High efficient column chromatographic extraction of curcumin from curcuma longa

High efficient column chromatographic extraction of curcumin from curcuma longa High efficient column chromatographic extraction of curcumin from curcuma longa High efficient column chromatographic extraction of curcumin from curcuma longa High efficient column chromatographic extraction of curcumin from curcuma longa High efficient column chromatographic extraction of curcumin from curcuma longa High efficient column chromatographic extraction of curcumin from curcuma longa High efficient column chromatographic extraction of curcumin from curcuma longa High efficient column chromatographic extraction of curcumin from curcuma longa High efficient column chromatographic extraction of curcumin from curcuma longa High efficient column chromatographic extraction of curcumin from curcuma longa High efficient column chromatographic extraction of curcumin from curcuma longa High efficient column chromatographic extraction of curcumin from curcuma longa

Short communication

High-efficient column chromatographic extraction of curcumin from Curcuma longa Pei-Yin Zhana, Xue-Hua Zenga, He-Ming Zhangb,⇑, Hai-Hang Lia,⇑

a

Guangdong Provincial Key Laboratory of Biotechnology for Plant Development, College of Life Sciences, South China Normal University, Guangzhou 510631, China

b Key Laboratory of China State Administration of Traditional Medicine for Chinese Medicine and Photonics Technologies, Institute of Traditional and Innovative Medicines, South China Normal University, Guangzhou 510631, China

a r t i c l e i n f o

Article history:

Received 16 December 2010

Received in revised form 20 February 2011

Accepted 22 April 2011

Available online 30 April 2011

Keywords:

Column chromatographic extraction

Curcumin

Turmeric

Curcuma longa

a b s t r a c t

Curcumin is an important food additive and a potential therapeutic agent for various diseases from tur-meric, the rhizome of Curcuma longa L High-efficient column chromatographic extraction (CCE) proce-dures were developed for the extraction of curcumin from turmeric Turmeric powder was loaded into

a column with 2-fold 80% ethanol The column was eluted with 80% ethanol at room temperature For quantitative analysis with a non-cyclic CCE, 8-fold eluent was collected as extraction solution For large preparation with a cyclic CCE, only the first 2-fold of eluent was collected as extraction and other eluent was sequentially circulated to the next columns More than 99% extraction rates were obtained through both CCE procedures, compared to a 59% extraction rate by the ultrasonic-assisted maceration extraction with 10-fold 80% ethanol The CCE procedures are high-efficient for the extraction of curcumin from tur-meric with minimum use of solvent and high concentration of extraction solution

Ó 2011 Elsevier Ltd All rights reserved

1 Introduction

Curcumin is one of the important food additives for spice and

natural pigment, from turmeric, the rhizomes of Curcuma longa L

(Green et al., 2008) Recently, it has been shown to possess various

pharmaceutical functions, such as anti-inflammatory (Jurenka,

2009), anticancer (Fu & Kurzrock, 2010; Ravindran, Prasad, &

Aggarwal, 2009), anti-ageing (Sikora, Scapagnini, & Barbagallo,

2010), neuro-protection in Alzheimer’s disease (Bandyopadhyay,

Huang, Lahiri, & Rogers, 2010; Kulkarni & Dhir, 2010) and many

other functions (Heng, 2010; Zhou, Beevers, & Huang, 2011) It

has received considerable interest as a potential therapeutic agent

for the prevention of various diseases (Zhou et al., 2011)

Extraction of curcumin from plant materials are mainly based

on the maceration method with the combination of circulation,

ultrasonic, microwave, heating, pressure or enzyme treatment

(Braga, Leal, Carvalho, & Meireles, 2003; Green et al., 2008; Mandal,

Dewanjee, Sahu, & Mandal, 2009; Mandal, Mohan, & Hemalatha,

2008; Manzan, Toniolo, Bredow, & Povh, 2003) These methods

re-quire long extraction times, high energy consumption and large

volume of organic solvent, have low extraction efficiency and are

unsafe for thermo-sensitive substances (Green et al., 2008)

