Home | About PM | Editorial board | Search | Ahead of print | Current Issue | Archives | Instructions | Subscribe | Advertise | Contact us |  Login 
Pharmacognosy Magazine
Search Article 
  
Advanced search 
 


 
  Table of Contents  
ORIGINAL ARTICLE
Year : 2020  |  Volume : 16  |  Issue : 72  |  Page : 730-732  

Isolation of new diterpene from methanolic extract of Capsicum annuum Linn. fruits


1 Faculty of Pharmacy, AIMST University, Bedong, Kedah, Malaysia
2 Dr. K.N. Modi Institute of Pharmaceutical Education and Research, Modinagar, Ghaziabad, Uttar Pradesh, India
3 School of Pharmaceutical Education and Research, Jamia Hamdard, New Delhi, India
4 Department of Pharmacy, School of Medical and Allied Sciences, Galgotias University, Greater Noida, Uttar Pradesh, India

Date of Submission10-Jun-2020
Date of Decision06-Jul-2020
Date of Acceptance15-Sep-2020
Date of Web Publication16-Feb-2021

Correspondence Address:
Pradeep Kumar Sharma
Dr. K.N. Modi Institute of Pharmaceutical Education and Research, Modinagar, Ghaziabad, Uttar Pradesh
India
Login to access the Email id

Source of Support: None, Conflict of Interest: None


DOI: 10.4103/pm.pm_250_20

Rights and Permissions
   Abstract 


Background: Capsicum annuum (CA) fruits are consumed in the human diet for more than six centuries. Less research evidence for critical isolation experiment over CA fruits methanolic extract (CAFME) is an important concern for the investigators. Objectives: The present investigation was intended to explore the CAFME for the presence of any new phytoisolate. Materials and Methods: The study involved the preparation of CAFME, followed by critical isolation through normal phase silica-gel based column chromatography using various combinations of petroleum ether and chloroform. The phytoisolate was characterized using ultraviolet, Fourier transformed infrared, Nuclear magnetic resonance (1H, 13C, Distortion less enhancement by polarization transfer and correlation spectroscopy), and mass spectrometry. Results: The chromatographic critical isolation and spectrometric experiment over CAFME offered a new phytoisolate characterized as 3,11,15-trimethyl-14 β-hydroxy-n-hexadeca-7-en-4,18-olide (CA-1). Conclusion: The present study concludes the isolation of a new diterpene 3,11,15-trimethyl-14 β-hydroxy-n-hexadeca-7-en-4,18-olide (CA-1) for the first time in CAFME. The present study also recommends that in future this new phytoisolate should be further standardized and explored for its therapeutic properties.

Keywords: Capsicum, characterization, chromatography, diterpene, phytoisolate


How to cite this article:
Fuloria NK, Fuloria S, Sharma VK, Ali M, Singh A, Sharma PK. Isolation of new diterpene from methanolic extract of Capsicum annuum Linn. fruits. Phcog Mag 2020;16:730-2

How to cite this URL:
Fuloria NK, Fuloria S, Sharma VK, Ali M, Singh A, Sharma PK. Isolation of new diterpene from methanolic extract of Capsicum annuum Linn. fruits. Phcog Mag [serial online] 2020 [cited 2021 Mar 9];16:730-2. Available from: http://www.phcog.com/text.asp?2020/16/72/730/309310



SUMMARY

  • In the present study, the critical column chromatography of methanolic extract of Capsicum annuum offered a new diterpene (CA-1), that was spectrometrically characterized as 3, 11, 15-trimethyl-14 β-hydroxy-n-hexadeca-7-en-4,18-olide (CA-1). This phyotoisolate is reported for the first time in Capsicum annum fruits methanolic extract.


Abbreviations used: CA: Capsicum annuum; CAFs: Capsicum annuum fruits; CAFME: Capsicum annuum fruits, extract; COSY: Correlation spectroscopy; d: Doublet; DEPT: Distortion less enhancement by polarization transfer; 1D: One dimension; 2D: Two dimension; FABMS: Fast atomic bombardment mass spectrometer; FTIR: Fourier transformed infrared; g: Gram; Kg: Kilogram; m: Multiplet; MHz: Mega Hertz; m/z: Mass to charge ratio; λmax: Maximum wavelength; NMR: Nuclear magnetic resonance; No.: Number; Rf: Retention factor; S: Singlet; Silica gel G: Silica gel gypsum; TLC: Thin layer chromatography; UV: Ultraviolet.




