- Email: [email protected]
Isolation and characterization of steroids from Calligonum polygonoides
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 3 4 6 e3 4 9
Available online at www.sciencedirect.com
journal homepage: www.elsevier.com/locate/jopr
Original Article
Isolation and characterization of steroids from Calligonum polygonoides Muhammad Qasim Samejo a,b, Shahabuddin Memon a,*, Muhammad Iqbal Bhanger a, Khalid Mohammed Khan c a
National Center of Excellence in Analytical Chemistry, University of Sindh, Jamshoro 76080, Pakistan Dr. M. A. Kazi Institute of Chemistry, University of Sindh, Jamshoro 76080, Pakistan c H. E. J. Research Institute of Chemistry, International Center Chemical and Biological Science, University of Karachi, Karachi 75270, Pakistan b
article info
abstract
Article history:
Aim: Calligonum polygonoides Linn. (Polygonaceae) is commonly known as ‘Phog’ contains
Received 30 January 2013
number of phytochemical constituents’ viz. flavonoids, alkaloids, proteins, tannins, ste-
Accepted 14 March 2013
roids, phenols, carbohydrates, terpenoids etc. The aim of the present study was to isolate
Available online 1 April 2013
and characterize steroids from the roots of C. polygonoides. Method: Methanol extract of the roots of plant was subjected to column chromatography
Keywords:
and eluted with solvent mixtures of increasing polarity, composed of chloroform, chloro-
Calligonum polygonoides
formeethyl acetate mixtures and ethyl acetate to isolate phytochemical constituents. The
Polygonaceae
identities of these compounds were checked by solubility, preliminary phytochemical test,
Steroids
melting point determination and TLC study. Finally the structure was elucidated by different chromatographic and spectroscopic methods. Results: On the basis of chemical and spectral evidence and upon comparison with the literature data, the isolated compounds were identified as campesterol (1), stigmasterol (2), (3b,5a,24S)-stigmastan-3-ol (3) and stigmast-4-en-3-one (4). Conclusion: The steroids were isolated from the roots of C. polygonoides may serve as a potential source of useful drugs in the near future. The steroids were isolated first time from this plant extract. Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved.
1.
Introduction
Natural products, such as plants extract, either as pure compounds or as standardized extracts, provide unlimited opportunities for new drug discoveries because of the unmatched availability of chemical diversity. The medicinal value of plants is due to the presence of chemical constituents such as flavonoids, alkaloids, terpenoids, tannins and
steroids.1,2 Steroids are terpenoids lipids identified by carbon skeleton with 4 fused rings. Steroids are differing due to their oxidation state of functional groups attached to the rings and oxidation state of rings. The major responsibilities of steroids (androgens, progestagens, estrogens, mineralocorticoids and glucocorticoids) are to salt balance, controlling metabolism and the improvement and function of the sexual organs as well as other biological differences between the sexes.
* Corresponding author. Tel.: þ92 (22) 9213430; fax: þ92 (22) 9213431. E-mail address: [email protected] (S. Memon). 0974-6943/$ e see front matter Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved. http://dx.doi.org/10.1016/j.jopr.2013.03.017
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 3 4 6 e3 4 9
Steroids in the form of bile salts (e.g., salts of deoxycholic and cholic acid and their taurine conjugates and glycine) facilitate in digestive processes. Synthetic steroids like glucocorticosteroids, estrogens, methylprednisolone, corticosteroids, androgens, squalamine and hydrocortisone are also used for the treatment of various diseases such as arthritis, malignancies, allergic reactions, and diseases resulting from abnormal production or hormone deficiencies.3 Campesterol (rapeseed, soy and wheat-germ oils) is the most familiar plant sterols in nature along with stigmasterol and b-sitosterol, it show cholesterol lowering and anticarcinogenic effects. Since angiogenesis is essential for cancer, it was surmise that an antiangiogenic effect may be involved in the anticancer action of this compound.4 Stigmasterol may be useful in prevention of certain cancers, including ovarian, prostate, breast, and colon cancers. It possesses potent antioxidant, hypoglycemic and thyroid inhibiting properties.5,6 Stigmast-4-en-3one show orally hypoglycaemic agent and necessary intermediate in the metabolism of b-sitosterol.7 (3b,5a,24S)-stigmastan-3-ol also reduce the absorption of cholesterol from the diet.8 The genus Calligonum belongs to the family Polygonaceae, comprises of about 80 species and is found in many countries such as Northern Africa, Southern Europe and Western Asia. Calligonum polygonoides Linn. is known for its medicinal properties. The flowers of C. polygonoides are useful against cough, asthma and cold. The juice of shoot is applied to the eyes as an antidote to scorpion sting, a roots decoction mixed with catechu is used as gargle for sore gum, and the latex is used for treating eczema, to cure bites of rabid dogs and to induce abortion. Methanol extract of the C. polygonoides showed strong toxicity in brine-shrimp lethality test.9 Phytochemical screening of C. polygonoides shows positive results for flavonoids, alkaloids, proteins, tannins, steroids, phenols, carbohydrates and terpenoids.10 The essential oil from buds and roots of C. polygonoides contain a complex mixture of terpenoids, hydrocarbons, phenolic compounds, acid derivatives and ketones. The literature survey revealed that the Calligonolides, tetracosan-4-olide, steroidal ester, b-sitosterol, b-sitosterol glucoside and ursolic acid isolated from C. polygonoides.9 The aim of present study was to isolate and identify the steroids from the roots of C. polygonoides. To the best of our knowledge, these steroids (1e4) were found for the first time from this species.
