile of callus from treatment 7 (4.52 µM 2,4-D + 4.44 µM BAP) in the absence of light - Peaks 1-2 = cinnamic acid derivatives, Peak 3 = benzoic acid derivative, Peak 4 = gallic acid derivative, Peak 5 = benzoic acid derivative, Peaks 6-7 = gallic acid derivatives, Peaks 8-9 = benzoic acid derivatives, Peaks 10-11 = p-coumaric acid derivatives

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Table 1
Induction, fresh and dry weight, color and consistency of Byrsonima verbascifolia’s callus, in the presence of light after 45 days cultivation

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Table 2
Induction, fresh and dry weight, color and consistency of B. verbascifolia’s callus, in the absence of light after 45 days cultivation

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Table 3
Phenolic compounds in hydromethanolic extracts in Byrsonima verbascifolia’s callus using HPLC-DAD

Treatment	Growth regulators	Induction (%)	Fresh weight (g)	Dry weight (g)	Color	Consistency
Control	Absence of regulator	-	-	-	-	-
1	4.52 μM 2,4-D	53d	0.71 ± 0.09 b	0.07 ± 0.01 b	Yellow/Red	Friable
2	9.05 μM 2,4-D	87b	0.77 ± 0.08 b	0.04^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Yellow/Red	Friable
3	18.10 μM 2,4-D	68c	0.70 ± 0.07 b	0.04^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Yellow/Red	Friable
4	4.44 μM BAP	7f	0.47 ± 0.09 c	0.03^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Green/Red	Compact
5	8.88 μM BAP	35e	0.48 ± 0.10 c	0.08 ± 0.01 b	Green/Red	Compact
6	17.75 μM BAP	94b	0.21 ± 0.05 d	0.04^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Green/Red	Compact
7	4.52 μM 2,4-D + 4.44 μM BAP	100a	1.77 ± 0.09 a	0.17 ± 0.04 a	Green	Compact
8	4.52 μM 2,4-D + 8.88 μM BAP	100a	1.58 ± 0.12 a	0.15 ± 0.05 a	Green/Red	Compact
9	4.52 μM 2,4-D + 17.75 μM BAP	100a	0.69 ± 0.07 b	0.07 ± 0.01 b	Red	Compact
10	9.05 μM 2,4-D + 4.44 μM BAP	100a	1.55 ± 0.15 a	0.14 ± 0.05 a	Green/Red	Compact
11	9.05 μM 2,4-D + 8.88 μM BAP	100a	0.92 ± 0.20 b	0.07 ± 0.01 b	Green	Compact
12	9.05 μM 2,4-D + 17.75 μM BAP	100a	0.99 ± 0.21 b	0.09 ± 0.02 b	Green	Compact
13	18,10 μM 2,4-D + 4.44 μM BAP	100a	0.77 ± 0.10 b	0.08 ± 0.02 b	Green/Red	Compact
14	18.10 μM 2,4-D + 8.88 μM BAP	100a	1.02 ± 0.25 b	0.08 ± 0.01 b	Green/Red	Compact
15	18.10 μM 2,4-D +17.75 μM BAP	100a	0.28 ± 0.06 d	0.03^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Green/Red	Compact
F		9.456^* * Significant value at 5% level by F test; means in the column followed by the same letter are not significantly different at p ≤ 0.05 by the Tukey test;	5.227^* * Significant value at 5% level by F test; means in the column followed by the same letter are not significantly different at p ≤ 0.05 by the Tukey test;	7.253^* * Significant value at 5% level by F test; means in the column followed by the same letter are not significantly different at p ≤ 0.05 by the Tukey test;