   Introduction Top


Capsicum annuum (CA) Linn. belonging to the genus capsicum with Solanaceae family has been consumed in the human diet for more than six centuries.[1] It is a perennial herb that is commonly found in central and south America. Based on flavor and size, the CA is called with several names. The larger and sweeter varieties of CA are called sweet pepper in the UK and red or green or bell pepper in the USA; whereas the smaller and hotter varieties are known as chile, chillis, or chili peppers.[2] The CA Linn. is an annual herbaceous plant that possesses glabrous, lanceolate leaves, white color fruits, and flowers.[3] The CA fruits (CAFs) are commonly used in food preparations as spices attributed to their pungent taste (due to capsaicin presence in fruits, seeds, and placental tissue). The CAFs are indicated for various medicinal purposes, such as stomach pain, asthma, gout, arthritis, anorexia, seiatica, dyspepsia, flatulence, cardiac debility, cough, malaria, cholera, muscle spasm, neuralgia, lumbago, and chilblains.[4],[5] The CAFs are reported to contain capsaicinoids (0.1%–2%), that are responsible for characteristic pungent taste and possess high therapeutic value in gastric ulcer and rheumatoid arthritis.[6],[7] Study suggests most varieties of CAFs to comprise key capsaicinoids like capsaicin and dihydrocapsaicin (1:1); and few capsaicinoids like homodihydrocapsaicin, homocapsaicin, and nordihydrocapsaicin (ranging from 1% to 38%).[8] The crude extract of CAFs known as capsicum oleoresin contains at least 100 different volatile chemical constituents.[9] Other chemical constituents found in CAFs include carotenoids (like capsanthin, capsorubin, carotene, lutein, etc.), saponin alkaloid (capsicidin), fats (9%–17%), protein (12%–15%), vitamins (A, B and C), xanthins (Cycloviolaxanthin, (8S)-capsochrome, 5,6-epoxycapsanthin, karpoxanthin, cucurbitaxanthin A and B, violaxathin, 3,6-epoxycapsanthin), steroidal alkaloidal glycosides (solanine, solanidine, solasodine), steroidal glycosides (capsicoside A-D), coumarins (Scopoletin, a coumarin) and volatile oils.[10],[11] Although several studies reported the presence of a wide range of phytochemicals in CAFs using various solvents system, still very less research data is available over the critical isolation of CAFs methanolic extract (CAFME) using narrow range of eluting solvents system, especially over petroleum ether-chloroform. Based on these facts, it was hypothesized that by performing critical isolation over CAFs, the present study would explore some newer phytoisolate. Hence based on these findings, the present study was designed to prepare methanolic extract of CAFs, followed by its critical isolation (by normal phase column chromatography using various combinations of petroleum ether and chloroform) and spectrometric characterization (using ultraviolet (UV), Fourier transformed infrared (FTIR), Mass and 1D and 2D nuclear magnetic resonance [NMR] spectroscopy).

Experimental

General

The reagents, solvents, and chemicals used in the present study were of analytical standard, obtained from Merck and Sigma Aldrich and were used without purification. The normal phase column chromatography of phytosiolate involved the use of Silica gel G (60–120 mesh). The thin-layer chromatography (TLC) of phytoisolate was performed over silica gel G plates. The UV spectrum of phytoisolate was recorded at 200–400 nm using Shimadzu UV-160A UV visible spectrophotometer. Fourier transformed infra-red spectrum was recorded at 500–4000 cm-1, using Bruker FTIR spectrometer The 1D and 2D NMR spectra (1H,13C, Distortion less enhancement by polarization transfer [DEPT] and correlation spectroscopy [COSY]) were recorded on Avance 300 MHz spectrometer using CDCl3 (solvent) and TMS (internal standard) expressing coupling constants in Hertz. The mass spectrum was recorded using fast atomic bombardment mass spectrometer (FABMS) spectrometer.