2.
Materials and methods
2.1.
Collection and identification of plant materials
Roots of C. polygonoides were collected from Village MehendriJo-Par (longitude: N 25 340 200 and latitude: E 70 110 2000 ), District Umerkot in Sindh Province of Pakistan in January 2012. A voucher specimen (15173) of the plant was deposited in the herbarium of Institute of Plant Sciences, University of Sindh Jamshoro, Pakistan. The plant sample was identified by a Taxonomist of the same institution. The plant material was air dried under normal conditions and ventilated.
2.2.
347
Extraction and isolation
About 300 g powdered roots of C. polygonoides were macerated in methanol for three days. Occasional shaking and stirring was done. Then extract was filtered using Whatman filter paper. The filtrate was concentrated to dryness under the vacuum. Chemical tests (Salkowski and LiebermanneBurchard reaction) were performed to detect the steroids in the extract.6 The dried methanol extract was subjected to column chromatography over silica gel (particle size 0.2e0.5 mm, 35e70 mesh ASTM) and gradient elutions were carried out with eluents chloroform, chloroformeethyl acetate mixtures and ethyl acetate. Each fraction was collected in 20 ml portion and was monitored by TLC (Pre-coated aluminum silica gel plates). The chloroform fraction was further purified by preparative TLC using hexane:chloroform (40:60) solvent system. TLC result shows the four spots with different retention time. Each spot (showing compound) was scratched separately and dissolved in hexane then filtered using Whatman filter paper. The isolated compounds were again confirmed of their identity by chemical tests. For further characterization UV, FT-IR and GCeMS was done.
2.3.
Gas chromatographyemass spectrometry (GCeMS)
GCeMS analysis of plant sample was performed on Agilent 6890 N GC instrument coupled with MSe5975 inert XL mass selective detector and auto sampler 7683-B injector was used. The HPe5MS column with dimensions of 30 m 0.25 mm i.d., film thickness 0.25 mm was used for the analysis. Initial temperature 150 C, maintained for 2 min, final temperature 230 C, kept for 5 min, ramp rate 4 C/min. 1.0 ml sample was injected, using split mode (split ratio, 10:1). Helium gas was used as a carrier gas at a flow rate of 0.8 ml/min. An electron ionization mode with ionization energy of 70 eV was used for MS detection. The injector and MS transfer line temperatures were set at 240 and 270 C, respectively.
2.4.
FT-IR analysis
FT-IR spectra were obtained using a Thermo Nicolet Avatar 330 FT-IR spectrometer controlled by OMNIC software (Thermo Nicolet Analytical instruments, Madison, WI, USA) station with a deuterated triglycine sulfate (DTGS) detector and KBr optics. The sampling station was equipped with overhead ATR accessory (Spectra-Tech, Shelton, CT) comprising of transfer optics with in the chamber through which infrared radiation is directed to a detachable ATR zinc selenide crystal mounted in a shallow trough for sample containment. A single beam spectrum (4000-650 cm1) of the sample was obtained against air as a background at a resolution of 4 cm1 and a total of 32 scan.11
3.
