European Food Research and Technology (2020) 246:167–177

https://doi.org/10.1007/s00217-019-03396-9

ORIGINAL PAPER

Effect of light intensity and water availability on plant growth,

essential oil production and composition in Rosmarinus officinalis L.
Antonio Raffo1   · Eric Mozzanini2 · Stefano Ferrari Nicoli1 · Elisabetta Lupotto1 · Claudio Cervelli2

Received: 15 July 2019 / Revised: 17 September 2019 / Accepted: 26 October 2019 / Published online: 16 November 2019
© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Abstract
The effect of light intensity (LI) and water availability (WA) on rosemary (Rosmarinus officinalis L.) plant growth, essen-
tial oil (EO) production and composition was investigated by a two-factorial field experiment, where the first factor was LI
(100%, 50% or 25% of natural sunlight) and the second factor was WA (irrigation set at 85%, 70% or 55% of field capacity
during plant growing). The EO obtained by steam distillation of the dried aerial part of the plant was analysed by GC/MS.
Reduction of LI from 100 to 25% of natural sunlight markedly lowered plant biomass production, whereas reduction of WA
from 85 to 55% had a smaller lowering effect on plant growth. High shading (25% of LI) markedly reduced EO yield on a
plant basis (− 43%), whereas intermediate shading (50% of LI) increased EO yield as % content of the fresh biomass (+ 29%)
when compared to full solar radiation. WA markedly influenced EO yield, as expressed on a plant basis, but only in plants
exposed to 100% LI. Moreover, changes in LI and WA seemed to have an opposite effect on the relative abundance of EO
constituents that are formed through the activity of two groups of enzymes, pinene synthases (α- and β-pinene, camphene
and myrcene) and, respectively, bornyl diphosphate synthases (borneol, camphor and bornyl acetate). Accurate management
of light conditions and water availability, in greenhouse as well as open field conditions, may allow to optimize rosemary
EO yield and modulate EO profile in view of different potential uses.

Keywords  Rosemary · Terpenoids · Monoterpene synthases · Solar radiation · Aroma · Irrigation

Introduction applications as ingredient of functional foods [4] or active

Rosemary is a perennial aromatic herb and member of the
Lamiaceae family, native and widely cultivated in the Medi-
terranean basin, where it is traditionally used as a spice herb
for culinary purposes [1]. Rosemary essential oil (EO) is
reported to be commercially used in quantities between 100
and 500 tons per year [2], namely as fragrance in cosmetic
products [3], flavouring and active ingredient in products for
aromatherapy and food preservation [1]. Innovative potential
Electronic supplementary material  The online version of this
article (https​://doi.org/10.1007/s0021​7-019-03396​-9) contains
supplementary material, which is available to authorized users.
* Antonio Raffo
antonio.raffo@crea.gov.it
1
CREA-Research Centre for Food and Nutrition, Via
Ardeatina, 546, 00178 Rome, Italy
2
CREA-Research Centre for Vegetable and Ornamental
Crops, Corso Inglesi, 508, 18038 Sanremo, Italy
packaging for food products [5] are focus of current intense
research.
It is well established that, not only plant growth and mor-
phology, but also EO production are strongly influenced in
medicinal and aromatic plant (MAP) species by climatic
conditions [6–12]. Indeed, even though the composition
of EO is primarily under genetic control [13], its forma-
tion is also highly dependent on environmental conditions,
such as day length, irradiance, temperature and water sup-
ply [14]. Light intensity (LI) is, in general, strongly asso-
ciated with plant photosynthetic activity and, thus, carbon
fixation, vegetative growth and dry matter accumulation in
several species, while secondary metabolites, including EO
constituents, are formed from photosynthetic carbon [15].
However, not all experimental studies on the effect of LI on
EO production in MAP species showed univocal results: in
some cases, a higher LI was associated to a higher EO yield,
as observed on Thymus vulgaris [16], Origanum vulgare
[17], Ocimum basilicum [18] and Mentha piperita [19]. On
the contrary, in other experiments on Salvia officinalis [16],