Plant material and extract preparation

The CAFs were procured from the province of Ghaziabad, Uttar Pradesh, India, and were authenticated by Dr. K. C. Bhatt, National Bureau of Plant Genetic Resources, Pusa Campus, Delhi, India. A voucher specimen (NHCP/NBPGR/2009/2/551) of CAFs was deposited in the herbarium of the Department of Pharmacognosy, R. V. Northland Institute, Dadri, Greater Noida, Uttar Pradesh, India, for future reference. The CAFs were collected, air-dried under the shade, and powdered. The dried powder of CAFs (1 kg) was subjected to exhaustive extraction with 95% methanol in a Soxhlet apparatus for 50 h. The obtained CAFs crude was concentrated with rotary evaporator to offer 96 g of dark brown CAFME.

Isolation and purification

The CAFME (90 g) was subjected to critical isolation by dissolving in a minimum amount of methanol in a China dish and then adsorbed on silica gel (60–120 mesh) slowly for preparation of a slurry. The CAFME was air-dried, powdered and passed through sieve (No. 8) to get uniform particle size. A clean dried column plugged on the lower side with a piece of nonabsorbent cotton was fitted in vertical position on a stand. Column was then half filled with petroleum ether. Silica gel for column chromatography (60–120 mesh) was then poured in small portions and allowed to settle down to form the stationary phase. The dried CAFME slurry was loaded over the column and elution of the column was carried out successively with various combinations of petroleum ether:chloroform (10:90, 9:91, 8:92, 7:93, 6:94, 5:95, 4:96, 3:97, 2:98, 1:99, 0:100). The homogeneity of collected fractions was checked using TLC. The fractions having the same retention factor (Rf) values were combined and concentrated. The concentrate was purified using a suitable solvent system to offer CA-1. The purified phytoisolate was subjected to UV, FT-IR, NMR (1H, 13C, DEPT, and COSY) and FAB-Mass spectrometric characterization studies. The structure of phytoisolate CA-1 was established based on the physical and characterization data.


   Results Top


The isolation experiment offered a new isolate CA-1 using petroleum ether and chloroform (2:98) solvent system. Results of physical and characterization studies over CA-1 are presented as follows:

Physical data

Quantity: 0.0018% (167 mg from 90 g CAFME); colour: pale yellow; state: semisolid; eluent system: 9th fraction of petroleum ether and with Chloroform (2:98); Rf value: 0.46 (chloroform:methanol, 3:1).

Characterization data

UV maximum wavelength (MeOH): 229 nm; IR νmax (KBr) in cm-1: 3409, 2926, 2846, 1733, 1645, 1442, 1383, 1256, 1048 cm-1; 1H NMR (CDCl3) δ value in Hz: 0.85 (3H, d, J = 6.4, CH3-1), 0.99 (3H, d, J = 6.8, CH3-19), 1.03 (3H, d, J = 6.6, CH3-17), 1.10 (3H, J = 6.7, CH3-20), 1.15 (3H, d, J = 6.5, CH3-16), 1.19–1.80 (9H, m, CH-5 and 4 x CH2), 2.01 (2H, m, CH2-5), 2.05 (1H, m, CH-3α), 2.23 (1H, m, CH-11), 2.36 (2H, m, CH2-9), 2.72 (2H, m, CH2-6), 3.60 (1H, m, CH-14α), 4.06 (1H, m, CH-4α) and 5.3 (1H, m, CH-8); 13C NMR (CDCl3) δ value in Hz: 14.5 (C1), 22.2 (C2), 39.6 (C3), 78.9 (C4), 58.3 (C5), 50.0 (C6), 140.2 (C7), 120.2 (C8), 30.5 (C9), 30.6 (C10), 42.7 (C11), 21.2 (C12), 21.4 (C13), 68.2 (C14), 31.5 (C15), 18.3 (C16), 16.5 (C17), 166.4 (C18), 20.2 (C19), 18.3 (C20); FABMS m/z: 324 [M] + (C20H36O3), 86 (base peak), 281, 267, 251, 239, 157, 129, 57.


