Docstoc

ANTIOXIDANT ACTIVITY OF ESSENTIA

Document Sample
ANTIOXIDANT ACTIVITY OF ESSENTIA Powered By Docstoc
					                                          Bulletin of the Chemists and Technologists of Macedonia, Vol. 20, No. 1, pp. 61-66 (2001)
GHTMDD - 376                                                                                                     ISSN 0350 - 0136
Received: October 13, 2000                                                                         UDC: 582.929.4-113.55 (497.7)
Accepted: March 22, 2001
                                                                                                                  Short communication




               ANTIOXIDANT ACTIVITY OF ESSENTIAL OILS OF DIFFERENT
                             WILD THYMUS L. SPECIES

                               SvetIana Kulevanova1*, Tatjana Kadifkova Panovska2
                     IJnstitute of Pharmacognosy and 2Jnstituteof Toxicological Chemistry,
                    Faculty of Pharmacy, Vodnjanska 17, 1000Skopje, Republic of Macedonia

             Antioxidant activity of fifteen essential oils of Thymus L. (Lamiaceae) that are growing wild in Republic of
       Macedonia was investigated. Chemical composition of the oils, obtained by hydrodistillation of dried herbs, were de-
       termined by GC and GC-MS methods. Samples of oils were divided into six groups denoted as oils rich in: thymol,
       thymol+carvacrol, carvacrol, linalool+citral, linalool+carvacrol and linalool+geraniol+carvacrol. Antioxidant effects
       of these oils on thermal autooxidation of lard at 60 QCwere tested by measuring the content of produced peroxides
       during the period of 16 days. Antioxidant effects of oils were strongly dependent on the content of phenolics, follow-
       ing this order: carvacrol> thymol+carvacrol > thymol. Phenolic-rich essential oils were obtained from T. tosevii, T.
       tosevii var. degenii, T. tosevii var. longifrons, T. tosevii subsp. substriatus and T. longidens var. lanicaulis, but from
       the specimens of herbs collected in very particular area. Essential oils from other samples of Thymus, from the above
       mentioned or other species, with different origin, contained trace amounts of carvacrol and/or thymol and possessed
       low potential for antioxidant activity.

       Key words: Thymus; wild thyme; essential oils; antioxidant activity; peroxide value



                                                        INTRODUCTION

      Thymus vulgaris (thyme) (like some other                          antioxidant activities of combination of plant ex-
wild Thymus species (wild thyme)) posseses a wide                       tracts of thyme, oregano, marjoram and sage with
range of biological activities including expectorant,                   primary antioxidants or citric acid in lard stored at
spasmolitic, sedative [1-4], antibacterial [3], anti-                   75 DC,has found minor synergistic efficiency of
fungal [5], antioxidant [6-12] etc. In general, in-                     citric acid with thyme extracts [8]. Deighton et al.
vestigations of biological activity in most of the                      [9] investigated formation of stable free radicals
studies were carried out on extracts, essential oils or                 upon reaction of the essential oils of ten plant spe-
pure compounds isolated from dried herbs of Thy-                        cies with ultraviolet radiation and the superoxide
mus specIes.                                                            radical anion, by electron paramagnetic resonance
      Nowadays, there is great world-wide interest                      (EPR) spectroscopy. His research group found that
in finding new and safe antioxidants from natural                       oils obtained from oregano, savory and thyme pro-
sources, to prevent oxidative deterioration of foods                    duced EPR spectra accounted for free radicals pro-
and to minimise oxidative damage of living cells.                       duced by phenolics, carvacrol and thymol. The
Within Lamiaceae species, examples of new anti-                         conclusion was based on previously known anti-
oxidants include phenolic diterpenes, phenolic car-                     oxidant properties of these molecules [9, 10].
boxilic acids, biphenils, and flavonoids isolated                       Deans et at. [11] found that volatile oils from
from rosemary, sage, oregano and thyme [6, 7]. In                       thyme were the effective in the protection of poly-
many cases phenolic compounds have antioxidant                          unsaturated fatty acids in mice liver after ingestion
activities more effective than ex-tocopherol and ac-                    of volatile oil obtained by hydrodistillation.
tivity comparable to that of synthetic antioxidants,
                                                                        Madsen et al. [12] found that total phenol content
BRA and BRT [6, 7]. Banias et al. [8], studying                         in the extracts of different Lamiaceae species (in-
62                                        S. Kulevanova,   T. Kadijkova Panovska



