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Journal of Chemistry
Volume 2013, Article ID 493870, 4 pages
http://dx.doi.org/10.1155/2013/493870

Research Article
Isolation and Characterization of Antitumor Alkaloid from
Poppy Capsules (Papaver somniferum)

Ibrahim Bulduk1 and Fulya Taktak2

1
Department of Chemistry, University of Uşak, 64200 Uşak, Turkey
2
Department of Chemical Engineering, University of Uşak, 64200 Uşak, Turkey

Correspondence should be addressed to Fulya Taktak; [email protected]

Received 17 May 2012; Accepted 30 July 2012

Academic Editor: Malgorzata Baranska

Copyright © 2013 I. Bulduk and F. Taktak. is is an open access article distributed under the Creative Commons Attribution
License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly
cited.

Noscapine, a phthalideisoquinoline alkaloid derived from opium, has been used in the treatment of various cancer types. Its low-
toxicity pro�le has increased attention to this alkaloid. �ith regard to increasing demand for this compound, we developed a new
method for isolation of noscapine from dried capsules of Papaver somniferum. Noscapine was successfully isolated from poppy
capsules for the �rst time and the purity of the isolated compound was determined to be over 99.59% by HPLC analysis. e
structure of noscapine was con�rmed by 1 H NMR, 13 C NMR, FT-IR, elemental analysis, and HR-ESI-MS methods.

1. Introduction has an excellent pharmacological pro�le, therefore the pure

yield of this alkaloid from natural plants is very important.
Important class of pharmaceutical compounds, they are Isolation of noscapine from opium was studied in litera-
alkaloids form, which can be found in natural products ture. Sim showed the separation of alkaloids from opium in a
such as the opium poppy, Papaver somniferum. Alkaloids step-wise process and produced noscapine as the last product
are normally removed from natural products using solvent [9]. Also Ramanathan and Chandra report several processes
extraction. However, the extraction of very small quantities for purifying narcotine which is known as an impure form
of alkaloids in natural plants required the development of of “Noscapine” [10]. In these methods are used opium as the
new and improved experimental methods. Noscapine, (3S)- starting material.
6,7-Dimethoxy-3-[(5R)-4-methoxy-6-methyl-5,6,7,8-tetrahy- On the other hand, the poppy straw is an alternative
dro-1,3-dioxolo[4,5-g]isoquinolin-5-yl]isobenzofuran-1(3H)- source which is chemically used to extract the alkaloids [11].
one, is a natural phthalideisoquinoline alkaloid obtained For this method, the capsules of P. somniferum are cut and
from the opium poppy which has been used for many dried in air in the �eld then harvested and crushed to straw.
decades as a very safe cough suppressant with no side effects In many countries this method is present as an alternative
[1, 2]. Unlike other alkaloids, noscapine is not addictive. route to obtain alkaloids like noscapine due to the fact that
Recently, antitumor properties of noscapine have been the usage of crude opium which contains the controlled
reported by Joshi�s group for the �rst time [3] and research substance morphine may be limited by governmental rules.
on the therapeutic effect of noscapine in inhibiting tumor Because of this restriction, noscapine producers have been
progression in experimental animals is ongoing [4–6]. searching for a new source and poppy straw, which is
Moreover, histological studies have shown that noscapine is harvested when it is fully mature and dry, which is suitable
not toxic to normal tissues [1, 7, 8]. Generally, alkaloids are for this. Also, it is important to say that poppy straw may
obtained from opium poppy, are structurally complex, and contain much higher quantities of noscapine relative to
most of them cannot be economically synthesized. Noscapine morphine content [12]. However, P. somniferum has been
2 Journal of Chemistry

O give 200 cm3 of brown aqua phase. e pH of the solution was

NCH3 set 3.5 by adding acetic acid. 5.0 g of activated charcoal were
O added into this solution and contents was heated to 60○ C and
H3 CO stirred mechanically for �een minutes. is solution was
O �ltered. e �ltrate was extracted with chloroform (4 times
250 cm3 ) and the extracts are combined. e chloroform
OCH3
O extract was concentrated to give 15 g of dark brown residue.
OCH3
is residue was dissolved in 400 mL of 10% acetic acid
solution and heated to 50○ C. e pH of this solution was
made 9.3 by adding 25% ammonia slowly. Crude noscapine
F 1: Chemical structure of noscapine. was precipitated. e precipitate was �ltered and dried in an
oven at 105○ C.