13
Vol.:(0123456789)

168
Mentha piperita [20], Eucalyptus citriodora [21], plants sub-
jected to a reduced level of solar irradiance produced higher
yields of EO, suggesting that effects on EO production may
not simply parallel those on photosynthetic activity. Also
water availability (WA) has been observed to markedly influ-
ence formation of EO constituents in several species [22].
Water shortage generally leads to an increase of EO concen-
tration in the plant organs, even though due to the marked
concurrent decline of biomass production, total EO content
on a whole plant basis generally decreases, as observed in
Salvia officinalis [22, 23], Lavandula angustifolia [23],
and Mentha spicata [24]. As regards rosemary, while it has
been thoroughly assessed that EO composition is mainly
dependent on genetic background and origin [25], it has also
been observed that severe water stress conditions signifi-
cantly enhance EO concentration within plant biomass, even
though due to the marked biomass reduction slight changes
in EO yield result on a plant basis [24]. On the contrary, no
information is available about the effect of more moderate
water stress conditions, which would be compatible with
crop cultivation management, nor about the influence of LI.
In addition no information have been reported on rosemary
about the interaction between the two factors, LI and WA,
while it has been emphasized that interference of multiple
factors may be relevant when analysing the influence of
environmental conditions on EO production [22].
Cultivation of rosemary generally takes place in the open
field, where solar irradiance may significantly differ depend-
ing on latitude, field slope and orientation, or geographical
features, such as the shading of tall hedge. Thus an improved
knowledge of the effects of light conditions on EO produc-
tion could have useful implications on management of prod-
uct quality and standardization, both in open field and in
greenhouse conditions [17]. Moreover, WA may be delib-
erately modulated in the field to optimize cultivation strate-
gies aimed at improving product quality and yield [22]. In
fact, the importance of both light and drought-related stress
conditions to be used in elicitation methods to enhance the
synthesis of secondary metabolites in MAP species is gener-
ally recognized [26].
In the present study the effects of light intensity (LI)
and water availability (WA), along with their interactions,
on rosemary (Rosmarinus officinalis L.) plant growth and
EO production and composition were investigated through
a two-factorial field experiment, where the first factor was
LI (100%, 50% and 25% of natural sunlight) and the sec-
ond factor was WA (by setting irrigation regime at 85%,
70% and 55% of field capacity during plant pot growing).
Aim of the study was to provide an in-depth knowledge of
rosemary plant growth response to these key environmental
factors along with their influence on EO yield and composi-
tion, with the purpose of optimizing cultivation strategies
to improve crop productivity and EO quantity and quality.
13
European Food Research and Technology (2020) 246:167–177
Materials and methods
Plant materials and growing conditions
The trial was performed at Sanremo (latitude 43°49′05.23″N,
longitude 7°45′26.12″E), Italy, in the experimental fields
of CREA-Research Centre for Vegetable and Ornamental
Crops, located about 80 m a.s.l., in a climatic area suita-
ble for citrus cultivation in the open air. A rosemary (Ros-
marinus officinalis L.) variety widely cultivated in Italian
aromatic pot plant production, commonly named “erect
rosemary” and characterized by an α-pinene/1,8-cineole
chemotype, was used. Plants propagated by a local grower
were potted in 7.5-L plastic pots on 3 April 2017 in a com-
mercial substrate (Terflor ­Hochmoor® Vulcan Invernale
pumex) suitable for nursery crops. A controlled release
fertilizer (Geogreen Horti-Cote®Plus, 6  months release
period, N–P–K–Mg 16–6–11–2 plus microelements) was
added in the amount of 5 g L−1 of substrate. Two fertiga-
tions were accomplished after 7 and 12 days with Ferty
1 (Planta Düngemittel GmbH, N–P–K–Mg 20–7–10–2,
1.5 g L−1). Application of different levels of LI and WA
started on 15 April. Three light environments in the open air
were arranged, where reduction of full sunlight was obtained
by shading the plants with black plastic nets of suitable cut-
off. The three LI treatments corresponded to (a) 100%, or
full, sunlight; (b) 50% sunlight and (c) 25% sunlight. The
plants subjected to each light treatment were grown under an
arched metallic structure 2.3 m tall; each structure was com-
pletely covered (except for 100% sunlight) by the appropriate
shading net and, only on the top, by a polythene transparent
film to avoid rainfall into the pot substrate (all treatments).
In each light environment the plants were subjected to three
different WA regimes: (a) 85%, (b) 70%, (c) 55% of field
capacity. To accomplish these regimes during plant growth,
water was not supplied until the scheduled field capacity per-
centage was reached, restoring the 100% field capacity level
by irrigation after the threshold was overcome. The water
amount in each pot and the corresponding percentage of field
capacity was calculated daily by weighing the pots, then sub-
tracting the weight of the pot (fixed), of the dried substrate
(a representative sample was measured before experiment)
and of the plant (on sample plants a destructive relief was
carried out monthly). In that way a two-factorial scheme was
obtained, with 9 treatments (3 LI levels × 3 WA levels), and
a completely randomized experimental design was arranged;
15 plants for each treatment were used. Measurements on
biomass growth and plant structures were performed. Data
on temperature of leaves and pot external surface were also
collected by an infrared thermometer (Minolta Cyclops
Compac 3, Osaka). The trial lasted 154 days (15 April–15
September 2017).
European Food Research and Technology (2020) 246:167–177 169
Essential oil extraction
For each treatment, the aerial part of 3 plants (3 replicates),
without the woody base (about 7 cm tall) carrying the branch
insertions, was harvested, reduced to small pieces and air
dried at room temperature (20–28 °C) in a shaded environ-
ment for 15 days after the trial end. Weight loss was deter-
mined for each plant replicate. The resulting biomass from
each plant replicate was accurately mixed and transferred
into a 2-L round-bottomed flask, 1 L of water was added and
the mixture was steam distilled for 2 h using a Clevenger-
type apparatus, according to the method recommended by
the Italian Pharmacopoeia [27]. At the end of distillation
the volume and weight of the obtained EO was determined.
GC–MS analysis of essential oil profile
Extracted EO samples were dried over ­Na2SO4, then 1 mL
of oil was diluted to total 100 mL with tert-butyl methyl
ether, containing butyl benzene as internal standard
(2 μL mL−1). GC/MS analyses were performed on an Agi-
lent 6890 GC 5973 N MS system equipped with a quad-
rupole mass filter for mass spectrometric detection (Agi-
lent Technologies, Palo Alto, CA) and a DB1-MS column
(0.25 mm i.d. × 60 m, 0.5 μm film thickness; J&W, Agilent
Technologies, Palo Alto, CA) for GC separation. Chro-
matographic conditions were as follows: split (50:1 ratio)
injection, injector temperature at 250 °C, oven temperature
programme from 70 to 200 °C at 4 °C min−1, and then to
270 °C (5 min) at 50 °C min−1, constant flow of He carrier
gas was 1.5 mL min−1 corresponding to a linear velocity
of 32 cm s−1. The MS detector operated in the electronic
impact ionization mode at 70 eV; transfer line, source, and
quadrupole temperatures were set, respectively, at 300, 230,
and 150 °C. Detection was performed in the full scan mode,
over the mass range 30–250 amu. Identification of EO con-
stituents was accomplished by comparison of linear retention
indexes (LRI) and mass spectra of chromatographic peaks
with those obtained on standards solution of pure reference
compounds (purchased from Sigma-Aldrich, Milan, Italy).
Linear retention indexes were determined by analysing, in
the same conditions used for EO samples, a standard solu-
tion of C7–C30 saturated alkanes, and by applying the equa-
tion proposed by van den Dool and Kratz. When a pure com-
pound was not available tentative identification was based
on the comparison of determined linear retention indexes
with those reported in the NIST Chemistry WebBook data-
base [28], and on the comparison of mass spectra with those
reported in the NIST/EPA/NIH Mass Spectra Library 2005.
To evaluate the relative abundance of each EO constituent
(expressed as percentage content) peak area normaliza-
tion measurement was carried out, based on average data
obtained by a duplicate GC run on the same EO diluted
sample.
Statistical analysis
A 2-way ANOVA was performed on plant growth data and
on EO (yield and composition) data to highlight significant
effects associated to each of the experimental factors and to
their interactions. A 1-way ANOVA was conducted on plants
temperature data. When significant effects were observed,
the Tukey’s multiple comparison test was performed to high-
light significant differences between the levels of the two
factors. ANOVA and Tukey’s test were performed by using
the XLStat software (ver. 2016.05; Addinsoft, New York,
NY).
Dataset on EO composition was formed by average (n = 2)
data on 28 constituents determined on the 27 EO samples (9
treatments × 3 plant replicates): percentage data on relative
content of EO constituents were transformed by the arcsin
function before statistical analysis. A principal components
analysis (PCA) was carried out on EO constituents data by
using the PAST software [29].
Results and discussion
Effects on plant growth
Both whole plant fresh weight and dry weight were nega-
tively affected by LI reduction (Table 1). In plants exposed
to a 25% LI level, a 55% and a 59% decrease of whole plant
fresh and dry weight was observed with respect to 100% LI.
A similar effect of LI reduction was found on fresh and dry
weight of the plant aerial part (49% and 57% decrease) and
an even more strong effect on roots fresh and dry weight
(77% and 72% decrease). Reduction of LI also decreased
stem diameter, the number of total shoots, even though
plants exposed to the highest LI showed a decrease of the
nodes developed on the trunk during the last 2 months of
cultivation. As regards the effects on leaf characteristics,
reduction of LI induced an increase of the leaf area and a
decrease of specific leaf weight, both fresh and dry.
Differently from the effect of LI, only plant fresh weight,
but not dry weight, was affected by WA, with a significant
reduction at the lowest WA level (55% of field capacity)
(Table 1). This effect was much smaller than that due to
reduction of LI, amounted approximately to a 11% plant
fresh weight decrease and could be ascribed to a reduced
plant water content. The effect of WA reduction on the aerial
part fresh weight (17% decrease) was similar to that one
observed on the whole plant. On the contrary, reduction of
WA promoted roots biomass accumulation, the lowest val-
ues being recorded at the highest WA level (85% of field
13

