VARUT Renata-Maria1, BELU Ionela1
1University of Medicine and Pharmacy of Craiova, Pharmacy, 2-4 Petru Rares Str., 200349, Craiova, Romania
rennata_maria@yahoo.com, ionelabelu@gmail.com
Abstract
The presence of specific functional groups to certain classes of compounds, guides us on the potential therapeutic effects, being able to make a prediction on the pharmacological effects of vegetable extracts. FT-IR is most useful in providing information about the presence or absence of specific functional groups and can provide a molecular fingerprint that very useful when comparing samples composition. In the present study we realised the qualitative characterization of the vegetable extracts from the specific FT-IR vibrations, for a series of six vegetable extracts: Acanthi balcanici herba (ABH), Cardui acanthoiditis folium (CAF), Dorycnii pentaphylli herba (DPH), Tamaricis ramosissimae folium et flos (TRFF), Tragoponis pratensis folium (TPF), Myrtilli folium (M-fol.), Myrtilli fructus (M-fr.). Vibrations from the FT-IR spectra of vegetable extracts are attributed to polyphenol compounds (flavonoids, polyphenolcarboxylic acids), carotenoids (β-carotene, carotenoid esters), triglycerides, phytosterols, amino acids.
Keywords: IR, hypoglycemic extracts
Introduction
The IR spectrum is basically a plot of transmitted (or absorbed) frequencies vs. intensity of the transmission (or absorption). Frequencies appear in the x-axis in units of inverse centimeters (wavenumbers), and intensities are plotted on the y-axis in percentage units. Although the entire IR spectrum can be used as a fingerprint for the purposes of comparing molecules, the 600 – 1400 cm-1 range is called the fingerprint region. This is normally a complex area showing many bands, frequently overlapping each other. Not all covalent bonds display bands in the IR spectrum, only polar bonds do so, being referred to as IR active. The intensity of the bands depends on the magnitude of the dipole moment associated with the bond in question. The first IR spectral analysis of the residue obtained by evaporation of some plant extracts (maceration in petroleum ether) were carried out for the first time by Grlić L. & Grlić Scaroni. Based on the differences between fingerprint vibration transmissions, the authors have drawn up a scheme for the characterization of Cannabis sativa var. indica from different regions [].
In the present study we proposed the qualitative characterization of the vegetable extracts from the specific vibrations in IR point of view, for a series of six vegetable extracts: Acanthi balcanici herba (ABH), Cardui acanthoiditis folium (CAF), Dorycnii pentaphylli herba (DPH), Tamaricis ramosissimae folium et flos (TRFF), Tragoponis pratensis folium (TPF), Myrtilli folium (M-fol.), Myrtilli fructus (M-fr.).
Materials and methods
The analyzed plant materials were harvested from “Alexandru Buia” University Botanical Garden Craiova species, during 2011-2014. For each of the investigated plant products in the Collection of the Pharmacognosy Laboratory of the Faculty of Pharmacy, U.M.F. Craiova are stored witness samples harvested in different periods.
The vegetable extracts were obtained by simple percolation, in a vegetable / solvent ratio (ethanol 70o) of 1: 5 (F.R. X). Vegetable products naturally dried and pulverized have been made to the specific sieve degree of fineness IV, using a electrical mill-type grinder.
IR spectra of residues obtained by evaporation of vegetable extracts were determined under the following experimental conditions:
▪ apparatus: spectrometer FT-IR Nicolet–Avatar ESP 360;
▪ source of IR radiation, with the field of work 4000–400 cm-1: sample spectra were recorded in the field 4000–800 cm-1;
▪ detector DTGS KBr;
▪ conditioned samples in KBr, achieved by compression until a translucent film;
▪ computerized data analysis software EZ Omnic E.S.P. 5.1 / 32 bit.
Results and discussions
The FT-IR spectral analysis was performed on the six plant extracts to guide our future research. The presence of functional groups specific to certain classes of compounds guides us on the potential therapeutic effects, being able to make a prediction on the pharmacological effects of plant extracts.
Secondary plant metabolites can be identified by FT-IR due to the vibration characteristic of certain functional groups in their structure. Thus, by this method, compounds belonging to the classes can be identified: phenolic acids, flavonoids, terpenoids, alkaloids, iridoids, chlorophylls, phytosterols, fatty acids []. Table 1 and Figures 1-6 show vibrations obtained from the FT-IR analysis of vegetable extracts compared to the extraction solvent (ethanol, Figure 7).