We have reported a high-efficient column chromatographic

extraction (CCE) procedure for the extraction of compounds from

biological materials (Ni, Zhou, Li, & Huang, 2009) The CCE method,

based on chromatographic theory and practice, combined tradi-tional and recent new technologies in the extraction field, such

as maceration, leaching, dynamic and circulation Target sub-stances in biological materials are dissolved and eluted in columns with minimum volume of solvent Using this method, curcumin was completely extracted from turmeric materials with only 2-fold (through cyclic CCE) or 8-fold (through non-cyclic CCE) 80% etha-nol at room temperature The highly concentrated extraction solu-tion can be easily concentrated with low energy cost

2 Materials and methods 2.1 Materials

Turmeric, dried rhizomes of C longa L., is products of Guizhou Province, China, and is purchased from Guangzhou market of Chinese medicine The dried turmeric was ground and sieved; material between 833 and 350lm was used for experiments Cur-cumin standard compound was purchased from Sigma company (St Louis, MO, USA) HPLC-grade solvents were purchased from Burdick & Jackson Inc (Muskegon, MI, USA) Food-grade of 95% ethanol was used in all extraction experiments Other

analytical-or biochemical-grade analytical-organic solvents and chemical reagents were purchased from local suppliers

2.2 Extraction methods For ultrasonic-assisted macerating extraction as a control for traditional extraction methods, 10 g of turmeric powder was

0308-8146/$ - see front matter Ó 2011 Elsevier Ltd All rights reserved.

⇑Corresponding authors Tel./fax: +86 20 8521 7701 (H.-M Zhang), tel./fax: +86

20 8521 2630 (H.-H Li).

E-mail addresses: d_zhm@163.com (H.-M Zhang), li_haihang@yahoo.com (H.-H.

Li).

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j o u r n a l h o m e p a g e : w w w e l s e v i e r c o m / l o c a t e / f o o d c h e m

extracted with a 10-fold excess volume of solvents (w/v) with

100-W ultrasonic treatment for 1 h, unless otherwise stated The

extraction solution was centrifuged at 5000 g for 10 min, filtered

through a 0.45-lm membrane filter and analysed by HPLC

CCE procedure was performed in glass chromatographic

columns as described previously (Ni et al., 2009) Given amounts

of solvents were loaded into a column (1.5 cm diameter) Turmeric

powder (20 g) was then gradually added and was allowed to sink

freely down into the solvent A total of 2-fold volume of solvent,

based on the weight of the turmeric powder (v/w), was then loaded

onto the column After 1 h, the column was eluted by common

procedures at a flow rate of 2 bed volumes (BVs)/h, and eluent

were collected in fractions, each with a solvent volume equal to

two times the sample weight (w/v) Curcumin content in each

fraction were then analyzed by HPLC

For cyclic CCE, only the first fraction of eluent was collected as

the final extraction solution The second fraction was used to

extract the next material, and the third fraction was used for

eluting the next column Through the cyclic CCE method, the total

volume of final extracting solution was only two times the weight

of the dry material (v/w), while the columns were eluted three

times

2.3 Determination of curcumin

HPLC determination of curcumin is based on our previous

report (Wu, Ni, Li, & Li, 2008) with some modification A Shimadzu

SPD-20A HPLC system (Shimadzu, Japan) with LC-20AT UV

detec-tor, YMC-packed ODS column (250 mm  4.6 mm, 5lm) and

ana-lytical software was used for the analysis of curcumin The mobile

phase was acetonitrile: 5% acetic acid aqueous solution (50:50, v/ v) UV detection wavelength is 425 nm All samples were filtered through 0.45-lm membrane filters before injection into the HPLC The curcumin peak in samples were identified by its retention time and co-injection test with standard curcumin (Fig 1A) Quantita-tive analysis of curcumin was performed using the peak area based

on the standard curve

All experiments were repeated at least three times, and the results are given as the mean of three independent experi-ments ± standard error

3 Results and discussion The CCE method consists of two steps, dissolving target compounds into the extraction solvent and eluting them from the material in a chromatographic column The key point for the first step is to find a good solvent to dissolve target compounds (Ni et al., 2009)

Using the traditional macerating method, the extraction efficiencies of different concentrations of ethanol was tested As shown inFig 1B, the extraction rates of curcumin increased with ethanol concentrations from 40% to 80%, with the highest extrac-tion at 80%, but decreased when the ethanol concentraextrac-tion was higher than 80% The best solvent for dissolving curcumin from turmeric was 80% ethanol

To determine a minimum volume of solvent for the extraction

of curcumin from turmeric, five-fold excesses of 80% ethanol was added to and mixed with the turmeric powder, the remained solvent (not absorbed by the material) was determined each