170 European Food Research and Technology (2020) 246:167–177

Table 1  Data on rosemary plant growth

Variable 2-way ANOVA (p values)1 Light intensity (LI) levels Water availability
(WA) levels
Light intensity (LI) Water Interaction LI × WA 100% 50% 25% 85% 70% 55%
availability
(WA)

Plant growth and architecture

 Whole plant fresh weight (g) 0.000*** 0.002** 0.198 475 ­a2 398 b 216 c 377 a 376 a 334 b
 Whole plant dry weight (g) 0.000*** 0.404 0.062 157 a 126 b 64 c 116 118 113
 Aerial part fresh weight (g) 0.000*** 0.000*** 0.002** 375 a 337 b 193 c 325 a 310 a 270 b
 Aerial part dry weight (g) 0.000*** 0.067 0.014* 137 a 112 b 59 c 105 104 99
 Roots fresh weight (g) 0.000*** 0.039* 0.169 100 a 60 b 23 c 52 b 66 a 64 ab
 Roots dry weight (g) 0.000*** 0.014* 0.137 19.6 a 13.7 b 5.4 c 11.0 b 13.7 a 14.0 a
 Aerial part/roots fresh weight ratio 0.000*** 0.000*** 0.066 3.75 c 5.61 b 8.39 a 6.25 a 4.70 b 4.22 b
 Aerial part/roots dry weight ratio 0.000*** 0.000*** 0.168 7.01 b 8.20 b 10.85 a 9.55 a 7.59 b 7.07 b
 Stem diameter (mm) 0.000*** 0.000*** 0.000*** 16.3 a 12.2 b 10.3 c 14.3 a 12.7 b 11.9 b
 Total shoots (number) 0.000*** 0.000*** 0.035* 474 a 478 a 447 b 485 a 471 a 442 b
 Nodes developed on the trunk in 0.000*** 0.096 0.095 1.25 b 9.92 a 6.83 a 4.58 5.50 7.92
the last 2 months (number)
 Leaf area (­ cm 2) 0.000*** 0.047* 0.075 0.46 c 0.59 b 0.67 a 0.59 a 0.59 a 0.54 a
 Leaf specific fresh weight 0.046* 0.631 0.195 0.069 a 0.059 b 0.061 b 0.061 0.064 0.063
(g cm−2)
 Leaf specific dry weight (g cm−2) 0.000*** 0.387 0.012* 0.027 a 0.019 b 0.016 c 0.020 0.022 0.021