   Discussion Top


The present study was intended to explore any new phytoisolate present in the CAFs. For this 90 g of prepared CAFME was successively eluted with petroleum ether and chloroform using normal phase column chromatography. This yielded 10 fractions (each fraction of 500 ml) each with the following eluent system (chloroform:methanol ratio): fraction 1–5 (10:90), fraction 6–10 (9:91), fraction 11–15 (8:92), fraction 16–20 (7:93), fraction 21–25 (6:94), fraction 26–30 (5:95), fraction 31–35 (4:96), fraction 36–40 (3:97), fraction 41–45 (2:98), fraction 46–50 (1:99), fraction 51–55 (0:100). The fractions 46–50 were combined and exposed to TLC, which offered a new spot with Rf value of 0.46 (chloroform:methanol, 3:1). The fraction 46–50, when dried and subjected to preparative TLC, offered semisolid pale yellow mass CA-1. Based on the characterization techniques used in other standard investigations present study involved characterization of phytoisolate (CA-1) using UV, FT-IR, NMR (1H, 13C, DEPT, and COSY) and FAB-Mass spectrometric characterization studies.[12],[13],[14] Characteristic IR bands at 3411, 1737, and 1646 cm-1 in CA-1 FTIR spectrum revealed the presence of OH, δ-lactone, and unsaturation, respectively in CA-1. The FAB mass spectrum of CA-1 exhibiting M+ ion signal at m/z 324 established its molecular formula as: C20H36O3. The presence of fragmented ions signals at m/z 281, 251, 239, and 129; corresponding to C14-C15⌉+, C13-C14⌉+, C11-C12+, and C10-C11+ fission fragment ions revealed the presence of OH group at C14 position. The fragment ion signals at m/z 57, 267, 157 attributed to C3-C4+ fission and C8-C9+ fission fragment ion revealed the presence of δ-lactone nucleus at C4 (18) and vinyl linkage at C7. The 1H NMR spectrum of CA-1 exhibited one three-proton triplet at δ 0.85 and showed the presence of primary C1 methyl protons. Four three proton doublet signals at δ 0.99, 1.03, 1.10, and 1.15 revealed the presence of C19, C17, C29, and C16 secondary methyl protons, respectively. The two one proton multiplets at δ 3.67 and 4.06 revealed the presence of H-14α carbinol and H-4α-oxymethine protons. One proton multiplet signal at δ 5.35 showed the presence vinylic H-8 proton. Remained methylene and methine protons exhibited signals from δ 1.19 to 2.72. The 13C NMR spectrum of CA-1 exhibited characteristic signals at δ 68.19, 78.85, and 166.42 attributed to the presence of C14 carbinol, C4 oxymethine and C18 lactone carbons. The signals at δ 120.15 and 140.22 were attributed to C8 and C7 vinylic carbons. The spectrum signaled for methyl carbons at δ 14.48, 16.51, 18.27, 18.33, 20.17, corresponding to C1, C17, C20, C16, and C19, respectively. The DEPT spectrum of CA-1 also supported the presence of 05 methyl, 07 methylene, 06 methine, and 02 quaternary carbons. The 1H-1H COSY spectrum of CA-1 showed the characteristic correlation of H2-5 with H2-6; H4 and H3; H8 with H2-6 and H2-9; and correlation of H14 with H2-13, H15, H3-16, and H3-20. The confirmation of spectrometric characterization data results of the present study was based on the agreement with the results of other standard findings.[15],[16] Based on spectrometric characterization data, the structure of phytoisolate CA-1 is elucidated as 3, 11, 15-trimethyl-14 β-hydroxy-n-hexadeca-7-en-4,18-olide [Figure 1]. This diterpene lactone is reported for the first time in CAFME.
Figure 1: Chemical structure of Capsicum annuum-1

Click here to view



   Conclusion Top


The findings of the present research conclude and reports the isolation of a new diterpenic lactone 3, 11, 15-trimethyl-14 β-hydroxy-n-hexadeca-7-en-4,18-olide for the first time in the methanolic extract of CA Linn. fruits. The present study also recommends that in future this new phytoisolate should be further standardized and explored for its therapeutic properties.