genetically modi�ed by gene insertion to either enhance 2.�. Puri�cation of Crude Noscapine. Crude noscapine was
alkaloid production or to “switch off ” the genes responsible suspended in 50 cm3 of toluene. e suspension was heated
for production of nonrequired alkaloids. to 70–80○ C. 5.0 g of activated charcoal was added and
A simple process for isolating the alkaloids is highly stirred mechanically for �een minutes at room temperature.
desirable to evaluate product quality. is paper describes a e solution was �ltered. e �ltrate was concentrated
method for the extraction and puri�cation of noscapine that under reduced pressure to give white precipitate and stirred
is present in the poppy straw. In this method, noscapine was mechanically for thirty minutes in an ice bath. e white
obtained in purity suitable for use in pharmacopy (99.59%). precipitate was �ltered and dried in an oven at 105○ C. e
e chemical structure of this alkaloid is shown in Figure 1. resulting white solid corresponded to a yield of 90% and
purity (by HPLC) of 99.59%. M.p.: 174–177○ C (decomp);
[𝛼𝛼]𝐷𝐷 = −176○ (𝑐𝑐 = 0.5 g/100 cm3 , CHCl3 ). 1 H NMR (500
22
2. Experimental
MHz, CDCl3 ): 𝛿𝛿 = 1.88–1.93 (m, 1H), 2.32–2.38 (m, 2H),
2.1. General. Melting point was determined in open glass 2.55 (s, 3H), 2.58–2.61 (m, 1H), 3.86 (s, 3H), 4.05 (s, 3H), 4.09
capillary tube by means of a BUCHI Melting Point B- (s, 3H), 4.40 (d, 𝐽𝐽 = 3.8 Hz, 1H), 5.58 (d, 𝐽𝐽 = 3.8 Hz, 1H), 5.94
540 apparatus. Infrared (FT-IR) spectra were taken using (s, 𝐽𝐽 = 1.2 Hz, 1H), 6.08 (d, 𝐽𝐽 = 8.2 Hz, 1H), 6.31 (s, 1H), 6.96
a PerkinElmer Spectrum One FT-IR spectrometer, 𝜈𝜈max in (d, 𝐽𝐽 = 8.2 Hz, 1H); 13 C NMR (125 MHz, CDCl3 ): 𝛿𝛿 = 28.0
cm−1 . e 1 H and 13 C nuclear magnetic resonance (NMR) (t), 46.3 (q), 50.0 (t), 56.9 (d), 59.4 (d), 60.9 (d), 62.2 (d), 81.8
were taken with a Bruker 500 NMR spectrometer. Chemical (d), 100.7 (t), 102.3 (d), 117.2 (s), 117.6 (d), 118.4 (d), 120.2
shis are reported in ppm (𝛿𝛿). High resolution electrospray (s), 132.1 (s), 134.1 (s), 140.5 (s), 141.3 (s), 147.9 (s), 148.4 (s),
ionization mass spectra (HR-ESI-MS) were obtained with 152.2 (s), 168.0 (s).
MeOH on a Bruker micrOTOF-Q. Elemental analyses (EA)
were performed using a ermo Finnigan FLASH 1112
2.5. Determination of Alkaloid Content of Poppy Capsule,
SERIES EA instrument. e speci�c rotation was measured
on an Optical Activity Ltd. A55 polarimeter instrument Crude Noscapine and Pure Noscapine by HPLC
using 2 cm3 cell with a 0.5 dm path length and the sample 2.5.1. Sample Preparation. 200 mg of homogenized, pow-
concentration is given in g/100 cm3 unit. Chemical reagents dered, dried poppy capsule was shaken with 50.0 cm3 0.1 M
for isolation were of analytical grade and purchased from hydrochloric acid for 2 hours. An aliquot of the extract was
Merck (Darmstadt, 90 Germany). All solvents were �ltered �ltrated and the �ltrate was �lled to a vial.
through 0.45 𝜇𝜇m membrane �lters before being injected into 50 mg of crude noscapine or pure noscapine was dis-
the HPLC. solved in 5 cm3 of 10% acetic acid solution is added approx-
imately and 100 cm3 of demineralised water, sonicated to
2.2. Plant Material. Poppy capsules were procured from dissolve, then diluted to 250 cm3 with demineralised water.
Turkish Grain Board in August 2009. For this method, the It was mixed well and �lled to a vial.
capsules of P. somniferum are cut and dried in air in the
�eld then harvested and crushed to straw. ese capsules are 2.6. Chromatographic System. An Agilent 1200 HPLC with
grown from genetically engineered seeds and contain 1.5% UV-vis detection capabilities and a 0.01 cm3 injection volume
(w/w) noscapine. was used. e wavelength used during the analysis was
288 nm. A Eurospher-100 column (KNAUER) measuring
2.3. Extraction of Crude Noscapine. A three-neck round 120 × 4 mm, 5 𝜇𝜇m particle size and with C18 stationary
bottom �ask was charged with 500 g of air-dried and ground phase was used. e column was heated and maintained at
poppy straw, 2000 cm3 of ethanol, and 250 cm3 of dem- 40○ C. e mobile phases consisted of (A) water with 0.67%
ineralised water. e contents of the �ask were stirred tri�uoroacetic acid (TFA) and (B) 0.67% TFA in a mixture
mechanically. e suspension was heated and re�uxed for of methanol, acetonitrile, and water (500/16/129 cm3 ) using
two hours at 80–85○ C. e suspension was �ltered. e a gradient system. e �ow rate was 1.0 mL/min and the
alcohol extract was concentrated under reduced pressure to applied elution program is described in Table 1.
Journal of Chemistry 3