Results of 2-way ANOVA and mean values of samples belonging to the same factor (light intensity and water availability) level. Tukey’s multi-
ple comparison test (at p = 0.05)
1
 Statistical significance of ANOVA: *p < 0.05; **p < 0.01; ***p < 0.001
2
 When a factor was significant at ANOVA the Tukey’s multiple comparison test was performed to compare data related to different levels of
each factor. In the box of light intensity or water availability, different letters within the same row denoted significant differences between differ-
ent levels of the factor, according to Tukey’s test (at p < 0.05)

capacity). Consequently, while reduction of LI increased the
aerial part/roots weight ratios (fresh and dry weight ratios),
the opposite effect was associated to WA reduction. Reduc-
tion of WA, similarly to LI, was associated to lowered stem
diameter and number of total shoots, whereas leaf character-
istics seemed to be scarcely affected by WA changes.
ANOVA results also showed the occurrence of significant
interactions between the two factors for some plant growth
parameters (Table 1), in particular for the aerial part weight.
Evaluation of data related to individual treatments (Fig. 1)
highlighted that the effect of WA reduction on formation of
the aerial part biomass occurred only in plants subjected to
100% LI, whereas the influence of WA was negligible in
plants exposed to lower LI levels. This observation was even
more clear when evaluating the plant foliage biomass weight
(Fig. 1S), instead of the global weight of the aerial part. In
a similar way, the effect of WA reduction on formation of
roots biomass occurred only in plants exposed to 100% LI,
whereas no effect was observed at lower LI levels (Fig. 2).
As reported in the literature for many species, a lower LI
generally reduces plant growth (in terms of biomass pro-
duction), extent of branching, stem diameter, leaf thickness
and specific leaf weight, and these effects may be linked
to reduced photosynthetic activity, and resulting lowered
carbon fixation [6–8, 10, 11]. In addition, a reduction of LI
is associated to an increase of the aerial part/roots weight
ratio, mainly due to a remarkable reduction of the root bio-
mass, and of leaf size. This adaptation can be interpreted as a
strategy to maximize the capture rate of radiant energy [30].
Similar changes in plant growth have also been found in
MAP species, such as Salvia sclarea [31], Salvia officinalis
[32] and Ocimum basilicum [18]. Results reported in this
paper show that rosemary plant responds in a similar way to
the other species reported above when subjected to increas-
ing levels of shading. Moreover, reduction of biomass of the
aerial part at the 25% LI level was higher than that observed
on Salvia officinalis, another aromatic species typically cul-
tivated in the open air [32], confirming the sun-loving nature
of rosemary plant, which, accordingly, in a wild environment
tends to behave like a pioneer species.
In general, plant growth is adversely affected by water
stress due to a decrease in stomatal opening, which restricts
­CO2 diffusion into leaves and, thus, reduces photosynthetic
activity [12]. Minor effects on plant growth have been

13
European Food Research and Technology (2020) 246:167–177 171

(a) Light intensity (LI) level (a) Light intensity (LI) level
100% LI 50% LI 25% LI 100% LI 50% LI 25% LI
450
115
400
Aerial part fresh weight (g)

Roots fresh weight (g)

95
350
75
300

250 55

200 35

150 15
85 70 55 85 70 55
Water availability (WA) level Water availability (WA) level

Light intensity (LI) level

(b) Light intensity (LI) level
(b) 100% LI 50% LI 25% LI
100% LI 50% LI 25% LI
25
150
Aerial part dry weight (g)

Roots dry weight (g)

130 20

110 15

90 10

70 5

50
0
85 70 55
85 70 55
Water availability (WA) level Water availability (WA) level

Fig. 1  Aerial part fresh (a) and dry (b) weight (g) of rosemary plants Fig. 2  Roots fresh (a) and dry (b) weight (g) of rosemary plants
grown under different light intensity (LI) and water availability (WA) grown under different light intensity (LI) and water availability (WA)
conditions conditions

obtained in this study by applying decreasing levels of WA,
mainly consisting of a decrease of whole plant and aerial
part biomass, stem diameter and number of total shoots,
and of an increase of root biomass. In analogy to LI effects,
changes associated to variation in WA can be interpreted as
a plant strategy for growth optimization: in this case, under
conditions of reduced WA, plant response involves enhanced
investment of dry matter into structures that absorb water,
while structures that consume water are downsized. This is
shown also by the reduction of the aerial part/root weight
ratio, similarly to that previously reported in the literature
on various species, including rosemary, under conditions
of water stress [9, 33]. This kind of effects occurred in the
present study in spite of the relatively moderate water stress
imposed: it must be remarked that this study was designed
to evaluate plant growth in normal cultivation conditions,
where water stresses can sometimes occur but are not usu-
ally severe. Interestingly, in the present study the combined
application of two factors, LI and WA, allowed to highlight
also that the above effects due to WA occurred only in plants
exposed to the full solar radiation. In essence the moder-
ate water stress condition caused a moderate effect on plant
growth and this effect was observable only under conditions
of maximum solar radiation and photosynthetic activity. This
was in accordance with previous observations of water stress
effects on rosemary plant growth [24], where it was observed
that decreasing levels of WA reduced biomass formation
through declining photosynthetic activity and that the effect
of reduction of photosynthetic activity was higher at higher
levels of photosynthetic active radiation.
Effects on EO yield
Essential oil yield, expressed as percentage of fresh bio-
mass weight, ranged in the 9 treatments between 0.29 and
0.43% (Fig. 3), being relatively low when compared to Ital-
ian national production standard, 0.5–0.6% [34], or with data
reported in the international literature, 0.5–0.9% [1]. This
relatively low yield could be due to the harvest season in
the present study, in mid September, whereas the highest
yields are known to be obtained during late spring and early
summer. Secondly, dried material used for EO extraction
included almost all the upper aerial parts of the plant, com-
prising a higher fraction of ligneous parts than in common
practice. This procedure for plant material sampling was