Results and discussion
The methanol extract of C. polygonoides roots was subjected to different phytochemical tests and it gives highly positive results for steroids. The extract was subjected to column chromatography over silica gel. The column was eluted in different solvent
348
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 3 4 6 e3 4 9
isolated compounds are identified as campesterol (1), stigmasterol (2), (3b,5a,24S)-stigmastan-3-ol (3) and stigmast-4en-3-one (4) (Fig. 1) from methanol extract of the roots of C. polygonoides.
HO
HO
Conflicts of interest Stigmasterol (2)
Campesterol (1)
All authors have none to declare. (C29H48O; Mol.Wt. 412.69)
(C28H48O; Mol.Wt. 400.68)
Acknowledgments Financial support and necessary facilities offered by National Centre of Excellence in Analytical Chemistry (NCEAC), University of Sindh, Jamshoro, Pakistan is gratefully acknowledged.
O
HO
Stigmast-4-en-3-one (4) (C29H52O; Mol.Wt. 416.72)
Fig. 1 e Sterols identified in roots of Calligonum polygonoides (1e4). system (CHCl3, CHCl3eEtOAc mixtures and EtOAc) with gradient elutions. Each fraction was monitored by TLC. The chloroform fraction was further purified by preparative TLC using hexane:chloroform (40:60) solvent system. The TLC result leads to the isolation of campesterol (1), stigmasterol (2), (3b,5a,24S)stigmastan-3-ol (3) and stigmast-4-en-3-one (4) (shown in Fig. 1). The FT-IR spectra of isolated compounds exhibit the diagnostic peaks relating to CeH stretching at 2950 cm1 and 2860 cm1. The OeH stretching and C]C absorption peak appears at 3360 cm1 and 1630 cm1, respectively. Other absorption peaks includes 1445 cm1 (CH2); 1371 cm1 (OH def), 1050 cm1 (cycloalkane) verify the required data regarding the structures of steroids. UV spectrum (recorded in methanol) of stigmasterol exhibited peak at 256 nm because it contains two isolated double bond. While, stigmast-4-en-3-one and campesterol exhibited peaks at 231 and 251 nm respectively. GCeMS is the most useful method for the characterization of steroids.12,13 Each compound was analyzed by GCeMS and identified by comparison of their mass spectra with the reference compounds in the data systems of Wiley and National Institute of Standards and Technology (NIST) spectra libraries matching. Compounds were identified with a resemblance percentage above 90%. Further conformation of these compounds was done by comparison of their and mass spectra with data in literature.14e19 Results show good agreement for the structure of campesterol (1), stigmasterol (2), (3b,5a,24S)-stigmastan-3-ol (3) and stigmast-4-en-3-one (4) as reported in the literature.
4.
references
(C29H48O; Mol.Wt. 412.69)
Conclusion
On the basis of chemical and spectral evidence and upon comparison of obtained data with the literature data, the
1. Kardong D, Upadhyaya S, Saikia LR. Screening of phytochemicals, antioxidant and antibacterial Activity of crude extract of Pteridium aquilinum Kuhn. J Pharm Res. 2012;5(11):5194e5196. 2. Sasidharan S, Chen Y, Saravanan D, Sundram KM, Latha LY. Extraction, isolation and characterization of bioactive compounds from plants’ extracts. Afr J Tradit Complement Altern Med. 2011;8(1):1e10. 3. Bhawani SA, Sulaiman O, Hashim R, Ibrahim MNM. Thinlayer chromatographic analysis of steroids. Trop J Pharm Res. 2011;9:301e313. 4. Choi JM, Lee EO, Lee HY, et al. Identification of campesterol from Chrysanthemum coronarium L. and its antiangiogenic activities. Phytother Res. 2007;21:954e959. 5. Gabay O, Sanchez C, Salvat C, et al. Stigmasterol: a phytosterol with potential anti-osteoarthritic properties. Am J Clin Nutr. 2010;18:106e116. 6. Panda S, Jafri M, Kar A, Meheta BK. Thyroid inhibitory, antiperoxidative and hypoglycemic effects of stigmasterol isolated from Butea monosperma. Fitoterapia. 2009;80:123e126. 7. Jamaluddin F, Mohamed S, Lajis MN. Hypoglycaemic effect of Stigmast-4-en-3-one from Parkia speciosa empty pods. Food Chem. 1995;54:9e13. 8. Batta AK, Xu G, Honda A, Miyazaki T, Salen G. Stigmasterol reduces plasma cholesterol levels and inhibits hepatic synthesis and intestinal absorption in the rat. MetabolismClinical Exp. 2006;55:292e299. 9. Samejo MQ, Memon S, Bhanger MI, Khan KM. Chemical composition of essential oil from Calligonum polygonoides Linn. Nat Prod Res; 2012. http://dx.doi.org/10.1080/14786419.2012. 686904. 10. Samejo MQ, Memon S, Bhanger MI, Khan KM. Preliminary phytochemical screening of Calligonum polygonoides Linn. J Pharm Res. 2011;4:4402e4403. 11. Burdi DK, Samejo MQ, Bhanger MI, Khan KM. Fatty acid composition of Abies pindrow (West Himalayan fir). Pak J Pharm Sci. 2007;20:15e19. 12. Radulovic NS, Dordevic ND. Steroids from poison hemlock (Conium maculatum L.): a GCeMS analysis. J Serb Chem Soc. 2011;76:1471e1483. 13. Lacikova L, Zapletal J, Masterova I, Grancai D. GCeMS analysis of leaves petrol ether extracts from four Staphylea L. species. Acta Facult Pharm Univ Comenianae. 2007;54:104e108.