cluding thyme) correlated linearly with the anti-                 tivities such as antioxidant should be investigated.
oxidant activity as measured by oxygen depletion,                 In our previous work was found that ether and
but not with the free radical scavenging effects.                 ethylacetate extracts of four wild Thymus species
The results obtained showed that investigated ex-                 possessed antioxidant activity in lard. All investi-
tracts contain components with at least two differ-               gated extracts contained various contents of fla-
ent antioxidative mechanisms [12].                                vonoids. The correlation between the antioxidant
      All studies of antioxidant activity of Thymus               activity and the flavonoid content was poor, mean-
were carried out upon the extracts, essential oils or             ing that, b~sides flavonoids, other components in
isolated compounds of thyme (Thymus vulgaris).                    the extracts were also responsible for the antioxi-
In available literature no data for antioxidant activ-            dant activity [13]. These components could be
ity of wild Thymus species can be found. Chemical                 phenol acids or phenolics from the essential oils. In
composition of wild thyme species is similar to                   the present work antioxidant activity of wild thyme
that of thyme but it is not identical and certain ac-             essential oils was investigated.


                                                EXPERIMENTAL

                     Materials                                    in our investigation were: thymol, carvacrol, gera-
                                                                  niol, linalool and citral, present in different quanti-
       Fifteen essential oils were obtained by steam              ties. According to the abundance of these compo-
distillation in Klevenger type apparatus using                    nents, the oils were divided into six groups denoted
standard procedure [14]. Air dried herbal parts of                as: "thymol + carvacrol", "thymol", "carvacrol",
plants were cut to pieces and put into the distilla-              citral + linalool", linalool + carvacrol" and "gera-
tion balloon, mixed with water (l: 10, m/V) and                   niol + linalool + carvacrol" type of oils. The con-
distilled in duration of 3 h. The water was sepa-                 tent of the components as well the biological origin
rated from the supernatant oil and the oil was dried              of the oils are presented in Table 1.
over anhydrous Na2S04. After filtration oils were
stored in small, closed, glass containers at low                                              Chemicals
temperature (in refrigerator). Small parts of the oils
were separated and dissolved in ethanol (1.0 %
                                                                       Butyl hydroxyl anisole (BHA), butyl hy-
solutions) for determination of the chemical com-
                                                                  droxyl toluene (BHT) and thymol were purchased
position. Analysis of the oils were performed by
                                                                  from Merck (Germany). All other chemicals, pur-
GC-FID on fused silica capillary column PONA
                                                                  chased from Alkaloid (Macedonia), were of pro
(50 m x 0.2 mm), coated with crosslinked methyl                   analyse quality. Lard was a kind gift from "Zem-
silicone gum (0.5 ).lm film thickness). Hewlett-                  jodelsko stopanstvo", Stip (Macedonia).
Packard, model 5890 Series II gas chromatograph
equipped with split-splitless injector was used. In-
strumental parameters were previously reported                                     Thermal autooxidation of lard
[15]. The relative amounts of individual compo-
nents were based on the peak areas obtained, with-                      Standard substances of BHA, BHT and thy-
out FID response factor correction. The GC-MS                     mol dissolved in ethyl acetate were added to the
                                                                  lard in amount of 0.02 % in final concentration.
analysis was carried out on a HP 5890 Series II gas
                                                                  Each of the essential oils were added to the lard in
chromatograph equipped with HP 5971 mass detec-
                                                                  the final concentration of 0.04 %. The blank sam-
tor working in electron impact mode (70 eV). The
chromatographic conditions were the same as in GC-                ple (control) contained ethyl acetate in concentra-
FID analysis. Transfer line was heated at 280 QC.
                                                                  tion of 0.04 %. Antioxidativ effects of oils on
The identification of the components was based on a               thermal autooxidation of lard was analyzed by the
comparison of their retention times with those of                 method described by Mimica-Dukic et al. [16]. All
analytical standards of available terpenoids, and                 samples of lard were stored in a thermostat cham-
matching mass spectral data of oil constituents                   ber at 60 QC. At definite time intervals samples
with those from Wiley/NBS library of MS spectra.                  were removed and the peroxide value (Pb) was
                                                                  determined by the Wheeler method [17]. The
      According to the previously published results
                                                                  experiment was carried out in triplicate.
[15], the main components of fifteen oils included