13

172
(a)
0.5
EO yield as % of fresh biomass weight
0.45
0.4
0.35
0.3
0.25
0.2
85
Water availability (WA) level
(b)
1.5
EO yield as amount per plant (g)
1.3
1.1
0.9
0.7
0.5
85
Water availability (WA) level
Fig. 3  Essential oil (EO) yield of rosemary plants grown under dif-
ferent light intensity (LI) and water availability (WA) conditions. EO
yield expressed as % of fresh biomass (a) and as amount per plant (b)
preferred to obtain enough material for each sample to be
used for EO distillation.
Results from the two-factorial experiment showed that
both LI and WA had a significant effect on EO yield when
expressed as amount per plant, whereas only LI significantly
affected EO yield when expressed as % of fresh biomass
weight (Table 2). The interaction between the two factors
was significant only in the case of EO yield expressed as
amount per plant. Globally, the extent of the effect due to
changes in LI was higher than the one related to variations
in irrigation conditions. As regards the effect of LI, plants
exposed to 50% sunlight produced, on average, the highest
EO level when expressed as % content on fresh biomass.
This situation partly changed when considering EO yield on
a plant basis, due to the negative effect of decreasing levels
of LI on fresh biomass production (Table 1): for this reason,
EO yields on a plant basis were not significantly affected by
LI reduction from 100 to 50%, whereas they were markedly
reduced in plants grown under 25% LI. WA significantly
affected EO yield on a plant basis, by reducing it at the low-
est irrigation level (55%). All these effects may be observed
in detail in Fig. 3, where mean values for each treatment
are reported. In particular, the interaction between the two
investigated factors in affecting EO yield on a plant basis
(Fig. 3b) is also clearly highlighted. EO yield on a plant
13
European Food Research and Technology (2020) 246:167–177
Light intensity (LI) level basis became lower and lower as WA decreased, but this
100% LI 50% LI 25% LI
effect only occurred on plants exposed to 100% LI; under
this light condition, EO yield on a plant basis was reduced
by approximately one-third when reducing WA from 85 to
55% of field capacity (Fig. 3b).
Terpenoids, which are the constituents of rosemary EO,
are secondary metabolites synthesized in leaf protuberances
termed peltate glandular trichomes from freshly fixed car-
bon and their accumulation is thought to depend on ­CO2
acquisition from air and formation of photosynthesis inter-
70 55 mediates [15]. Differently from what observed in the present
study, a decreased % EO content of the fresh biomass in
Light intensity (LI) level
plants exposed to reduced solar radiation has been observed
100% LI 50% LI 25% LI
in Thymus vulgaris [16], Origanum vulgare [17], Ocimum
basilicum [18], Mentha piperita [19], and has been gener-
ally attributed to reduced photosynthetic activity and cor-
responding level of basic metabolites. Results from other
studies also suggested that more intense solar radiation can
lead to higher accumulation of secondary metabolites as part
of a plant defence mechanism against exceeding level of
radiation [21]. On the contrary, in other cases, shading of
70 55
solar radiation was associated to increased % EO content
of the fresh biomass as observed in Salvia officinalis [16],
in Eucalyptus citriodora [21] and Mentha piperita [20]. In
these two last cases, this was explained based on the protec-
tion conferred by shading against the severe radiation stress
imposed on the plants by exposure to very intense and harm-
ful irradiance level. This explanation, however, did not seem
to apply to the enhanced EO yield observed at the 50% sun-
light level in the present study, because in this case plants
exposed to the highest LI also produced the highest level of
biomass, thus not denoting a condition of severe radiation
stress. In accordance with this observation, leaf temperatures
recorded throughout a day in August (Table 3) suggests that
the aerial part of plants grown at 100% sunlight level did not
experience a heat stress condition more than the other plants.
Data from the present study suggest that at a low level of
LI (25% of natural sunlight) the reduced amount of radiation
is the dominant factor in determining a lowered production
of biomass and EO, according to most results reported in the
above cited literature. On the contrary, at the highest level of
LI, production of EO seems to be decoupled from biomass
production. For this unexpected result one possible expla-
nation may be proposed. Results reported in Table 1 show
that the number of nodes developed on the trunk in the last
2 months of growth was much lower in plants exposed to full
sunlight than under shading. This means that in the final time
of cultivation leaves on plants under full sunlight were, on
the whole, older than the ones on plants exposed to the other
LI treatments, and so volatiles produced on these leaves
could have been already partially lost at the harvest time.
The growth rate reduction of these plants during the last
phase of growth (mid and late summer) may be associated
European Food Research and Technology (2020) 246:167–177 173

Table 2  Data on rosemary EO yield and composition (% level of each EO constituent)

Variable 2-way ANOVA (p values)1 Light intensity (LI) levels Water availability (WA) levels

Light intensity (LI) Water avail- Interaction 100% 50% 25% 85% 70% 55%
ability (WA) LI × WA

EO yield
 % Content of the fresh biomass 0.000*** 0.460 0.232 0.31 ­b2 0.40 a 0.34 b 0.35 0.36 0.34
 Amount per plant (g) 0.000*** 0.010* 0.030* 1.09 a 1.25 a 0.62 b 1.00 a 1.04 a 0.86 b
EO composition

