International Journal of Agricultural

Science and Research (IJASR)

ISSN(P): 2250-0057; ISSN(E): 2321-0087
Vol. 4, Issue 6, Dec 2014, 139-146
TJPRC Pvt. Ltd

COMPARATIVE STUDY OF CHEMICAL COMPOSITION AND ANTIOXIDANT

ACTIVITY OF ETHANOLIC EXTRACTS FROM ALGERIAN LAVANDULA STOECHAS L.
AND ROSMARINUS TOURNEFORTII DE NO
FOUAD MENACEUR1 & MOHAMED HAZZIT2
1

Department of Food Technology, The School of Agricultural and Life Sciences, El-Harrach, Algeria
1,2

Mohand University Akli Ouelhadj, Faculty of Nature and Life and Earth Sciences,
Department Science and Nature of Life, Bouira, Algeria

ABSTRACT
The purpose of the study was the analysis of extracts from Rosmarinus tounefortii de No and Lavandula stoechas
L. and the assessment of their in vitro antioxidant. The ethanolic extracts of studied plants recorded yields of 19.3 and
30.6% for L stoechas and R. tournefortii, respectively. These extracts had high percentages of polyphenols and flavonoids.
Antioxidant activity was assessed in vitro using two different tests. Free radical scavenging activity (DPPH) test revealed
that ethanolic extracts of rosemary (IC50= 11.6 0.1 mg/l) and lavender (IC50= 18.3 0.3 mg/l) exhibited higher DPPH
scavenging activity compared to that of the synthetic antioxidant BHT (IC50= 28 0.7 mg/l), and studied essential oils.
For reducing power test, the strongest effect was exhibited by butylated hydrxytoluene (BHT).

KEYWORDS: Rosmarinus tournefortii, Lavandula stoechas, Ethanol Extracts, Antioxidant Activity

INTRODUCTION
In recent years, increasing attention has been paid to the exploration of naturally-occurring antioxidants because
of the growing consumer demand for food products free from synthetic chemical additives. The plant kingdom has
attracted special interest because of its remarkable diversity in producing natural compounds (Wang et al, 2010).
Lamiaceae is a relatively common botanical family, members of which are found in the temperate regions
worldwide (Cantino, 1992). It includes approximately 220 genera and about 3,500 to 4,000 species (Almeida and
Albuquerque, 2002).
Lavandula genus is an important member of Labiatae (Lamiaceae) family. Lavandula species are widely
distributed in the Mediterranean region, and have great commercial value due to their intense and pleasant aroma. Both the
plant material and its essential oil are mainly demanded in perfumery, cosmetic, and food industries (Giray et al, 2008).
Rosmarinus is a small Mediterranean genus of aromatic shrubs. Present taxonomic accounts are both
unsatisfactory and confused by the many different treatments which have been produced. This has resulted from the
tremendous variation found within some of the species and hybridisation in the genus. As many as 14 specific names and
over 50 subspecific combinations exist in the literature, which have been reduced to three species; Rosmarinus officinalis
L, R. tournefortii L. (syn. R. eriocalyx Jordan & Four) and R. tomentosus Hub. Mor & Maire.

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Fouad Menaceur & Mohamed Hazzit

Rosmarinus tournefortii occurs in the Province of Almeria in Spain and across part of North Africa, differing
principally from R. officinalis in having a hairy calyx, in its habit and in corolla characters (Upson, 1992).
One of the major concerns in food technology is lipid oxidation due to the formation of oxidation products such as
fatty acid hydroperoxides and secondary degradation products (alkanes aldehydes, alkenes). Antioxidants have been
reported to prevent oxidative damage caused by free radical; they can interfere with the oxidation process by reacting with
free radicals, chelating, catalytic metals and also by acting as oxygen scavengers (Buyukokuroglu et al, 2001).
The utilization of antioxidants such as gallates, butylated hydroxytoluene (BHT), tert-butyl hydroxyanisole
(BHA) and tert-butyl hydroquinone (TBHQ) can prevent food oxidation or cell damage. Howerver, the safety of using
these synthetic antioxidants has become a concern among scientists and leading to current interest in uncovering natural
antioxidants. Nevertheless, strict legislation on the use of synthetic food additives and consumer preferences has shifted the
attention of manufacturers from synthetic to natural antioxidants (Dapkevicius et al, 1998). Hence, strong restrictions have
been placed on their application and there is a trend to substitute them with naturally occurring antioxidants. Natural plantbased antioxidants especially phenolics and avonoids have been exploited commercially either as antioxidant additives or
as nutritional supplements (Schuler, 1990)
The aim of the present study was to analyze the chemical composition of ethanol extracts of R. tournefortii and
L. stoechas from Algeria and to evaluate their antioxidant ability.

