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Occurrence of curcuminoids in Curcuma longa : A quality standardization by HPTLC

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A simple high performance thin layer chromatographic (HPTLC) method has been developed for the simultaneous determination of the pharmacologically important active curcuminoids viz. curcumin, demethoxycurcumin and bis-demethoxycurcumin in Curcuma longa L. The assay combines the separation and quantification of the analytes on silica gel 60 GF 254 HPTLC plates with visualization under UV and scanning at 425 nm. Using this technique, the alkaloidal content of different parts of the title plant has been determined.
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A journal of the Bangladesh Pharmacological Society (BDPS)
Bangladesh J Pharmacol 2008; 3: 55-58

Journal homepage:;
Indexed in Bangladesh Journals Online, Directory of Open Access Journals, Google Scholar and HINARI
ISSN: 1991-007X (Printed); 1991-0088 (Online); DOI: 10.3329/bjp.v3i2.833
Occurrence of curcuminoids in Curcuma longa: A quality
standardization by HPTLC
1M. Paramasivam, 1Md. Wasim Aktar, 1R. Poi, 1H. Banerjee and 2A. Bandyopadhyay
1Regional Analytical Laboratory of Medicinal and Aromatic Plants, Department of Agricultural Chemicals;
2Department of Spices and Plantation Crops, Bidhan Chandra Krishi Viswavidyalaya, Mohanpur 741252,
Nadia, West Bengal, India.

Article Info
Received: 24 April 2008
A simple high performance thin layer chromatographic (HPTLC)
Accepted: 9 May 2008
method has been developed for the simultaneous determination of the
Available online: 11 May 2008
pharmacologically important active curcuminoids viz. curcumin,
demethoxycurcumin and bis-demethoxycurcumin in Curcuma longa L.
Curcuma longa
The assay combines the separation and quantification of the analytes on
silica gel 60 GF254 HPTLC plates with visualization under UV and
scanning at 425 nm. Using this technique, the alkaloidal content of
different parts of the title plant has been determined.
Number of Tables: 2
Number of Figure: 1

Number of Refs:
Correspondence: MP


the determination of these alkaloids, are very
cumbersome and time-consuming and also not
Curcuma longa (Zingiberaceae), commonly called economically viable (Khurana and Ho, 1988;
Haldi, is a well-known plant drug in Ayurvedic Taylor and McDowell, 1992; Schieffer, 2002).
and Unani medicine (Chopra et al., 1956: Kapoor,
2001). It has been used for the treatment of various Therefore it was thought worthwhile to develop a
diseases and disorders particularly for urticaria, simple and high-precision HPTLC method for
skin allergy, viral hepatitis, inflammatory condi-
simultaneous analysis of curcumin, demethoxy
tions of joints, sore throat, and for wounds curcumin and bis-demethoxycurcumin occurring
(Chattopadhyay et al., 2004).
in roots of C. longa.
Curcumin, demethoxycurcumin and bis-demetho-

xycurcumin, three major pharmacologically Materials and Methods
important curcuminoids, have been isolated from Plant material: Rhizomes of C. longa were collected
C. longa (Gupta et al., 1999) and has been shown to from the Experimental Farm for the Development
possess anti-oxidant, anti-inflammatory, anti-
of Medicinal and Aromatic Plants (BCKV,
carcinogenic, anti-mutagenic, anti-fungal, anti-
Mohanpur, India). The plant specimens were
viral and anti-cancer (Chattopadhyay et al., 2004; authenticated and a voucher specimen is deposited
Ahsan et al., 1999). Methods, so far available for in the herbarium.