T 1: Parameters of gradient elution program. ×106

414.1532
Time (min) Flow rate (cm3 /min) Eluent A (v/v) Eluent B (v/v)
3
0.0 1.0 84.0 16.0

Intensity
14.4 1.0 45.2 54.8 2
14.5 1.0 0.0 100.0
16.4 1.0 0.0 16.0 1
16.5 1.0 84.0 16.0 409.16 435.1339
0
395 400 405 410 415 420 425 430 435
५ॸ
1.946

14.565-noscapine
F 3: HR-ESI-MS data of pure noscapine product.
3.275-morphine

12.523-thebaine
10.658

13.46
6.517
7.066

1
14.593-noscapine 0.8

Absorbance
12.559-thebaine

0.6
0.4
0.2
14.586-noscapine

O 0
NCH3
O
H3 CO
4000 3500 3000 2500 2000 1500 1000 500
Wavenumbers (cm−1 )
O
OCH3
O
OCH3
Noscapine
Noscapine standard
0 2 4 6 8 10 12 14
(min) F 4: FT-IR spectrums of noscapine product and noscapine
standard (purchased from Turkish Grain Board).
F 2: HPLC chromatograms of step by step noscapine isolation
and puri�cation (a) alkaloids in poppy straw sample. (b) Chro-
matogram of crude noscapine (the crude extract contains 49.411%
noscapine) aer extraction process. (c) Chromatogram of pure noscapine standard (purchased from Turkish Grain Board).
(99.59%) noscapine. IR spectra show numerous sharp bands between 700 and
1500 cm−1 which are assigned to deformation and stretching
vibrations of the alkaloid ring system. Also, the strong band
3. Results and Discussion at 1756 cm−1 is the most characteristic for C=O stretching
vibration.
Noscapine was isolated as a white crystalline solid with e NMR spectrum of noscapine shows two one-proton
negative rotation [𝛼𝛼]𝐷𝐷 = −176○ (𝑐𝑐 = 0.5 g/100 cm3 , CHCl3 ).
22 doublets at 6.96 and 6.08 ppm assigned to the aromatic
Several elution systems were tested in HPLC separation. protons in the phthalidyl ring. e one-proton singlet signal
e results indicated that when methanol-acetonitrile-water at 6.31 ppm is attributed to aromatic proton in the isoquino-
was used as mobile phase in gradient mode (Table 1), line ring. e three-proton singlets at 3.86 ppm, 4.05, and
good seperation results could be obtained. HPLC analysis of 4.09 ppm are ascribed to methoxy protons in isoquinoline
poppy straw, crude extract, and pure noscapine show that and phthalidyl rings, respectively. e signal at 5.94 which
noscapine was clearly seperated and exhibited well-de�ned is given as a two-proton singlet is accounted the methylene-
chromatogram with retention times of 14.586 min in the dioxy protons. e singlet signal at 2.55 ppm is due to the
selected experimental conditions (Figure 2). e content of –N–CH3 protons. e –CH2 –CH2 – protons of isoquinoline
noscapine in the crude extracts was 49.4%. Aer puri�cation ring are appeared as a multiplet at 2.32–2.38 ppm.
treatment, the purity of the collected fraction peak 1 was e 13 C NMR spectrum of compound exhibited signals
found to be 99.59%. for aromatic carbons between 117.2–134.1 ppm, methylene-
Its molecular formula was determined as C22 H23 NO7 dioxy carbon at 100.7 ppm, and carbonyl carbon at 168.0
on the basis of HR-ESI-MS (𝑚𝑚/𝑧𝑧 414.1553 [M+ H]+ , calcd ppm. e signals at 56.9, 59.4, and 62.2 ppm are ascribed
for C22 H23 NO7 414.1532). Figure 3 shows the expected to methoxy carbon atoms. e carbon atom which bonded
molecular ion peak at 𝑚𝑚/𝑧𝑧 = 414.1553, indicating formation nitrogen atom is observed at 46.3 ppm.
of [M+ H]+ .
e chemical structure of peak fraction in Figure 2(c) was 4. Conclusions
identi�ed according to its FT-IR, 1 H NMR data 13 C NMR.
ese were well-matched with the literature [12–15]. Figure Prior to now, noscapine is isolated from opium. Recently,
4 shows infrared spectra of pure noscapine product and following seed development and gene studies, poppy seed
4 Journal of Chemistry

including capsules that are rich in noscapine have been [14] H. Schulz, M. Baranska, R. Quilitzsch, and W. Schütze, “Deter-
developed. To our knowledge, this is the �rst report on mination of alkaloids in capsules, milk and ethanolic extracts of
the isolation of noscapine from poppy capsules. e isola- poppy (Papaver somniferum L.) by ATR-FT-IR and FT-Raman
tion method is very short compared with other production spectroscopy,” Analyst, vol. 129, no. 10, pp. 917–920, 2004.
methods from opium. Hence, poppy straw could be used [15] R. L. Prasad, S. N. akur, and G. C. Bhar, “CO2 laser photoa-
as an accessible source of noscapine product with high coustic spectra and vibrational modes of heroin, morphine and
yield. Its chemical structure was clearly con�rmed by using narcotine,” Pramana, vol. 59, no. 3, pp. 487–496, 2002.
various instrumental techniques. e obtained product has
pharmacological purity and does not include impurities.

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