MATERIALS AND METHODS

Plants
R. tournefortii was collected from Tablat (Longitude: 36o4127" Latitude: 3o 07 71", 60 Km south Algiers),
while L. stoechas was collected from Azzazga (Longitude: 364441". Latitude: 4722 20", 100 Km east Algiers).
The plants were harvested at the beginning of their flowering period.
Preparation of the Extracts
Dried leaves and flowers at room temperature in the dark were grounded into fine powder. 20 g of powder of each
plant were extracted with 100 ml of solvent (ethanol 95%) for 6 hours using Soxhlet extractor. The solvent was removed
under reduced pressure at 60 C using rotary evaporator. Finally, the residue was lyophilised, weighed and kept in the dark
at + 4-6C until further analysis.
Total Phenolic Content
Total phenolic constituents of plant extracts were performed employing the literature methods involving
Folin-Ciocalteu reagent and gallic acid as standard (Singleton et al, 1999). 0.25 ml of each sample (three replicates) was
mixed with 1.25 ml 1/10 dilution of Folin-Ciocalteaus reagent. After 3 minutes, 1 ml of Na 2CO3 (7.5%, w/v) were added
and incubated for 30 min. The same procedure was repeated to all standard gallic acid solutions (0.02-0.12 mg. ml-1) and
standard curve was obtained.
The absorbance of all samples was measured at 765 nm using a UV-VIS UNICAM Helios spectrophotometer
against a blank in which we replace extract solution with ethanol. The total phenolic content was expressed in mg of gallic
acid equivalents (mg GAE) / g of dried extract.

Impact Factor (JCC): 4.3594

Index Copernicus Value (ICV): 3.0

Comparative Study of Chemical Composition and Antioxidant Activity of Ethanolic

Extracts from Algerian Lavandula stoechas L. and Rosmarinus tournefortii De No

141

Total Flavonoids Contents

The total flavonoids content was determined using the modified AlCl3 method (Lamaison and Carnet, 1990). 1 ml
of aluminum trichloride (AlCl 3) in ethanol was mixed with the same volume of the extract solution. Absorption readings at
420 nm were taken after 1 hour against a blank sample consisting of a 1 ml of AlCl 3 with 1 ml ethanol without extract
solution. (The total flavonoids content was determined using a standard curve with quercetin (0.05-0.25 mg.ml-1) as the
standard. Total flavonoids content is expressed as mg of quercetin equivalents (mg QE/g of dried extract), by comparison
with the quercetin standard curve, which was made under the same conditions.

ANTIOXIDANT ACTIVITY

DPPH Assay
The antiradical activity of ethanolic extracts was determined using the stable 2, 2-diphenyl-1-picrylhydrazyl

radical (DPPH) (Hazzit et al, 2009). 25 L samples of various concentrations of ethanolic extracts were added to 975 L
of ethanolic solution containing DPPH radicals (60M) while butylated hydroxytoluene (BHT) acted as a positive control,
The absorbance of DPPH radical solution without sample was measured as blank. All test tubes were incubated in a dark
place at room temperature for 30 minutes. Then the absorbance was measured at 517 nm. All determinations were carried
out in triplicates. The disappearance of DPPH was recorded and the percent inhibition of the DPPH radical by sample is
calculated as follows:
Inhibition Percent = [(Ab As) / Ab] x 100.
Where Ab is the absorbance of blank and As is the absorbance of positive control or sample. Extract concentration
providing 50% inhibition (IC50) was calculated from the graph plotting inhibition percentage against extract concentration.