Table 1. Results of FT-IR analysis of vegetable extracts
| Sample / Characteristic Frequencies in IR [cm-1] | Assignment | ||||||
| ABH | CAF | DPH | TRFF | TPF | M-fr. | Etanol | |
| 3418 | 3442 | 3439 | 3408 | 3442 | 3432 | 3432 | O-H and N-H stretching vibrations (amino acids, amines, amides). |
| 2933 | 2974 | – | 2929 | 2981 | 2981 | – | ν C-H (asym.) (Lipids, fatty acids). |
| – | 2929 | 2936 | 2929 | 2929 | – | – | Asymmetric stretching vibrations -CH3, -CH2- (carboxylic acids). |
| – | – | – | 2858 | – | – | – | The symmetrical stretching vibration of C-H from the group -CH3 (fatty acids). |
| – | – | – | 2331, 2355 | – | 2355 | 2355 | C≡C triple bond. Vibration of stretching of the P-H group (phosphorus compounds). |
| – | – | – | 1723 | – | – | – | Stretching vibration of C = O (ester). |
| 1630 | 1647 | 1634 | 1613 | 1630 | 1651, 1630 | 1651, 1630 |
Vibration of the C-O group of the secondary amides. |
| 1490 | 1504 | 1511 | 1446, 1511 | 1452 | – | – | ν C-H (bending) (the -CH2 group in proteins). |
| 1401 | 1384 | 1350 | 1350 | 1385 | 1381 | 1347 | Deformation vibrations (C-H) of -CH3, -CH2 groups in aliphatic compounds. |
| – | 1258 | 1203 | 1220 | 1254 | 1278 | 1257 | ν C-O, NO-H def. (polyphenolic compounds). |
| – | – | – | – | 1169 | – | – | ν C-O-C, ring. |
| 1080 | – | 1073 | – | 1076 | 1083 | – | νC-H (deformation),νC-O, νC-C (stretch) – carbohydrates. |
| – | 1042 | 1046 | 1042 | 1042 | 1049 | 1042 | νC-O (stretch) polysaccharides. |
| 772 | 878 | – | 837 | – | – | – | νC-H (bending out of the plane). |
| – | – | – | 532, 666 |
618 | 550 | 550 | Vibration skeleton molecule. |
Vegetable extracts: ABH – Acanthi balcanici herba; CAF – Cardui acanthoiditis folium; DPH – Dorycnii pentaphylli herba; TRFF – Tamaricis ramosissimae folium et flos; TPF – Tragoponis pratensis folium; M-fr. – Myrtilli fructus.
They were analyzed particular strip of 750-3500 cm-1 region, knowing that the region reflects the biochemical composition of plant [].
Distinctive peak at 1630-1650 cm-1 was attributed to the stretching vibration of C = O group in the secondary amides, while the peak at 1340-1350 cm-1 corresponding to the bending vibrations (C-H) groups -CH3 , -CH2- in the aliphatic compounds.
The peak at 1723 cm-1 is due to the stretching vibration of the C = O group, which may be part of some secondary compounds molecules, of the triglyceride classes or more likely of the carotenoid esters [].
The peak at 1377 cm-1 appears in the literature as deformation characteristic vibration (C-H) groups -CH 3, -CH 2 from β-carotene [].
In the region of 3400-3450 cm-1 was observed the appearance of broad band vibration representative for -OH or -NH- groups, which may be derived from the polyphenolic compounds, amino acids, amines, amides.
Peaks in the range 1220-1260 cm-1 are representative for symmetric C-O and O-H vibrations, which could also be due to the polyphenols in the samples. At frequencies 2930-2980 cm-1, peaks are characteristic for a class more difficult to de characterize by FT-IR but whose presence has been identified previously, namely phytosterols.
From the IR spectra of residuals obtained from vegetable extracts evaporation, were removed the characteristic vibrations of chlorophyll [cm-1]: 2925(i) – νCH2as. (–CH2–CH2–); 2854(i) – νCH2sim. (–CH2–CH2–); 2346(s) – νC=NH* (the “ammonium” band); 1670(m) – νC=O (ketones α-, β-unsaturated); 1654(m) şi 1518(m) – νC=N (–CH=N–, pirol); 1261(m) – νC-O (–C=C–COOH) [].
| Figure 1 – FT-IR spectrum of the ABH (Acanthi balcanici herba) sample. |
| Figure 2 – FT-IR spectrum of the CAF (Cardui acanthoiditis folium) sample. |
Figure 3 – FT-IR spectrum of the DPH (Dorycnii pentaphylli herba) sample.
| Figure 4 – FT-IR spectrum of the TRFF (Tamaricis ramosissimae folium et flos) sample. |
| Figure 5 – FT-IR spectrum of the TPF (Tragoponis pratensis folium) sample. |
| Figure 6 – FT-IR spectrum of the M-fr. (Myrtilli fructus) sample. |
| Figure 7 – FT-IR spectrum of the extraction solvent (ethanol). |
In literature we have found articles on the pharmacognostic characterization of all the plants under study, with the exception of Acanthus balcanicus. For Dorycnium pentaphyllum subsp. herbaceum, Georgios Kazantzoglou et al. in 2003, isolates and elucidates the structure of doricnioside, a new phenylbutanone glucoside, together with another 2 known phenybutanone glucosides, five flavonoids, a cyanogenic glucoside, a cyclitol and a hydroquinone glucoside [].
In 2012, the chemical composition of the Tamarix ramosissima plant species was partially discovered by Mao Z.J., the extract containing tamarixetin, eugenol, vanillin, ferulic acid and polyphenolic acids [].
The chemical composition of Tragapogon pratensis has been elucidated since 1992 by spectral and chemical techniques: it contains nine triterpenic saponosides, called tragoponoids A-I, along with five other triterpenic glycosides [].
The chemical composition of Carduus acanthoides plant species was highlighted in 2009 following physico-chemical determinations and contains flavones (apigenin and luteolin) and saturated fatty acids [].
For Vaccinium myrtillus L. in a study in Poland published in 2010, plants harvested from 11 natural habitats show that although they contain active principles such as arbutin, polyphenolic acids in the leaves and anthocyanosides and organic acids in fruits sometimes differ significantly in the amount of their habitat. It influences quality and causes problems in standardization. The increased level in arbutin causes strong antiseptic and anti-inflammatory effects. In some areas of Poland fruit has a high content of anthocyanin dyes [].
Conclusions
In order to identify the chemical composition, vegetable extracts were first characterized in terms of specific vibrations in IR. Vibrations from the FT-IR spectra of vegetable extracts are attributed to polyphenol compounds (flavonoids, polyphenolcarboxylic acids), carotenoids (β-carotene, carotenoid esters), triglycerides, phytosterols, amino acids. In order to identify, the vegetable extracts were first characterized in terms of specific vibrations in IR.