Compound Retention
index

 Tricyclene 925 0.006** 0.073 0.860 0.4 b 0.5 a 0.4 b 0.5 0.4 0.4
 α-Pinene 935 0.001** 0.029* 0.777 30.5 b 37.8 a 31.2 b 36.2 a 32.4 ab 31.0 b
 Camphene 948 0.022* 0.054 0.940 6.9 ab 7.9 a 6.8 b 7.8 7.0 6.8
 Thuja-2,4-diene 950 0.001** 0.074 0.761 0.9 a 1.1 a 0.7 b 1.0 0.9 0.8
 1-Octen-3-ol 961 0.113 0.292 0.178 0.2 0.2 0.2 0.2 0.2 0.2
 3-Octanone 965 0.001*** 0.373 0.087 2.9 a 3.0 a 2.3 b 2.8 2.8 2.6
 β-Pinene 974 0.002** 0.651 0.814 1.8 b 2.3 a 1.8 b 2.0 1.9 1.9
 Myrcene 983 0.000*** 0.033* 0.709 3.3 b 3.8 a 3.0 b 3.7 a 3.2 b 3.3 ab
 α-Phellandrene 1000 0.000*** 0.051 0.481 0.2 b 0.3 a 0.2 b 0.3 0.2 0.2
 α-Terpinene 1012 0.000*** 0.055 0.464 0.6 c 0.8 a 0.7 b 0.7 0.7 0.6
 p-Cymene 1014 0.000*** 0.033* 0.068 1.4 a 1.1 b 0.9 c 1.2 a 1.1 ab 1.1 b
 1,8-Cineole 1023 0.009** 0.204 0.492 13.5 ab 13.8 a 12.7 b 13.7 13.2 13.1
 γ-Terpinene 1051 0.000*** 0.073 0.380 0.8 b 1.0 a 0.9 a 1.0 0.9 0.9
 Terpinolene 1081 0.000*** 0.126 0.252 1.1 b 1.4 a 1.4 a 1.3 1.3 1.3
 3-Carene 1084 0.005** 0.072 0.360 2.8 a 2.3 b 2.9 a 2.4 2.7 2.8
 Chrysanthenone 1103 0.020* 0.414 0.532 1.2 a 0.9 b 1.0 ab 0.9 1.0 1.1
 Camphor 1125 0.001*** 0.085 0.830 7.4 a 5.6 b 8.1 a 6.3 7.2 7.5
 Unknown 1 1130 0.008** 0.089 0.677 0.4 a 0.2 b 0.5 a 0.2 0.4 0.4
 Borneol 1153 0.000*** 0.049* 0.561 8.8 a 4.6 c 6.8 b 5.5 b 7.1 ab 7.6 a
 1-Terpinen-4-ol 1164 0.001*** 0.070 0.708 1.2 a 0.7 b 1.0 a 0.8 1.0 1.1
 α-Terpineol 1174 0.002** 0.077 0.849 1.7 a 0.9 b 1.6 a 1.1 1.5 1.6
 Verbenone 1183 0.014* 0.149 0.766 4.4 ab 2.4 b 5.2 a 3.0 4.5 4.5
 Unknown 2 1220 0.002** 0.050 0.703 0.8 a 0.4 b 0.8 a 0.5 0.7 0.8
 Unknown 3 1226 0.002** 0.054 0.729 1.2 a 0.5 b 1.1 a 0.7 1.0 1.1
 Bornyl acetate 1271 0.000*** 0.000*** 0.200 2.8 b 2.9 b 4.7 a 2.9 b 3.5 b 4.1 a
 β-Caryophyllene 1423 0.001** 0.311 0.326 2.1 b 3.1 a 2.1 b 2.7 2.4 2.3
 Humulene 1459 0.011* 0.517 0.365 0.3 b 0.4 a 0.3 ab 0.3 0.3 0.3
 Unknown 4 1576 0.008** 0.047* 0.671 0.7 a 0.2 b 0.7 a 0.3 b 0.5 ab 0.7 a

Results of 2-way ANOVA and mean values of samples belonging to the same factor (light intensity and water availability) level. Tukey’s multi-
ple comparison test (at p = 0.05)
1
 Statistical significance of ANOVA: *p < 0.05; **p < 0.01; ***p < 0.001
2
 When a factor was significant at ANOVA the Tukey’s multiple comparison test was performed to compare data related to different levels of
each factor. In the box of light intensity or water availability, different letters within the same row denoted significant differences between differ-
ent levels of the factor, according to Tukey’s test (at p < 0.05)

to a heat stress occurred at the roots level, only during this
phase, as the temperature of pot external surface during the
day tended to be higher at full sunlight than under shading,
rising up to more than 48 °C in the afternoon (Table 3).
When considering the effects of WA, differently from what
observed in the present study, reduction of WA strongly
increased EO % content of fresh biomass in a previous
study on rosemary [24]. However, in that experiment [24],
the extent of WA reduction was much higher (50 to 100%
reduction of estimated plant water use was compared to 0%
reduction) than in the present study, where the purpose was
to explore irrigation conditions within ranges compatible
with crop cultivation management rather than evaluation of
extreme environmental stress conditions. As a consequence,
the smaller extent of WA reduction may explain the absence
of an effect on EO % content of fresh biomass in the present
study. In the previous study, the strong increases in EO %
content of fresh biomass corresponded to slight increases or
reduction of EO yield when expressed on a plant basis, due
to the parallel reduction of produced biomass [24]. For the