Reducing Power
The reducing power was determined according to the method of Oyaizu (1986). 0.125 ml of ethanolic extracts and

BHT at different concentrations were mixed with 2.5 ml of sodium phosphate buffer (0.2 M, pH=6.6), and 2.5 ml of
potassium ferricyanide (1%). The mixture was incubated at 50C for 20 min. Then, 2.5 ml of 10% trichloroacetic acid was
added to the mixture which was centrifuged for 10 min at 1500 x g. The upper layer (2.5 ml) was mixed with 2.5 ml of
distilled water and 0.5 ml of 0.1% ferric chloride. The absorbance was measured at 700 nm. Blank absorbance was read by
replacing sample by ethanol. BHT was used as a positive control. The reducing power increases with the increase of
absorbance. All determinations were carried out in triplicates.

RESULTS AND DISCUSSIONS

Yield, Total Phenolic Contents and Flavonoids Contents of Ethanolic Extracts
Dried ethanol extracts were weight out and the percent yields of ethanol extracts were calculated and expressed in
g of extract /100 g of dry plant material (Table 1).
R. tournefortii was found to have higher yield than that of L. stoechas (30.6 and 19.3%, respectively). Soxhlet extraction
provided good yield of ethanolic extract from L. sotechas compared to that reported by Giray et al. (2008) working on
Turkish lavender (9.4%). Total phenolic assay was carried out based on the absorbance values of the various extract
solutions, reacted with FolinCiocalteu reagent and compared with the standard solutions of Gallic acid as described above
(Table 1). The content of total phenolics in R. tournefortii (221 mg GAE/g of dry extract) was found to be much higher

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Fouad Menaceur & Mohamed Hazzit

than in L. stoechas (132.3 mg GAE/g of dry extract). The variation in the total phenolic amount was attributed to many
factors including geno type, agronomic practices, maturity level at harvest, post-harvest storage, climatic and geographic
allocations. Total flavonoids contents of the extracts expressed as mg equivalent of quercetine /g of dry extract show that
ethanolic extract from L. stoechas contained significantly higher levels of flavonoids compared to R. tournefortii (41.6 and
26 mg equivalent of quercetine /g of dry extract, respectively).
Antioxidant Activity

Inhibition of DPPH Radical

Reduction of the DPPH radical, which accepts an electron of hydrogen radical to become a stable diamagnetic

molecule, was determined by the decrease in its absorbance at 517 nm by antioxidants. Figure 1 shows the scavenging
effect of essential oils; ethanol extracts samples and the chemical antioxidant (BHT). All the essential oils and ethanol
extracts increased with concentration. The results showed low inhibition values for the essential oils in comparison with
BHT. Thus, at the highest concentration tested (1000 mg/l), the scavenging activities were (24.11 0.34, 19.69 1.29 and
91.07 0.13%) for rosemary, lavender and BHT, respectively. On the other hand, ethanolic extracts exhibited good
scavenging activity (92.2 0.6% for R. tournefortii, 89.2 0.5% for L. stoechas and (73.2 0.9 %) for synthetic antioxidant
BHT at the concentration of 100 mg/l). In the current study, IC50 value of essential oils could not be determined with used
concentrations; their scavenging activity did not exceed 25% at the highest concentration (1000 mg/l).
Our ethanol extracts samples displayed solid inhibition activity; they strongly transformed the DPPH radical into
its reduced form. They have IC50 values of 11.59 0.07 and 18.30 0.31 mg/l for R. tournefortii and L. stoechas
respectively, which is lower than that of the antioxidant BHT (28.010.66 mg/l); thereby indicating their strong antioxidant
activity. Mohammedi (2006), reported lower IC50 value for methanol extract of Algerian L. stoechas (5.29 0.27 mg/l).