Bangladesh Pharmacological Society

Bangladesh J Pharmacol 2008; 3: 55-58
glass solvent development chamber well in
Table I: Rf values by HPTLC and linear regression equa-
advance to allow complete saturation which was
tions for the determination of curcumin, demethoxycurcumin
further enhanced by keeping one filter paper
and bis-demethoxycurcumin
along one wall of the twin trough chamber. The
Rf value
plate was then kept in a chamber and solvent
Y= 47.296+ 0.85X
front was allowed to develop at the height of 7 cm
on the plate. The TLC runs were made under
Y= 186.328+
laboratory conditions of 25 ± 50C and 50 % relative
humidity. After drying, the spots were visualized
under Camag UV cabinet (254 and 366 nm).
Y= 271.84+ 0.39X
Quantitative analysis of the compounds was done
by scanning the plates using Camag TLC scanner
Chemicals and standard alkaloids: Reagents used model 3 equipped with Wincats software (Camag)
were from Merck (Darmstadt. Germany). Analy-
applying the following conditions: slit width 6 x
tical standards of curcuminoids were obtained 0.45 mm, wavelength (?
from Ms. ChromaDex, Santa Ana, CA, USA.
max) 425 nm, absorption-
reflection scan mode. The identification of
Solvents used in entire study were from Merck, curcumin, demethoxycurcumin and bis-demetho-
India. The identities of curcumin, demethoxy-
xycurcumin in rhizomes were confirmed by
curcumin and bis-demethoxycurcumin were superimposing the UV spectra of samples and
confirmed by comparison of their spectral data standards within the same R
with those previously reported. (Govindarajan et
f window. In order to
prepare calibration curves, stock solutions of
al., 1980).
curcumin, demethoxycurcumin and bis-demetho-
Extraction of plant material for analysis: Air dried (35-
xycurcumin (1 mg/5 mL each) were prepared and
500C) rhizomes of seven germplasm of C. longa various volumes of these solutions were analyzed
and market turmeric powder samples (1 g each) by HPTLC exactly as described above. Then
were ultrasonically extracted separately in 20 mL calibration curves of peak area vs. concentration
HPLC grade methanol for 15 min (3 times) and were prepared.
filtered through Whatman No. 42 filter paper after
each extraction. Extracts were concentrated under
vacuum and finally made up to 20 ml with HPLC Results and Discussion
grade methanol and ready for HPTLC analysis.
Different compositions of the mobile phase for
Chromatographic conditions: Chromatography was HPTLC analysis were tested in order to obtain
performed on glass-backed silica gel 60 GF254 high resolution and reproducible peaks. The
HPTLC layers (20 x 20 cm, 300 µm layer thickness) desired aim was achieved using chloroform:
prepared using a Camag (Multenz, Switzerland) methanol (48:2, v/v) as the mobile phase. The
TLC plate auto-coater. Methanolic solutions of wave length of 425 nm was found to be optimal for
samples and standard compounds curcumin, the highest sensitivity (Figure 1). The calibration
demethoxycurcumin and bis-demethoxycurcumin curves for the alkaloids curcumin, demethoxy-
of known concentrations were applied to the layers curcumin and bis-demethoxycurcumin were linear
as 7 mm-wide bands positioned 15 mm from the in the range 100-1,000 ?g (Table I). The accuracy of
bottom and 20 mm from the side of the plate, using the determination of the recovery rate was
a Camag Linomat 5 automated TLC applicator determined by triplicate analyses of the rhizomes
with the nitrogen flow providing a delivery speed spiked with three different concentrations of stock
of 150 nL/s from the syringe. These parameters solution of curcumin, demethoxycurcumin and
were kept constant throughout the analysis of bis-demethoxycurcumin. The recovery rates were
97.3%, 92.9% and 95.4% for curcumin,
Detection and quantification of the alkaloids: After demethoxycurcumin and bis- demethoxycurcu-
sample application plates were developed in a min, respectively.
Camag twin trough glass tank pre-saturated with For the quantitative determination of curcumin,
the mobile phase chloroform:methanol (48:2, v/v) demethoxycurcumin and bis-demethoxycurcumin,
for one hour. It was then poured in twin trough the analyses of turmeric rhizome specimens of C.

Bangladesh J Pharmacol 2008; 3: 55-58

Figure 1: HPTLC chromatogram of the standard curcuminoids

reported here is very simple, sensitive, economic
Table II: Distribution of the alkaloids curcumin,
demthoxycurcu-min and bis-demethoxycurcumin in two
and suitable for rapid screening of large number of
cultivars of Curcuma longa
plant samples. Moreover, this analysis can be

performed without any special sample pre-
treatment and fifteen samples can be analyzed on a
single TLC laver (20 x 20 cm).
(%dry weight)a

(% dry weight) a
Ahsan H, Parveen N, Khan NU, Hadi SM. Pro-oxidant,
(% dry weight) a
antioxidant and cleavage activities on DNA of
a Mean values (n = 3)
curcumin and its derivatives demethoxycurcumin and
bis-demethoxycurcumin. Chem Biol Interact. 1999;
longa were repeated three times. The average
121: 161-75.
content of curcumin, demethoxycurcumin and bis-
demethoxycurcumin in the rhizomes are given in
Table II. It is clear that three alkaloids were present Chattopadhyay I, Biswas K, Bandyopadhyay U,
in two cultivars viz. Kalimpong and PTS-43 having
Banerjee RK. Turmeric and curcumin: Biological
actions and medicinal applications. Curr Sci. 2004;
their maximum concentrations in the Kalimpong
87: 44-50.
Chopra RN, Nayar SL, Chopra IC. Glossary of Indian
In summary, the HPTLC method for the simulta-
medicinal plants. New Delhi, CSTR, 1956, p 7.
neous analysis of curcumin, demethoxycurcumin Govindarajan VS. Turmeric- Chemistry, technology and
and bis-demethoxycurcumin from C. longa
quality. Crit Rev Food Sci Nutr. 1980; 12: 199-301.

Bangladesh J Pharmacol 2008; 3: 55-58
Gupta AP, Gupta MM, Kumar S. Simultaneous
determination of curcuminoids in curcuma samples
using High Performance Thin Layer Chromato-
Schieffer GW. Pressurized liquid extraction of curcumi-
graphy. J Liq Chromatogr Related Technol. 1999;
noids and curcuminoid degradation products from
22: 1561-69.
turmeric (Curcuma longa) with subsequent HPLC
assays. J Liq Chromatogr Related Technol. 2002; 25:
Kapoor LD. Handbook of Ayurvedic medicinal plants.
Boca Raton, FL, CRC Press, 2001, p 216.
Taylor SJ, McDowell IJ. Determination of the
Khurana A, Ho CT. High Performance Liquid
curcuminoid pigments in turmeric (Curcuma
Chromatographic analysis of curcuminoids and their
domestica Val.) by reversed phase High Performance
photo-oxidative decomposition compounds in
Liquid Chromatography. Chromatographia 1992; 34:
Curcuma longa L. J Liq Chromatogr. 1988; 11:

Occurrence of curcuminoids in Curcuma longa : A quality standardization by HPTLC



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