                                                                                    Bull. Chem. Technol. Macedonia, 20, 1,61-66 (2001)
                                       Antioxidant activity of essential oils of different wild Thymus L. species                            63

 Table        1
                     The content      of five components: thymol, carvacrol, linalool, geraniol                      and citral
                                    in the essential oils of the examined wild Thymus species

         Sample of oils                                           Thymol          Carvacrol           Citral        Linalool      Geraniol
                                                                    %                %                 %               %             %
         T. tosevii ssp. substriatus (Kavadarci)                   24.51             16.35             0.53           7.43          0.52
         T. tosevii var. degenii (Kitka)                           20.61             15.63            0.86            1.35          7.25
         T. tosevii var. longifrons (Kitka)                         17.25         ' 17.35             0.86            2.23         7.80


         T. tosevii var. degenii (Lazaropole)                      42.28             6.60             0.78           4.10          0.42
         T. longidens var. lanicaulis (Sonje)                      43.58             3.03             0.71           0.95          14.29

         T. tosevii var. tosevii (Kicevo)                           4.27            32.75             0.05           11.16         4.30
         T. tosevii var. longifrons (Mavrovo)                       8.64            21.06             0.27           0.44          0.11


         T. jankae var. patentipilus (Pelister)                     0.62                              25.10          31.16          1.25
         T. longidens var. dassareticus (Karadzica)                 3.36             0.45             10.87          24.27         16.27
         T.jankae var. pantotrichus (Pelister)                      0.48             0.29             16.96          30.35         2.30

         T. moesiacus (Bistra)                                      7.88             13.31            0.45           25.02         14.87
         T. alsharellsis (Alsar)                                    1.81             14.91            0.87           27.61         0.84

       T. tosevii var. longifrolls (Ponikva)                        0.30            10.00                            25.19         37.75
       T. moesiacus (Popova Sapka)                                  3.03            12.13             3.83           8.14          32.06
       T. longidens var. lanicaulis (Banjani)                       0.61             9.57             1.73           20.93         41.38




                                                      RESULTS AND DISCUSSION


      The effects of wild thyme essential oils, thy-                            "citral + linalool", "linalool + carvacrol" as well as
 mol and synthetic antioxidants, BRA and BRT, on                                oils from "geraniol + linalool + carvacrol" group
 thermal oxidation of lard are presented in Table 2.                            showed lower activity but in all measurements
The kinetic of oxidative changes in lard is pre-                               peroxide values were lower than that found in con-
 sented at Fig. 1. Essential oils with similar chemi-                          trol sample.
cal composition have shown very similar antioxi-                                      The results obtained confirmed antioxidant
dant effects. Remarkable differences were found in
                                                                               potential of phenolic components, thymol and car-
the activity of oils contained phenolic constituents                           vacrol. In one nonbiological model-system con-
in comparison to those containuing only traces of                              sisted of lard, lipid peroxidation was initiated by
thymol and carvacrol. As can be seen (Fig. 1), spe-                            higher temperature (60 QC),and the start of free
cies of wild thyme that contained high amounts of                              radicals reaction and production of peroxides was
phenolics and particularly carvacrol ("carvacrol"                              stimulated. The degree of oxidative changes of lip-
oils), provided very strong inhibition of lipid oxi-                           ids could be observed by measuring the value of
dation and kept the value of peroxides constant and                            created peroxides. Previously reported data showed
very low for 14 days. The effects were almost the                              that thymol and carvacrol produced stable free
same as those of BRA and much better than that of
                                                                               radicals, characterized by their EPR spectra [9, 10].
pure thymol. "Thymol" and "thymol + carvacrol"                                 These components can protect polyunsaturated
oils provided this inhibition for 7 days, very simi-                           fatty acids from oxidative changes, but they did not
lar to pure thymol. Other investigated oils from                               react as radical scavenging agents [12].


fJIac. XCM.TeXHOJI.MaKei10HHja, 20,1,61-66        (2001)
64                                                           S. Kulevanova, T. Kadifkova Panovska


Table         2
                      Peroxide   values in samples             of lard (at 60°C),          during the period                 of sixteen days