Acknowledgements

The authors are sincerely thankful to the RV Northland Institute; Galgotias University, IIT Delhi, India; and AIMST University, Malaysia, for providing necessary facilities and support for successful completion of the present study.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
   References Top

1.
Gebhardt C. The historical role of species from the Solanaceae plant family in genetic research. Theor Appl Genet 2016;129:2281-94.  Back to cited text no. 1
    
2.
Lin S, Chou Y, Shieh H, Ebert AW, Kumar S, Mavlyanova R, Rouamba A, et al. Pepper (Capsicum spp.) germplasm dissemination by AVRDC–the world vegetable center: An overview and introspection. Chron Hort 2013;53:21-7.  Back to cited text no. 2
    
3.
Sunil P, Sanjay Y, Vinod S. Pharmacognostical investigation and standardization of Capsicum annuum L. roots. Int J Pharmacogn Phytochem Res 2012;4:21-4.  Back to cited text no. 3
    
4.
Vijayalakshmi K, Shyamala R, Thirumurugan V, Sethuraman M, Rajan S, Badami S, et al. Physico-phytochemical investigation and anti-inflammatory screening of Capsicum annuum L. and Hemidesmus indicus Linn. R. Br. Anc Sci Life 2010;29:35-40.  Back to cited text no. 4
    
5.
Bosland PW. Capsicums: Innovative uses of an ancient crop. In: Progress in New Crops. Arlington VA: ASHS Press; 1996. p. 479-87.  Back to cited text no. 5
    
6.
Matucci-Cerinic M, Marabini S, Jantsch S, Cagnoni M, Partsch G. Effects of capsaicin on the metabolism of rheumatoid arthritis synoviocytes in vitro. Ann Rheum Dis 1990;49:598-602.  Back to cited text no. 6
    
7.
Satyanarayana MN. Capsaicin and gastric ulcers. Crit Rev Food Sci Nutr 2006;46:275-328.  Back to cited text no. 7
    
8.
González-Zamora A, Sierra-Campos E, Luna-Ortega JG, Pérez-Morales R, Rodríguez Ortiz JC, García-Hernández JL. Characterization of different capsicum varieties by evaluation of their capsaicinoids content by high performance liquid chromatography, determination of pungency and effect of high temperature. Molecules 2013;18:13471-86.  Back to cited text no. 8
    
9.
Cordell GA, Araujo OE. Capsaicin: Identification, nomenclature, and pharmacotherapy. Ann Pharmacother 1993;27:330-6.  Back to cited text no. 9
    
10.
Gupta D. An overview of capsicum. Int J Pharm Biol Sci Arch 2015;3:7-11.  Back to cited text no. 10
    
11.
Nong S, Yang X, Li D, Yang L, Xu Z, Chen Y, Jiejie L. Investigation of Capsicum annuum var. conoides in east central rural areas of Hainan province [J]. Res Sci 2010;32:2400-6.  Back to cited text no. 11
    
12.
Rahman SM, Pervin S, Quader MA, Hossain MA. Phytochemical studies of the petroleum ether extract of the leaves of Lagerstroemia speciosa Linn. Ind J Chem 2009;9:500-4.  Back to cited text no. 12
    
13.
Rahmana SM, Muktaa ZA, Hossainb MA. Isolation and characterization of β-sitosterol-D-glycoside from petroleum extract of the leaves of Ocimum sanctum L. As. J. Food Ag-Ind 2009;2:39-43.  Back to cited text no. 13
    
14.
Jutiviboonsuk A, Zhang HJ, Kondratyuk TP, Herunsalee A, Chaukul W, Pezzuto JM, et al. Isolation and characterization of cancer chemopreventive compounds from Barringtonia maunwongyathiae. Pharm. Biol. 2007;45:185-194.  Back to cited text no. 14
    
15.
Fuloria NK, Fuloria S. Spectroscopy Fundamentals and Data Interpretation. New Delhi: Studium Press; 2013.  Back to cited text no. 15
    
16.
Sultana S, Ali M, Mir SR. Chemical constituents from the roots of Oenothera biennis L. Pharma Biosci J 2018;6:29-35.  Back to cited text no. 16
    


    Figures

  [Figure 1]



 

Top
   
 
  Search
 
    Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
    Access Statistics
    Email Alert *
    Add to My List *
* Registration required (free)  

 
  In this article
    Abstract
   Introduction
   Results
   Discussion
   Conclusion
    References
    Article Figures

 Article Access Statistics
    Viewed230    
    Printed0    
    Emailed0    
    PDF Downloaded45    
    Comments [Add]    

Recommend this journal