Optimization of secoisolariciresinol diglucoside extraction from flaxseed (Linum usitatissimum L.) and isolation by a simple HPLC-UV method

Figures & data

Table 1. Flaxseed lignan extraction methods.

Tabla 1. Métodos de extracción de lignanos de linaza.

Extraction methods	Concentration	Temperature	Time	Reference
Alcoholic extraction	1,4-dioxane/95% ethanol	60°C	16 h	Johnsson et al. (2000)
Alkaline hydrolysis	0.3 mol L^−1 NaOH	Environment	48 h
Direct alkaline hydrolysis	1 NaOH	20°C	1 h	Eliasson et al. (2003)
Precipitation	60% ethanol
Alcoholic extraction	70% methanol	Environment	4 h	Li, Yuan, Xu, Wang, and Liu (2008)
Alkaline hydrolysis	20 mmol L^−1 NaOH	50°C	15 min
Alcoholic extraction	60% methanol	40°C	1 h	Chen et al. (2007)
Alkaline hydrolysis	1 mol L^−1 NaOH
Alcoholic extraction	1,4-dioxane/ethanol	40°C	2 h	Coran, Giannellini, and Bambagiotti-Alberti (2004)
Alkaline hydrolysis	0.1 mol L^−1 NaOH
Alcoholic extraction	70% ethanol	40°C	28 h	Zhang et al. (2007)
Alkaline hydrolysis	1 mol L^−1 NaOH	Environment	12 h
Microwave-assisted extraction	70% methanol	50 W	3 min	Beejmohun et al. (2007)
	1 mol L^−1 NaOH
Microwave-assisted extraction	40.9% ethanol	130 W	90.5 s	Zhang and Xu (2007)
Alkaline hydrolysis	0.25 mol L^−1 NaOH	25°C	2 h
Alcoholic extraction	70% methanol	Environment	2 h	Degenhardt, Habben, and Winterhalter (2002)
Alkaline hydrolysis	1 mol L^−1 NaOH		3 h

Figure 1. HPLC chromatogram of lignan SDG and other compounds extracted from flaxseed recorded at 280 nm. The injection volume was 100 µL and the mobile phase consisted of 1% aqueous acetic acid/acetonitrile (85:15 v/v) with a flow rate of 1 mL min⁻¹.

Figura 1. Cromatograma HPLC del lignano SDG y otros compuestos extraídos desde linaza registrado a 280 nm. El volumen de inyección fue 100 µL y la fase móvil consistió de 1% de ácido acético acuoso/acetonitrilo (85:15 v/v) con flujo de 1 mL min⁻¹.

Figure 2. Enzymatic hydrolysis with β-glucosidase to produce SECO from SDG.

Figura 2. Hidrolisis enzimatica con β-glucosidasa para producir SECO desde SDG.

Figure 3. HPLC chromatogram for the enzymatic test recorded at 280 nm. The injection volume was 20 µL and the mobile phase consisted of 1% aqueous acetic acid/acetonitrile (65:35 v/v) with a flow rate of 1 mL min⁻¹.

Figura 3. Cromatograma HPLC para la confirmación enzimática registrado a 280 nm. El volumen de inyección fue 20 µL y la fase móvil consistió de 1% de ácido acético acuoso/acetonitrilo (65:35 v/v) con flujo de 1 mL min⁻¹.

Figure 4. HPLC chromatogram of lignan SDG purified by collecting fractions, recorded at 280 nm. The injection volume was 20 µL and the mobile phase consisted of 1% aqueous acetic acid/acetonitrile (85:15 v/v) with a flow rate of 1 mL min⁻¹.

Figura 4. Cromatograma HPLC del lignano SDG purificado a través de recolector de fracciones registrado a 280 nm. El volumen de inyección fue 20 µL y la fase móvil consistió de 1% de ácido acético acuoso/acetonitrilo (85:15 v/v) con flujo de 1 mL min⁻¹.

Table 2. Box–Behnken design and its results.

Tabla 2. Diseño Box–Behnken y sus resultados.

Experimental run	Sodium methoxide concentration (mmol L^−1)	Temperature (ºC)	Time (h)	SDG yield (mg g^−1 whole flaxseed)
1	50	40	18	8.455
2	150	40	18	7.351
3	50	60	18	7.702
4	150	60	18	7.085
5	50	50	12	8.910
6	150	50	12	8.497
7	50	50	24	9.100
8	150	50	24	7.602
9	100	40	12	8.844
10	100	60	12	6.994
11	100	40	24	9.109
12	100	60	24	7.682
13	100	50	18	8.562
14	100	50	18	9.211
15	100	50	18	9.031

Table 3. Analysis of variance for the fitted model.

Tabla 3. Análisis de varianza para el modelo ajustado.