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 3 4 6 e3 4 9
14. Jain PS, Bari SB. Isolation of lupeol, stigmasterol, caempsterol from petroleum ether extract of woody stem of Wrightia tinctoria. Asian J Plant Sci. 2010;9:163e167. 15. Kamboj A, Saluja AK. Isolation of stigmasterol and bsitosterol from petroleum ether extract of aerial parts of Ageratum conyzoides (Asteraceae). Int J Pharm Pharm Sci. 2011;3:94e96. 16. Yang B, Koponen J, Kallio RTH. Plant sterols in seeds of two species of Vaccinium (V. myrtillus and V. vitis-idaea) naturally distributed in Finland. Eur Food Res Technol. 2003;216:34e38.
349
17. Alexander-Lindo RL, Morrison EY, Nair MG. Hypoglycaemic effect of stigmast-4-en-3-one and its corresponding alcohol from the bark of Anacardium occidentale (Cashew). Phytother Res. 2004;18:403e407. 18. Johnsson L, Dutta PC. Characterization of side-chain oxidation products of sitosterol and campesterol by chromatographic and spectroscopic methods. JAOCS. 2003;80:767e776. 19. Patra A, Jha S, Murthy PN, Manik Sharone A. Isolation and characterization of stigmast-5-en-3b-ol (b-sitosterol) from the leaves of Hygrophila spinosa T. Anders. Int J Pharma Sci Res. 2010;1:95e100.
Available online at www.sciencedirect.com
journal homepage: www.elsevier.com/locate/jopr
Original Article
Isolation and characterization of steroids from Calligonum polygonoides Muhammad Qasim Samejo a,b, Shahabuddin Memon a,*, Muhammad Iqbal Bhanger a, Khalid Mohammed Khan c a
National Center of Excellence in Analytical Chemistry, University of Sindh, Jamshoro 76080, Pakistan Dr. M. A. Kazi Institute of Chemistry, University of Sindh, Jamshoro 76080, Pakistan c H. E. J. Research Institute of Chemistry, International Center Chemical and Biological Science, University of Karachi, Karachi 75270, Pakistan b
article info
abstract
Article history:
Aim: Calligonum polygonoides Linn. (Polygonaceae) is commonly known as ‘Phog’ contains
Received 30 January 2013
number of phytochemical constituents’ viz. flavonoids, alkaloids, proteins, tannins, ste-
Accepted 14 March 2013
roids, phenols, carbohydrates, terpenoids etc. The aim of the present study was to isolate
Available online 1 April 2013
and characterize steroids from the roots of C. polygonoides. Method: Methanol extract of the roots of plant was subjected to column chromatography
Keywords:
and eluted with solvent mixtures of increasing polarity, composed of chloroform, chloro-
Calligonum polygonoides
formeethyl acetate mixtures and ethyl acetate to isolate phytochemical constituents. The
Polygonaceae
identities of these compounds were checked by solubility, preliminary phytochemical test,
Steroids
melting point determination and TLC study. Finally the structure was elucidated by different chromatographic and spectroscopic methods. Results: On the basis of chemical and spectral evidence and upon comparison with the literature data, the isolated compounds were identified as campesterol (1), stigmasterol (2), (3b,5a,24S)-stigmastan-3-ol (3) and stigmast-4-en-3-one (4). Conclusion: The steroids were isolated from the roots of C. polygonoides may serve as a potential source of useful drugs in the near future. The steroids were isolated first time from this plant extract. Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved.