15 min after start As shown inFig 1C, turmeric material was fully

Fig 1 Conditions for the extraction of curcumin from turmeric A: Identification of curcumin peak in HPLC spectrum of extracted sample by co-injection test with standard compound Three peaks from left to right in the sample’s HPLC spectrum are bisdemethoxycurcumin, demethoxycurcumin and curcumin, respectively B: Extraction of curcumin from turmeric with different concentrations of ethanol C: Absorbance of the solvent by turmeric powder D: Effect of macerating hours on the extraction of

imbibed in 80% ethanol within 30 min, and it absorbed a maximum

volume of 1.2-fold its own weight For operating purpose, we use

2-fold volume of solvent as minimum volume for extraction of

curcumin from turmeric

Fig 1D shows the effect of pre-macerating time on the

dissolv-ing of curcumin in 80% ethanol Dissolution of curcumin from

turmeric into solvent reached a dynamic balance within 1 h

Based on the results above, the conditions for extraction of

curcumin from turmeric by CCE procedure was determined as

follows Turmeric material was loaded with 2-fold of 80% ethanol

into a column at a height-to-diameter (H/D) ratio of 10:1,

pre-dissolved target substances for 1 h and then eluted the column

with 80% ethanol at a flow rate of 2 BVs/h, all at room temperature

Eluent was collected in fractions each with 2-fold volume of the

material weight and analyzed by HPLC

As shown inFig 2A, curcumin was completely eluted from the

column within the first 4 fractions (8-fold volume of material

weight) of eluent After then, curcumin could not be detected by

HPLC in the following fractions Also, no curcumin could be

extracted from the material in the column by the ultrasonic-assisted

maceration extraction

To low down the extraction solution and increase the

concentra-tion of the target compounds in large and continuous producconcentra-tion of

extracts, a cyclic CCE procedure was tested In this procedure, only

the first fraction of 2-fold eluent, which contained more than 80% of

curcumin, was collected as extraction, the second and third

frac-tions were circulated sequentially to the next column As shown

inFig 2B (black bar) andTable 1, the extraction rate of curcumin

reached more than 99% This result indicate that curcumin was

completely extracted from the material through the cyclic CCE

procedure with only 2-fold 80% ethanol which was circulated three times among different columns from low to high contents of curcu-min The cyclic chromatographic extraction reduced extensively the volumes of the solvent and the final extraction solution with high extraction efficiency Similar results were obtained when the scale of extracting experiment was enlarged 100 times in the cyclic CCE procedure

Based on the results above, curcumin can be effectively extracted by the CCE or cyclic CCE method in simple columns at room temperature, with much higher extraction rates than the ultrasonic-assisted macerating extraction (Table 1) The CCE procedure, which used 8-fold solvent for complete extraction of curcumin, is good for non-continuous extraction of samples in quantitative analysis, while the cyclic CCE procedure has multiple advantages for large scale and continuous extraction of curcumin from turmeric materials

4 Conclusions

A highly efficient CCE procedure was developed for the extrac-tion of curcumin from turmeric Turmeric material was loaded into

a column with 2-fold of 80% ethanol at a diameter-to-height ratio

of 1:10 After 1 h for dissolving target compounds, the column was eluted with 80% ethanol at a flow rate of 2 BVs/h For non-cyclic CCE procedure, 8-fold of eluent was collected, while for cyclic CCE procedure, only the first 2-fold of eluent was collected as extraction solution A more than 99% extraction rate for curcumin was obtained in both procedures, compared to a 59% extraction rate through the ultrasonic-assisted extraction with 10-fold of 80% ethanol The CCE procedure is good for non-continuous extrac-tion of samples in quantitative analysis, while the cyclic CCE proce-dure has multiple advantages for large scale and continuous extraction of curcumin Results indicated that the CCE procedures are highly efficient extraction methods for curcumin from turmeric, with minimum use of solvent, minimum volume and high concentration of extraction solution

Acknowledgments This work was supported by The Science and Technology Supporting Programs of Guangzhou Municipal Government (2008Z1-E591), Panyu District Science and Technology Programs

of Guangzhou City (2009-T-17-1) and Guangdong Natural Science Fund (10151063101000002)

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g)

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ethanol

20.82 ± 0.25 99.4 ± 1.2

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