Source	Sum of squares	Degrees of freedom	Mean squares	F-value	P-value
X1 (concentration, mmol L^−1)	1.6489	1	1.6489	14.69	0.0618
X2 (temperature, °C)	2.3070	1	2.3070	20.55	0.0454
X3 (Time, min)	0.0077	1	0.0077	0.07	0.8180
	0.7752	1	0.7752	6.91	0.1194
	0.0593	1	0.0593	0.53	0.5429
	0.2943	1	0.2943	2.62	0.2468
X1 X2	2.5327	1	2.5327	22.56	0.0416
X1 X3	0.0447	1	0.0447	0.40	0.5924
X2 X3	0.0095	1	0.0095	0.08	0.7983
Lack of fit	0.9855	3	0.3285	2.93	0.2650
Pure error	0.2245	2	0.1123
Total (corr.)	8.7586	14
R^2 = 0.8619	Standard error of Est. = 0.4919		Durbin–Watson statistic = 2.1887 (p = 0.4205)
Adjusted R^2 = 0.6132	Mean absolute error = 0.2555

Figure 5. Estimated response surface after 18 h of the extraction process of flaxseed lignans.

Figura 5. Superficie respuesta estimada de extracción de lignanos de linaza con 18 h de tiempo de reacción.

Figure 6. SDG content of flaxseed in four varieties according to their location of cultivation (Cajón and Gorbea, harvested in 2009) and year of harvest (2008). Bar graphs with the same letter are not significantly different (p < 0.05).

Figura 6. Contenido de SDG de linaza en cuatro variedades de acuerdo a su localidad de cultivo (Cajón y Gorbea, cosechada en 2009) y año de cosecha (2008). Gráficos de barras con la misma letra no muestra diferencias significativas (p < 0.05).

Johnsson, P., Kamal-Eldin, A., Lundgren, L. N., & Åman, P. (2000). HPLC method for analysis of secoisolariciresinol diglucoside in flaxseeds. Journal of Agricultural and Food Chemistry, 48(11), 5216–5219. doi:10.1021/jf0005871

 PubMed Web of Science ®Google Scholar

Eliasson, C., Kamal-Eldin, A., Andersson, R., & Åman, P. (2003). High-performance liquid chromatographic analysis of secoisolariciresinol diglucoside and hydroxycinnamic acid glucosides in flaxseed by alkaline extraction. Journal of Chromatography A, 1012(2), 151–159. doi:10.1016/s0021-9673(03)01136-1

 PubMed Web of Science ®Google Scholar

Li, X., Yuan, J.-P., Xu, S.-P., Wang, J.-H., & Liu, X. (2008). Separation and determination of secoisolariciresinol diglucoside oligomers and their hydrolysates in the flaxseed extract by high-performance liquid chromatography. Journal of Chromatography A, 1185(2), 223–232. doi:10.1016/j.chroma.2008.01.066

 PubMed Web of Science ®Google Scholar

Chen, J., Liu, X., Shi, Y. P., & Ma, C. Y. (2007). Determination of the lignan secoisolariciresinol diglucoside from flaxseed (Linum usitatissimum L.) by HPLC. Journal of Liquid Chromatography & Related Technologies, 30(4), 533–544. doi:10.1080/10826070601093853

 Web of Science ®Google Scholar

Coran, S. A., Giannellini, V., & Bambagiotti-Alberti, M. (2004). High-performance thin-layer chromatographic-densitometric determination of secoisolariciresinol diglucoside in flaxseed. Journal of Chromatography A, 1045(1–2), 217–222. doi:10.1016/j.chroma.2004.06.042

 PubMed Web of Science ®Google Scholar

Zhang, Z. S., Li, D., Wang, L. J., Ozkan, N., Chen, X. D., Mao, Z. H., & Yang, H. Z. (2007). Optimization of ethanol-water extraction of lignans from flaxseed. Separation and Purification Technology, 57(1), 17–24. doi:10.1016/j.seppur.2007.03.006

 Web of Science ®Google Scholar

Beejmohun, V., Fliniaux, O., Grand, É., Lamblin, F., Bensaddek, L., Christen, P. … Mesnard, F. (2007). Microwave-assisted extraction of the main phenolic compounds in flaxseed. Phytochemical Analysis, 18(4), 275–282. doi:10.1002/pca.973

 PubMed Web of Science ®Google Scholar

Zhang, W. B., & Xu, S. Y. (2007). Microwave-assisted extraction of secoisolariciresinol diglucoside from flaxseed hull. Journal of the Science of Food and Agriculture, 87(8), 1455–1462. doi:10.1002/jsfa.2793

 Web of Science ®Google Scholar

Degenhardt, A., Habben, S., & Winterhalter, P. (2002). Isolation of the lignan secoisolariciresinol diglucoside from flaxseed (Linum usitatissimum L.) by high-speed counter-current chromatography. Journal of Chromatography A, 943(2), 299–302. doi:10.1016/S0021-9673(01)01467-4

 PubMed Web of Science ®Google Scholar