1.
Introduction
Natural products, such as plants extract, either as pure compounds or as standardized extracts, provide unlimited opportunities for new drug discoveries because of the unmatched availability of chemical diversity. The medicinal value of plants is due to the presence of chemical constituents such as flavonoids, alkaloids, terpenoids, tannins and
steroids.1,2 Steroids are terpenoids lipids identified by carbon skeleton with 4 fused rings. Steroids are differing due to their oxidation state of functional groups attached to the rings and oxidation state of rings. The major responsibilities of steroids (androgens, progestagens, estrogens, mineralocorticoids and glucocorticoids) are to salt balance, controlling metabolism and the improvement and function of the sexual organs as well as other biological differences between the sexes.
* Corresponding author. Tel.: þ92 (22) 9213430; fax: þ92 (22) 9213431. E-mail address: [email protected] (S. Memon). 0974-6943/$ e see front matter Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved. http://dx.doi.org/10.1016/j.jopr.2013.03.017
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 3 4 6 e3 4 9
Steroids in the form of bile salts (e.g., salts of deoxycholic and cholic acid and their taurine conjugates and glycine) facilitate in digestive processes. Synthetic steroids like glucocorticosteroids, estrogens, methylprednisolone, corticosteroids, androgens, squalamine and hydrocortisone are also used for the treatment of various diseases such as arthritis, malignancies, allergic reactions, and diseases resulting from abnormal production or hormone deficiencies.3 Campesterol (rapeseed, soy and wheat-germ oils) is the most familiar plant sterols in nature along with stigmasterol and b-sitosterol, it show cholesterol lowering and anticarcinogenic effects. Since angiogenesis is essential for cancer, it was surmise that an antiangiogenic effect may be involved in the anticancer action of this compound.4 Stigmasterol may be useful in prevention of certain cancers, including ovarian, prostate, breast, and colon cancers. It possesses potent antioxidant, hypoglycemic and thyroid inhibiting properties.5,6 Stigmast-4-en-3one show orally hypoglycaemic agent and necessary intermediate in the metabolism of b-sitosterol.7 (3b,5a,24S)-stigmastan-3-ol also reduce the absorption of cholesterol from the diet.8 The genus Calligonum belongs to the family Polygonaceae, comprises of about 80 species and is found in many countries such as Northern Africa, Southern Europe and Western Asia. Calligonum polygonoides Linn. is known for its medicinal properties. The flowers of C. polygonoides are useful against cough, asthma and cold. The juice of shoot is applied to the eyes as an antidote to scorpion sting, a roots decoction mixed with catechu is used as gargle for sore gum, and the latex is used for treating eczema, to cure bites of rabid dogs and to induce abortion. Methanol extract of the C. polygonoides showed strong toxicity in brine-shrimp lethality test.9 Phytochemical screening of C. polygonoides shows positive results for flavonoids, alkaloids, proteins, tannins, steroids, phenols, carbohydrates and terpenoids.10 The essential oil from buds and roots of C. polygonoides contain a complex mixture of terpenoids, hydrocarbons, phenolic compounds, acid derivatives and ketones. The literature survey revealed that the Calligonolides, tetracosan-4-olide, steroidal ester, b-sitosterol, b-sitosterol glucoside and ursolic acid isolated from C. polygonoides.9 The aim of present study was to isolate and identify the steroids from the roots of C. polygonoides. To the best of our knowledge, these steroids (1e4) were found for the first time from this species.
2.
Materials and methods
2.1.
Collection and identification of plant materials
Roots of C. polygonoides were collected from Village MehendriJo-Par (longitude: N 25 340 200 and latitude: E 70 110 2000 ), District Umerkot in Sindh Province of Pakistan in January 2012. A voucher specimen (15173) of the plant was deposited in the herbarium of Institute of Plant Sciences, University of Sindh Jamshoro, Pakistan. The plant sample was identified by a Taxonomist of the same institution. The plant material was air dried under normal conditions and ventilated.
2.2.