                                                                                           Peroxide       value (mmol O2 kg-i)
                                                                       1          2          4             7      9      11                        14     16
     Sample                                           Days ---7
     Control                                                          0        1.99        7.84          15.34             29.75       45.76     55.79   65.67
     0.02% BRA                                                        0        1.69         1.72         1.73              1.97        1.97       1.99   1.99
     0.02% BRT                                                        0        1.71         1.71         1.72              1.95        1.96       1.97   1.98

     0.02% thymol                                                     0        1.69         2.94         3.24              9.11        18.29     23.16   37.44

     "Thymol+carvacrol" oils
     T. tosevii ssp. substriatus (Kavadarci)                           0       1.73         2.94         3.48              10.56       17.80     22.10   35.04

     T. tosevii var. longifrons (Kitka)                                0       1.96         2.48         2.69              3.89        4.91      10.37   20.48

     T. longidens var. lanicaulis (Sonje)                              0       1.48         1.95         1.97              6.99        11.63     23.38   30.88

     "Thymol" oils
     T. tosevii var. degenii (Kitka)                                   0       1.73         2.69         2.96              8.08        13.37     17.86   29.40

     T. tosevii var. degenii (Lazaropole)                              0       1.49         2.48         2.49              9.26        16.19     24.17   35.46
     "Carvacrol" oils
     T. tosevii var. tosevii (Kicevo)                                  0       1.49         2.24         2.24              2.24        2.24       2.47   7.50

     T. tosevii var. longifrons (Mavrovo)                              0       1.73         2.40         2.72              3.99        5.38      11.29   22.44
     "Citral+linalool" oils

     T.jankae var. patentipilus (Pelister)                             0       1.95         7.44         14.49             26.92       39.45     44.29   64.36

     T. longidens var. dassareticus (Karadzica)                        0       1.98         3.97         13.99             27.75       41.66     50.96   58.90

     T. jankae var. pantotrichus (Pelister)                            0          1.94      7.97         15.33             20.24       26.04     39.28   63.54
     "Linalool+carvacrol" oils
     T. moesiacus (Bistra)                                             0          1.74      3.20         4.97              15.95       27.05     31.66   41.44

     T. alsharensis (Alsar)                                            0          1.99      2.96         7.88              18.17       28.83     39.10   51.18
     "Geraniol + linalool + carvacrol" oils
     T. tosevii var. longifrons (Ponikva)                              0          1.96      4.45         10.36             23.81       36.95     48.42   64.50

     T. moesiacus (Popova Sapka)                                       0          1.98      4.40         6.49               18.6       30.95     37.21   50.26

     T. longidens var. lanicaulis (Banjani)                            0          1.92      7.44         10.77             24.99       38.47     48.62   64.88


                                             70
                                                                                                                      I=-=c;;"troi--
                                             60   +
                                                                                                         /' /I    i   I--SHA
                                       -Cl 50                                                                     i I-*-     Thymol
                                        '"
                                        0
                                         -40.
                                        '0                                            p                      i
                                                                                                         /'11--       1-:%--'.   type
                                                                                                                             Thymol



                                        !20  30 1                          /          /7
                                                                                           .                     ':"
                                                                                                                      l-o-caNaerOltype
                                                                                                                             ""''''''"
                                             101

                                              OD
                                                  1
                                                        or
                                                        2
                                                                T
                                                                4
                                                                    ./'---" /' Y
                                                                          T
                                                                          7
                                                                               T
                                                                               9
                                                                                           -
                                                                                           11
                                                                                                   --;
                                                                                                     -
                                                                                                    14
                                                                                                                 ,l"-'"'.'"
                                                                                                                  T
                                                                                                                 16
                                                                                                                       -tr- erol type
                                                                                                                       -.----- Geraniol+linalool+eaN
                                                                                                                                               al

                                                                           Days



                  Fig. 1. Oxidative changes in lard with addition of essential oils, thymol and BRA, at 60 QCduring 16 days



                                                                                                          Bull. Chem. Technol. Macedonia, 20, 1,61-66 (2001)
                                         Antioxidant   activity of essential oils of different wild Thymus L. species                                   65