Reducing Power
Figure 2 shows the reducing power (as indicated by the absorbance at 700 nm) of ethanol extracts and essential oil

from R. tournefortii and L. sotechas compared with BHT as standard. The obtained results indicated gradually increase of
reducing power with increasing of concentration of all samples.
The reducing power of all samples increased with the concentrations. For ethanol extracts, the sequence of
reducing power was as follow: BHT>Lavender ethanol extract> Rosemary ethanol extract, (corresponding absorbances at
700 nm were 0.8630.011, 0.7390.035 and 0.5910.012 at 100 g/ml, respectively).
Despite its lower content of total phenol compounds, lavender ethanol extract showed higher reducing power than
that of rosemary, this may could be attributed to its important amount of flavonoids which have been shown to be highly
effective scavengers of most types of oxidizing molecules, including singlet oxygen and various free radicals (Buettner,
1993; Bravo, 1998). In the present study, essential oil of L. stoechas expressed slightly greater antioxidant activity than that
of R. tournefortii. Nevertheless, both essential oils displayed reducing power significantly lower than that of BHT, this
could be explained by their very low levels of phenol compounds and flavonoids which are the main responsible of the
strong antioxidant activity of plants extracts.
In general, ethanol extracts exhibited significantly higher reducing power than that of essential oil from the same
plant and expressed an effect even with low concentrations.

Impact Factor (JCC): 4.3594

Index Copernicus Value (ICV): 3.0

143

Comparative Study of Chemical Composition and Antioxidant Activity of Ethanolic

Extracts from Algerian Lavandula stoechas L. and Rosmarinus tournefortii De No

CONCLUSIONS
We were interested in the extraction, characterization and evaluation of antioxidant and insecticide of plant
extracts from R. tournefortii and L. stoechas. The studied extracts showed very dissimilar chemical profiles. A potential
use of essential oils and ethanol extracts, in preventing alterations of food was explored, and this by replacing the
commonly used chemicals that may have adverse effects on human health and on the environment.
Ethanol extracts of studied plants showed high percentage of polyphenols and flavonoids; in other hand they
exhibited strong radical-scavenging activity and a very high reducing power compared to those of essential oils from the
same plants. In addition their important antioxidant activity has been revealed even with low concentrations. This strong
antioxidant activity of ethanol extracts suggests their possible use as natural antioxidants or in a potential pharmaceutical
application. In view of the present results, this is the first report on the ethanolic extracts antioxidant activity of Algerian
R. tournefortii. Organic control by using essential oils is an alternative to chemical control that poses no persistence of
action, resistance and residue problems. However, their use must be accompanied of an appropriate choice of applied
doses, because very low or too high doses can lead to the opposite effect, and for the same oil we can see two reactions.
Further research, mainly on the interactions with other food ingredients, is still necessary.

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Table 1: Yield (%), Phenolic Contents and Flavonoids Contents of Ethanol Extracts
Sample
Rosmarinus tournefortii
Lavandula stoechas

Yield (%)
30.6
19.29

Phenolic Contents
Flavonoids
mg GAE/g
Contents mg QE/g
221.4
26.0
132.3
41.6

The values were shown as (%) percent yield of ethanol extracts (w/w).

Figure 1: DPPH Radical-Scavenging Activity of Ethanol Extracts (Ext) from Rosmarinus tournefortii
(Rosemary) and Lavandula stoechas (Lavender) Compared to that of BHT (mean S. D; n =3)
Impact Factor (JCC): 4.3594

Index Copernicus Value (ICV): 3.0

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Comparative Study of Chemical Composition and Antioxidant Activity of Ethanolic

Extracts from Algerian Lavandula stoechas L. and Rosmarinus tournefortii De No

Figure 2: Reducing Power of Ethanol Extracts (Ext) from Rosmarinus tournefortii (Rosemary) and
Lavandula stoechas (Lavender)) Compared to that of BHT (Mean S. D; n = 3)

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