347
Extraction and isolation
About 300 g powdered roots of C. polygonoides were macerated in methanol for three days. Occasional shaking and stirring was done. Then extract was filtered using Whatman filter paper. The filtrate was concentrated to dryness under the vacuum. Chemical tests (Salkowski and LiebermanneBurchard reaction) were performed to detect the steroids in the extract.6 The dried methanol extract was subjected to column chromatography over silica gel (particle size 0.2e0.5 mm, 35e70 mesh ASTM) and gradient elutions were carried out with eluents chloroform, chloroformeethyl acetate mixtures and ethyl acetate. Each fraction was collected in 20 ml portion and was monitored by TLC (Pre-coated aluminum silica gel plates). The chloroform fraction was further purified by preparative TLC using hexane:chloroform (40:60) solvent system. TLC result shows the four spots with different retention time. Each spot (showing compound) was scratched separately and dissolved in hexane then filtered using Whatman filter paper. The isolated compounds were again confirmed of their identity by chemical tests. For further characterization UV, FT-IR and GCeMS was done.
2.3.
Gas chromatographyemass spectrometry (GCeMS)
GCeMS analysis of plant sample was performed on Agilent 6890 N GC instrument coupled with MSe5975 inert XL mass selective detector and auto sampler 7683-B injector was used. The HPe5MS column with dimensions of 30 m 0.25 mm i.d., film thickness 0.25 mm was used for the analysis. Initial temperature 150 C, maintained for 2 min, final temperature 230 C, kept for 5 min, ramp rate 4 C/min. 1.0 ml sample was injected, using split mode (split ratio, 10:1). Helium gas was used as a carrier gas at a flow rate of 0.8 ml/min. An electron ionization mode with ionization energy of 70 eV was used for MS detection. The injector and MS transfer line temperatures were set at 240 and 270 C, respectively.
2.4.
FT-IR analysis
FT-IR spectra were obtained using a Thermo Nicolet Avatar 330 FT-IR spectrometer controlled by OMNIC software (Thermo Nicolet Analytical instruments, Madison, WI, USA) station with a deuterated triglycine sulfate (DTGS) detector and KBr optics. The sampling station was equipped with overhead ATR accessory (Spectra-Tech, Shelton, CT) comprising of transfer optics with in the chamber through which infrared radiation is directed to a detachable ATR zinc selenide crystal mounted in a shallow trough for sample containment. A single beam spectrum (4000-650 cm1) of the sample was obtained against air as a background at a resolution of 4 cm1 and a total of 32 scan.11
3.
Results and discussion
The methanol extract of C. polygonoides roots was subjected to different phytochemical tests and it gives highly positive results for steroids. The extract was subjected to column chromatography over silica gel. The column was eluted in different solvent
348
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 3 4 6 e3 4 9
isolated compounds are identified as campesterol (1), stigmasterol (2), (3b,5a,24S)-stigmastan-3-ol (3) and stigmast-4en-3-one (4) (Fig. 1) from methanol extract of the roots of C. polygonoides.
HO
HO
Conflicts of interest Stigmasterol (2)
Campesterol (1)
All authors have none to declare. (C29H48O; Mol.Wt. 412.69)
(C28H48O; Mol.Wt. 400.68)
Acknowledgments Financial support and necessary facilities offered by National Centre of Excellence in Analytical Chemistry (NCEAC), University of Sindh, Jamshoro, Pakistan is gratefully acknowledged.
O
HO
Stigmast-4-en-3-one (4) (C29H52O; Mol.Wt. 416.72)
Fig. 1 e Sterols identified in roots of Calligonum polygonoides (1e4). system (CHCl3, CHCl3eEtOAc mixtures and EtOAc) with gradient elutions. Each fraction was monitored by TLC. The chloroform fraction was further purified by preparative TLC using hexane:chloroform (40:60) solvent system. The TLC result leads to the isolation of campesterol (1), stigmasterol (2), (3b,5a,24S)stigmastan-3-ol (3) and stigmast-4-en-3-one (4) (shown in Fig. 1). The FT-IR spectra of isolated compounds exhibit the diagnostic peaks relating to CeH stretching at 2950 cm1 and 2860 cm1. The OeH stretching and C]C absorption peak appears at 3360 cm1 and 1630 cm1, respectively. Other absorption peaks includes 1445 cm1 (CH2); 1371 cm1 (OH def), 1050 cm1 (cycloalkane) verify the required data regarding the structures of steroids. UV spectrum (recorded in methanol) of stigmasterol exhibited peak at 256 nm because it contains two isolated double bond. While, stigmast-4-en-3-one and campesterol exhibited peaks at 231 and 251 nm respectively. GCeMS is the most useful method for the characterization of steroids.12,13 Each compound was analyzed by GCeMS and identified by comparison of their mass spectra with the reference compounds in the data systems of Wiley and National Institute of Standards and Technology (NIST) spectra libraries matching. Compounds were identified with a resemblance percentage above 90%. Further conformation of these compounds was done by comparison of their and mass spectra with data in literature.14e19 Results show good agreement for the structure of campesterol (1), stigmasterol (2), (3b,5a,24S)-stigmastan-3-ol (3) and stigmast-4-en-3-one (4) as reported in the literature.