13

174 European Food Research and Technology (2020) 246:167–177

Table 3  Temperature (°C) of Measured parameter Time of day of Recorded temperature (°C) 1-way
leaves and pot external surface measurement ANOVA (p
recorded throughout a day Light intensity (LI) levels values)2
during the last phase of growth
on plants subjected to the 100% 50% 25%
different levels of light intensity
Leaf temperature (°C)1 10.30 am 29.3 b 28.8 b 30.3 a 0.002**
1.30 pm 31.7 b 31.9 ab 33.4 a 0.020*
4.30 pm 29.9 b 29.7 b 31.4 a 0.000***
Pot surface temperature (°C)1 10.30 am 39.9 a 35.1 b 35.1 b 0.000***
1.30 pm 40.4 a 37.4 b 37.0 b 0.000***
4.30 pm 48.1 a 43.6 b 41.7 b 0.000***
1
 Temperature measurements recorded throughout a day, on August 24th 2017
2
  Statistical significance of ANOVA: *p < 0.05; **p < 0.01; ***p < 0.001

same reason, in the present study the absence of an effect
of water reduction on EO % content of fresh biomass cor-
responded to a reduction of EO yield per plant. This effect
occurred only in plants exposed to 100% LI, because only
these plants experienced a significant decrease of biomass
formation due to WA reduction (Fig. 2S), for the reason dis-
cussed above. In any case, results from plants exposed to full
sunlight highlighted a marked negative effect of reduction
of WA on productivity, in terms of EO yield, and this would
suggest taking into account this effect when applying water
shortage strategies in the cultivation of this crop.
Effects on EO composition
GC–MS analysis allowed to determine 28 major constitu-
ents of the EO of the “erect rosemary” variety used for the
experiment, whose profile resulted to be similar to that one
of the ecotype Cevoli from a not distant geographical area
[34]. Determined compounds belonged to the chemical
classes of monoterpene hydrocarbons (n. 12), oxygenated
monoterpenes (n. 10), sesquiterpenes (n. 2) and unidenti-
fied terpenoid compounds (n. 4), whereas major constitu-
ents were α-pinene (28.9–39.4%), 1,8-cineole (12.3–13.8%),
camphene (6.3–8.2%), camphor (5.1–9.0%) and borneol
(4.3–9.8%) (Table 2). Global effects of the investigated fac-
tors on the EO profile were shown by results of the PCA
reported in Fig. 4. A substantial percentage of the variance
within the dataset was explained by the first two PCs, 91.6%
and 4.8%, respectively. Similarly to what observed on EO
yield data, the effect of LI was more pronounced than that
associated to WA, as highlighted by the fact that treatments
corresponding to the same LI level can be grouped, as
shown in Fig. 4, whereas this was not possible for treatments
belonging to the same WA level. PC1 tended to discriminate
plants subjected to 50% LI from plants exposed to the other
light levels, whereas PC2 discriminated plants subjected to
100% and 25% LI. Data on PC loadings (Table 1S) high-
lighted a high contribution of α-pinene (positive loadings),
borneol, verbenone and camphor (negative loadings) to the
PC1, and a high contribution of borneol, 1,8-cineole (posi-
tive loadings), verbenone and bornyl acetate (negative load-
ings) to the PC2. A higher impact of the evaluated light
conditions than irrigation regimes was also confirmed by
results of ANOVA of data on EO composition (Table 2). LI
conditions affected the percent level of 27 out of the deter-
mined 28 EO constituents, whereas WA affected the level
of only 6 compounds. Interactions between the two factors
were not significant. Again, as regards the effect of LI, major
differences were observed between plants subjected to 50%
LI and the other two groups of plants. A group of com-
pounds, such α-pinene, 1,8-cineole, camphene, β-pinene,
myrcene, α-phellandrene, α-terpinene and β-caryophillene
showed enhanced relative abundances in plants exposed to
50% sunlight, whereas a second group of compounds, such
as 3-carene, camphor, borneol, 1-terpinen-4-ol, α-terpineol,
verbenone, humulene and three unknown compounds
(unknown 1, 2, 3), showed lowered abundances in these
plants. Major changes due to reduction of light intensity
from 100 to 50% were observed for α-pinene (on average
+ 24%), camphor (− 24%), borneol (− 44%) and verbenone
(− 45%). ANOVA results also allowed to highlight effects of
WA level on EO composition. Reducing water availability
significantly lowered the relative content of α-pinene (on
average − 14%), myrcene and p-cymene, while increased
the relative abundance of borneol (+ 38%), bornyl acetate
(+ 41%) and compound unknown 4 (Table 2).
Biosynthesis of terpenoids occurs through diverse path-
ways, by which precursors of both monoterpenoids (geranyl
diphosphate) and sesquiterpenoids (farnesyl diphosphate)
are formed. Then, from these precursors all the other mem-
bers of the groups are formed by complex series of reactions,
which are all under enzymatic control [35]. Some of the key
enzymes involved in monoterpene biosynthesis (monoter-
pene synthases) have been studied in detail in Salvia offici-
nalis, where some pinene synthases have been recognized
as responsible for the synthesis of α- and β-pinene and