                                                                    CONCLUSION


     Essential oils of Macedonian wild Thymus                                            very interesting natural resources with antioxidant
species showed antioxidant activity in strong de-                                        activity such as: T. tosevii var. tosevii (Kicevo), T.
pendence to the carvacrol and less to the thymol                                         tosevii var. degenii (Kitka), T. tosevii var. longi-
content. Carvacrol containing oils showed stronger                                      frons (Kitka) T. tosevii ssp. substriatus (Kavadarci)
inhibition on thermal autooxidation of lard, at 60                                       and T. longidens var. lanicaulis (Sonje).
QC.Few Thymus species could be proposed as a


                                                                     REFERENCES


[1] C. O. Van den Broucke, Fitoterapia, 54,171-174 (1983).                              [11] S. G. Deans, R. C. Noble, L Penzes, S. G. Imre, AGE,
[2] C. O. Van Den Broucke, J. A. Lemli, Pharmacelllisch                                      16,71-74 (1993).
     Weekblad Scientific Edition, 5, 9-14 (1983).                                       [12] H. L. Madsen, B. R. Nielsen, G. Bertelsen, L. H. Skib-
[3] G. Vampa, A. Albasini, A. Provvisionato, A. Bianchi, M.                                  sted, Food Chemistry, 57, 331-337 (1996).
     Melegari, Plai1l.Med. Phytother., 22, 195-222 (1988).                              [13] T. Kadifkova Panovska, Gj. Stefkov, S. Kulevanova,
 [4] B. J. Juven, J. Kanner, F. Schved, H. Weisslowicz, J.                                  Proceedings of the First Conference on Medicinal and
     Appl. Bacteriol., 76, 626-631 (1994).                                                  Aromatic Plants of Southeast European COlli1lries,
 [5] O. Kandil, N. M. Radwan, A. B. Hassan, A. M. Amer, H.                                   Arangelovac, FR Yugoslavia, 2000, p. 527-531.
     A. EJbanna, W. M. Amer, J. Ethnopharmacol., 44, 19-24                              [14] Pharmacopea Helvetica, editio septima, Eidgenossische
     ( 1994).                                                                               Drucksachen- und Materia]zentrale, 3000 Bcrn (1987),
                                                                                            V.4.5.8 Gehaltsbestimmung des Atherischen DIes in Dro-
 [6] N. Nakatani, ACS Symposium Series, 507, 72-86 (1992).
                                                                                            gen.
 [7] K. Miura, N. Nakatani, Agricul. BioI. Chem., 53, 3043-                             [15] C. KYJIeBaHOBa, npOY'IYBalbe Ha eiTieplI'lHOlUO ",WCAO II
     3045 (1989).                                                                                              BO
                                                                                             cjJAaBOHolU:)U/Ue iipelUCluaBli1l4U 00 pOOOlU Thymus L. BO
 [8] C. Banias, V. Oreopoulou, C. D. Thomopoulos, J. Am.                                     cjJAopmua lia Peay6mlKa Mak.'eooJluja, JJ:OKTopCKa)\l!cep-ra-
     Oil. Chem. Soc., 69, 520-524 (1992).                                                    l\l!ja, <I>apMaueBTCKl!cpaKYJITeT, CKonje (1997).
                                                                                        [16] N. M. Mimica-Dukic, M. M. Budincevic, B. A. Mihaj-
 [9] N. Deighton, S. M. Glidewell, S. G. Deans, B. A. Good-
                                                                                             lovic,O. S. Gasic,J. Serb. Chem.Soc., 42, 823-828 (1994).
     man, J. Sci. Food. Agric., 63, 221-225 (1993).
                                                                                        [17] J. Trajkovic, M. Miric, J. Baras, S. Shiler, Analiza zivot-
[10] N. Deighton, S. M. Glidewell, B. A. Goodman, S. G.
                                                                                             nih namimica, Tehnolosko-metalurski fakultet, Bcograd,
    Deans, Biolog. Sci. 102,247-252 (1994).                                                  1983, pp. 410.