4.
references
(C29H48O; Mol.Wt. 412.69)
Conclusion
On the basis of chemical and spectral evidence and upon comparison of obtained data with the literature data, the
1. Kardong D, Upadhyaya S, Saikia LR. Screening of phytochemicals, antioxidant and antibacterial Activity of crude extract of Pteridium aquilinum Kuhn. J Pharm Res. 2012;5(11):5194e5196. 2. Sasidharan S, Chen Y, Saravanan D, Sundram KM, Latha LY. Extraction, isolation and characterization of bioactive compounds from plants’ extracts. Afr J Tradit Complement Altern Med. 2011;8(1):1e10. 3. Bhawani SA, Sulaiman O, Hashim R, Ibrahim MNM. Thinlayer chromatographic analysis of steroids. Trop J Pharm Res. 2011;9:301e313. 4. Choi JM, Lee EO, Lee HY, et al. Identification of campesterol from Chrysanthemum coronarium L. and its antiangiogenic activities. Phytother Res. 2007;21:954e959. 5. Gabay O, Sanchez C, Salvat C, et al. Stigmasterol: a phytosterol with potential anti-osteoarthritic properties. Am J Clin Nutr. 2010;18:106e116. 6. Panda S, Jafri M, Kar A, Meheta BK. Thyroid inhibitory, antiperoxidative and hypoglycemic effects of stigmasterol isolated from Butea monosperma. Fitoterapia. 2009;80:123e126. 7. Jamaluddin F, Mohamed S, Lajis MN. Hypoglycaemic effect of Stigmast-4-en-3-one from Parkia speciosa empty pods. Food Chem. 1995;54:9e13. 8. Batta AK, Xu G, Honda A, Miyazaki T, Salen G. Stigmasterol reduces plasma cholesterol levels and inhibits hepatic synthesis and intestinal absorption in the rat. MetabolismClinical Exp. 2006;55:292e299. 9. Samejo MQ, Memon S, Bhanger MI, Khan KM. Chemical composition of essential oil from Calligonum polygonoides Linn. Nat Prod Res; 2012. http://dx.doi.org/10.1080/14786419.2012. 686904. 10. Samejo MQ, Memon S, Bhanger MI, Khan KM. Preliminary phytochemical screening of Calligonum polygonoides Linn. J Pharm Res. 2011;4:4402e4403. 11. Burdi DK, Samejo MQ, Bhanger MI, Khan KM. Fatty acid composition of Abies pindrow (West Himalayan fir). Pak J Pharm Sci. 2007;20:15e19. 12. Radulovic NS, Dordevic ND. Steroids from poison hemlock (Conium maculatum L.): a GCeMS analysis. J Serb Chem Soc. 2011;76:1471e1483. 13. Lacikova L, Zapletal J, Masterova I, Grancai D. GCeMS analysis of leaves petrol ether extracts from four Staphylea L. species. Acta Facult Pharm Univ Comenianae. 2007;54:104e108.
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 3 4 6 e3 4 9
14. Jain PS, Bari SB. Isolation of lupeol, stigmasterol, caempsterol from petroleum ether extract of woody stem of Wrightia tinctoria. Asian J Plant Sci. 2010;9:163e167. 15. Kamboj A, Saluja AK. Isolation of stigmasterol and bsitosterol from petroleum ether extract of aerial parts of Ageratum conyzoides (Asteraceae). Int J Pharm Pharm Sci. 2011;3:94e96. 16. Yang B, Koponen J, Kallio RTH. Plant sterols in seeds of two species of Vaccinium (V. myrtillus and V. vitis-idaea) naturally distributed in Finland. Eur Food Res Technol. 2003;216:34e38.
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