Treatment	Growth regulators	Induction (%)	Fresh weight (g)	Dry weight (g)	Color	Consistency
Control	Absence of regulator	-	-	-	-	-
1	4.52 μM 2,4-D	71 c	0.68 ± 0.16 b	0.06 ± 0.01 b	Yellow	Friable
2	9.05 μM 2,4-D	73 c	0.85 b ± 0.11	0.04^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Yellow/Red	Friable
3	18.10 μM 2,4-D	82 b	0.34 ± 0.05 c	0.03^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Yellow	Friable
4	4.44 μM BAP	53 d	0.15 ± 0.05 d	0.02^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Yellow/Red	Compact
5	8.88 μM BAP	74 c	0.19 ± 0.04 d	0.03^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Yellow/Red	Compact
6	17.75 μM BAP	59 d	0.32 ± 0.07 c	0.04^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Yellow	Compact
7	4.52 μM 2,4-D + 4.44 μM BAP	100 a	1.66 ± 0.18 a	0.15 ± 0.05 a	Yellow	Friable
8	4.52 μM 2,4-D + 8.88 μM BAP	100 a	1.49 ± 0.15 a	0.14 ± 0.05 a	Yellow/Red	Compact
9	4.52 μM 2,4-D + 17.75 μM BAP	100 a	0.88 ± 0.10 b	0.06 ± 0.01 b	Yellow/Red	Compact
10	9.05 μM 2,4-D + 4.44 μM BAP	100 a	0.81 ± 0.09 b	0.04^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Yellow/Red	Compact
11	9.05 μM 2,4-D + 8.88 μM BAP	100 a	1.58 ± 0.10 a	0.15 ± 0.05 a	Yellow	Compact
12	9.05 μM 2,4-D + 17.75 μM BAP	100 a	1.43 ± 0.10 a	0.13 ± 0.02 a	Yellow	Compact
13	18,10 μM 2,4-D + 4.44 μM BAP	100 a	0.84 ± 0.05 b	0.07 ± 0.01 b	Yellow	Compact
14	18.10 μM 2,4-D + 8.88 μM BAP	100 a	0.87 ± 0.09 b	0.05 ± 0.01 c	Yellow/Red	Compact
15	18.10 μM 2,4-D +17.75 μM BAP	100 a	0.34 ± 0.05 c	0.04^ Standard deviation values for these data were omitted, since they are less than 0.009. Legend: - (no callus induction) c	Yellow/Red	Compact
F		10.778^* * Significant value at 5% level by F test; means in the column followed by the same letter are not significantly different at p ≤ 0.05 by the Tukey test;	6.534^* * Significant value at 5% level by F test; means in the column followed by the same letter are not significantly different at p ≤ 0.05 by the Tukey test;	8.123^* * Significant value at 5% level by F test; means in the column followed by the same letter are not significantly different at p ≤ 0.05 by the Tukey test;