                                                                          Pe3HMe

     AHTMOKCIIJJ:AIJ;I10HA                   AKTIIBHOCT HA ETEPII1.IHII MACJIA OJJ:PA3JIIIqHII                                             BIIJJ:OBII
                                                     MAJqlIHA JJ:YIDIIQA

                                         CBeTJ13l1a KYJIeBalloBal*,          TaTjalla    Ka/l,mliKOBa     llallOBCKa    2

            1HHCiIilliTiyLTi 3a papMaK021l0311ja,       2HHCiIilltUyiIl      3a tUOKCUKO.flOUlKa xeMlIja, c[JapMGl!eaiIicKlI       paKY.fliIieiIi,
                                       BoofballcKa       17, MK-1000           CKollje, Pelly6.flltKa MGI:eooflua




          KJIY1JHH 36opOBH:    Thymus;    Maj'II:IHa ):IyuII:IlI,a; CTCpl1'IHO MaCJIO; aHTHOKCH):Iall,HOHa aKTHBHOCT;
                              nepOKCH):ICH 6poj



         11cnHTYBaHa c aHTHOKCH):Iall,HOHa aKTHBHOCT Ha neT-                            'ICHH KaKO MaCJIa 60raTH        co: THMOJI, THMOJI + KapBaKpOJI,
HaeceT    CTCPHqHH    MaCJIa 0):1 BH):IOBHTe Thymus            (Maj'II:IHa              KapBaKpOJI, JIHHaJIOJI + lI,HTpaJI, JIHHaJIOJI + KapBaKpOJI H
):IYlllHl\a) KOH ):IHBO pacTaT BO MaKe):lOHHja. XCMHCKHOT                               JIHHaJIOJI + repaHHOJI + KapBaKpOJI. AHTHOKCH):Iall,HOHHOT
COCTaB Ha MaCJIaTa ):I06HeHH co ):ICCTHJIall,Hja co BO):lHa na-                         CcpCKT Ha OBHe MaCJIa BP3 TCpMaJIHaTa OKCH):Iall,Hja Ha
pea 6ClllC onpC):ICJICH co MCTO):lHTC GC H GC-MS.               IIpHMc-                 CBHHCKaTa MaCT, TcpMOCTaTHpaHa Ha 60 "C, 6ClllC CJIC):IeH
pOll,HTe 0):1 MaCJIaTa 6ea    nO):leJICHH BO nCT rpynH          H 03Ha-                 npeKY   McpClbe     Ha CO):lp)[(HHaTa Ha C03):1a):lCHHTe ncpOK-




fJIac. xeM. TeXHOJI.MaKe)\OHl!ja, 20,1,61-66       (2001)
66                                                      S. Kulevanova,      T. Kadifkova Panovska



CII):\II BO MaCTa, BO IIepIIO):\OT 0):\ 16 ):\eHa. AHTlIoKclI):\a-                 substriatus   II T. longidens      var. lanicaulis,    HO 0):\ IIplIMepOll,ll
lI,lIOHIITe e<jJeKTII Ha MaCTa 6ea     BO CTpora       3aBIICHOCT 0):\             Ha pacTeHlIjaTa       KOII 6ea      c06paHII      0):\ TWIHO OIIpe):\eJIeHII
Co):\p)KIIHaTa Ha <jJeHOJUlTe, IIO'IIITYBajKII      ro CJIe):\HllOT pe-            JIOKaJIllTeTII.   0):\ ):\PyrlI IIplIMepOll,lI    Ha Thymus-oT,       0):\ IICTll
):\OCJIe):\: KapBaKpOJICKO > TllMOJI + KapBaKpOJICKO > TII-                        IIJIII ):\pyrlI Bll):\OBII, KOII llMaa pa3JIII'IHO IIOTeKJIO 0):\ IIpeT-
MOJICKO MaCJIO. nplIMepOll,ll    0):\ eTeplI'IHll MaCJIa 60raTII                   XO):\HIITe, eTepwIHllTe          MaCJIa IIOKa)KYBaa IIPllCYCTBO Ha
co <jJeHOJIHII KOMIIOHeHTII 6ea       ):\06lleHII   0):\ T. tosevii,   T.          TparlI    0):\ KapBaKpOJI       lI/llJIII   TllMOJI II IIoce):\YBaa     HII30K
tosevii var. degenii, T. tosevii var. longifrons, T. tosevii subsp.                IIOTeHlI,lljaJI 3a aHTIIOKClliWlI,llOHaTa          aKTllBHOCT.




                                                                                                      Bull. Chem. Technol. Macedonia, 20, 1,61-66 (2001)

				
DOCUMENT INFO
Shared By:
Categories:
Tags:
Stats:
views:68
posted:3/10/2010
language:English
pages:6