Sample	Peak number	Rt (min.)	λmax (nm)	Compound	Reference
Initial explant	1	3.491	264	Benzoic acid derivative	Maatta; Kamal-Eldin; Torronen (2003)MÄÄTTÄ, K. R.; KAMAL-ELDIN, A.; TÖRRÖNEN, A. R. High-performance liquid chromatography (HPLC) analysis of phenolic compounds in berries with diode array and electrospray ionization mass spectrometric (MS) detection: ribes species. Journal of Agricultural and Food Chemistry, v. 51, n. 23, p. 6736-6744, 2003.
	2	21.037	326	p-cumaric acid derivative	*Sakakibara et al.* (2003)**SAKAKIBARA, H. et al. Simultaneous determination of all polyphenols in vegetables, fruits, and teas. Journal of Agricultural and Food Chemistry, v. 51, n. 3, p. 571-581, 2003.
	3	23.264	326	p-cumaric acid derivative
Treatment 6 (presence of light)	1	3.506	265	Benzoic acid derivative	*Sakakibara et al.* (2003)**SAKAKIBARA, H. et al. Simultaneous determination of all polyphenols in vegetables, fruits, and teas. Journal of Agricultural and Food Chemistry, v. 51, n. 3, p. 571-581, 2003.
	2	10.433	258
	3	13.146	258
	4	17.780	279	Cathechin	Reference substance; *Sakakibara et al.* (2003)**SAKAKIBARA, H. et al. Simultaneous determination of all polyphenols in vegetables, fruits, and teas. Journal of Agricultural and Food Chemistry, v. 51, n. 3, p. 571-581, 2003.
	5	18.393	279
	6	19.125	279
	7	22.149	270	m-cumaric acid derivative	*Sakakibara et al.* (2003)**SAKAKIBARA, H. et al. Simultaneous determination of all polyphenols in vegetables, fruits, and teas. Journal of Agricultural and Food Chemistry, v. 51, n. 3, p. 571-581, 2003.
	8	23.226	270	m-cumaric acid derivative
Treatment 6 (absence of light)	1	2.163	274	Benzoic acid derivative	*Sakakibara et al.* (2003)**SAKAKIBARA, H. et al. Simultaneous determination of all polyphenols in vegetables, fruits, and teas. Journal of Agricultural and Food Chemistry, v. 51, n. 3, p. 571-581, 2003.
	2	3.509	272
	3	4.096	259
	4	4.780	262	Gallic acid derivative	*Abad-Garcia et al.* (2009)**ABAD-GARCÍA, B. et al.A general analytical strategy for the characterization of phenolic compounds in fruit juices by high-performance liquid chromatography with diode array detection coupled to electrospray ionization and triple quadrupole mass spectrometry. Journal of Chromatography A, v. 1216, n. 28, p. 5398-5415, 2009.
	5	5.055	262
	6	10.462	261
	7	13.164	257
	8	14.721	278	Cathechin	Reference substance; *Sakakibara et al.* (2003)**SAKAKIBARA, H. et al. Simultaneous determination of all polyphenols in vegetables, fruits, and teas. Journal of Agricultural and Food Chemistry, v. 51, n. 3, p. 571-581, 2003.
	9	17.351	279
	10	17.791	279
	11	18.384	279
	12	19.122	279
	13	22.127	273	m-cumaric acid derivative	*Sakakibara et al.* (2003)**SAKAKIBARA, H. et al. Simultaneous determination of all polyphenols in vegetables, fruits, and teas. Journal of Agricultural and Food Chemistry, v. 51, n. 3, p. 571-581, 2003.
	14	22.313	273
	15	23.195	273
Treatment 7 (absence of light)	1	2.225	277	Cinamic acid derivative	*Sakakibara et al.* (2003)**SAKAKIBARA, H. et al. Simultaneous determination of all polyphenols in vegetables, fruits, and teas. Journal of Agricultural and Food Chemistry, v. 51, n. 3, p. 571-581, 2003.
	2	2.878	278	Cinamic acid derivative
	3	3.477	269	Benzoic acid derivative
	4	5.051	261	Gallic acid derivative
	5	5.761	271	Benzoic acid derivative
	6	8.303	260	Gallic acid derivative	*Sakakibara et al.* (2003)**SAKAKIBARA, H. et al. Simultaneous determination of all polyphenols in vegetables, fruits, and teas. Journal of Agricultural and Food Chemistry, v. 51, n. 3, p. 571-581, 2003.
	7	10.426	261	Gallic acid derivative
	8	12.899	255	Benzoic acid derivative	*Sakakibara et al.* (2003)**SAKAKIBARA, H. et al. Simultaneous determination of all polyphenols in vegetables, fruits, and teas. Journal of Agricultural and Food Chemistry, v. 51, n. 3, p. 571-581, 2003.
	9	13.092	258	Benzoic acid derivative
	10	23.256	327	p-cumaric acid derivative
	11	29.490	308	p-cumaric acid derivative

[1] * Autor para correspondência

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Indução de calos e produção de compostos fenólicos bioativos em Byrsonima verbascifolia (L.) DC. (Malpighiaceae)

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