13
European Food Research and Technology (2020) 246:167–177 175
Fig. 4  PCA of data of composition of rosemary EOs obtained from
plants subjected to different levels of light intensity and water avail-
ability. Bi-plot (score and loadings plot) of the first two PCAs. Labels
in black character correspond to data points of the score plot (EO
samples). The first number of the label denotes the level of LI (100%,
50% or 25% of natural sunlight), the second number denotes the level
camphene, and to contribute to the synthesis of myrcene,
whereas bornyl diphosphate synthase has been recognized
as responsible for the synthesis of borneol and camphor,
and 1,8-cineole synthase for the synthesis of 1,8-cineole
[36]. Results from several studies have shown that changes
in LI and WA can have a significant effect on EO compo-
sition [17–19, 22, 37], whereas different mechanism have
been postulated to mediate these effects [19, 22, 37]. In
the present study, major changes in EO composition where
observed in plants subjected to 50% sunlight. Formation of
compounds catalysed by pinene synthase enzymes, as for the
major constituent α-pinene, but also β-pinene, camphene and
myrcene, seemed to be significantly promoted by the 50% LI
condition, when compared to the other LI levels, whereas the
contrary was observed for compounds formed through the
activity of bornyl diphosphate synthase, such as borneol and
camphor. Also the relative abundance of 1,8-cineole, which
is formed by 1,8-cineole synthase, is enhanced in plants
exposed to 50% LI, even though the extent of this effect was
smaller than that observed on α-pinene. An effect of the con-
sidered LI conditions on these enzymatic activities may be
involved in the observed changes in the EO profile and could
be confirmed by future investigations on other rosemary
of WA (85%, 70% or 55% of field capacity), the third number denotes
the treatment replicate (1, 2 or 3). Labels in blue character correspond
to data points of the loadings plot (EO constituents). Coloured area
are delimited by convex hulls, which are the smallest convex poly-
gon enclosing all the data points (EO samples) corresponding to each
level of LI (green: 25% LI; red: 50% LI; blue: 100 LI)
cultivars. Interestingly, verbenone, which may be produced
by conversion of α-pinene, was markedly influenced by
light conditions in an opposite way when compared to its
plausible precursor α-pinene. A similar opposite effect on
pinene synthase derived compounds and bornyl diphosphate
synthase derived compounds was also observed in relation
to irrigation regimes, where reduction of WA promoted the
accumulation of borneol and bornyl acetate (bornyl diphos-
phate synthase related compounds), while inhibiting the for-
mation of α-pinene and myrcene (pinene synthase related
compounds). Interestingly, the effects observed on borneol
and bornyl acetate, and also a similar though not significant
effect on camphor, seemed to confirm recent results by Rad-
wan et al. [37], who observed that a moderate drought stress
strongly up-regulates bornyl diphosphate activity in Salvia
officinalis, thus promoting the accumulation of camphor in
that species. On the contrary, in a previous study on rose-
mary, no changes in EO composition were observed in plants
subjected to severe drought stress [24]. In any case, in the
present experiment, it seemed that changes in both light and
water conditions affected in opposite ways the formation of
compounds derived from the activity of pinene synthase and
bornyl diphosphate synthase.
13

176
Conclusions
In the pot growing conditions considered in the present
experiment changes in LI markedly affected rosemary plant
growth and architecture as well as EO yield. High shading
of natural sunlight markedly hindered plant biomass produc-
tion and EO accumulation, whereas intermediate shading
conditions significantly promoted EO yield when expressed
as % content of fresh biomass, but not significantly when
expressed as amount per plant, with respect to 100% natu-
ral sunlight. Reduction of WA, within ranges compatible
with crop cultivation conditions, may markedly reduce crop
productivity in terms of EO yield on a plant basis, but only
when combined with exposure to full sunlight radiation.
This highlights the importance of taking into account the
possible occurrence of interactions between multiple factors
affecting EO yield when evaluating the influence of envi-
ronmental factors in crop management. In addition both
LI and WA levels significantly affected EO composition.
Changes in the level of the two factors seemed to have an
opposite effect on the relative abundance of volatiles that
are formed through the activity of two groups of enzymes,
pinene synthases and, respectively, bornyl diphosphate syn-
thases. This may have implications on EO sensory quality,
given that volatiles derived from the two enzyme groups,
such as α-pinene, camphene, borneol, camphor, are known
to be key odorants of rosemary EO and are characterized by
different aroma notes [38]. Moreover, variation in rosemary
EO composition is associated to changes in its biological
properties, such as antimicrobial and antioxidant activities
[39]. Thus results from the present study also suggest the
importance of a careful management of light and irrigation
conditions to standardize the composition of rosemary EO
or to modulate it in view of different potential uses.
Acknowledgements  The study was carried out with the financial sup-
port of the Ministry of Agriculture, Food and Forestry Policies and
Tourism, Italy, within the project “Implementation of the FAO Inter-
national Treaty on Plant Genetic Resources for Food and Agriculture”.
Compliance with ethical standards  vulgare L. ssp. hirtum (Link) Ietswaart. Ind Crops Prod 34:1516–
Conflict of interest  The author declares that they have no competing
interests.
Ethical approval  This article does not contain any studies with human
or animal subjects.
References
1. Sasikumar B (2012) In: Peter KV (ed) Handbook of herbs and
spices, vol 1, 2nd edn. Woodhead Publishing Limited, Cambridge
13
European Food Research and Technology (2020) 246:167–177
2. Brud WS (2009) In: Başer KHC, Buchbauer G (eds) Handbook of
essential oils: science, technology, and applications. CRC Press,
Boca Raton
3. Lubbe A, Verpoorte R (2011) Cultivation of medicinal and aro-
matic plants for specialty industrial materials. Ind Crops Prod
34:785–801. https​://doi.org/10.1016/j.indcr​op.2011.01.019
4. del Pilar Sánchez-Camargo A, Herrero M (2017) Rosemary (Ros-
marinus officinalis) as a functional ingredient: recent scientific
evidence. Curr Opin Food Sci 14:13–19. https:​ //doi.org/10.1016/j.
cofs.2016.12.003
5. Ribeiro-Santos R, Andrade M, de Melo NR, Sanches-Silva A
(2017) Use of essential oils in active food packaging: recent
advances and future trends. Trends Food Sci Technol 61:132–140.
https​://doi.org/10.1016/j.tifs.2016.11.021
6. Cervelli C, Fadelli PG, Tallone A (2001) Growth of Oreopanax
capitatus and Cocculus laurifolius in different protected environ-
ments. Acta Hortic 559:91–96
7. Conover C (1990) In: Novak J, Rudnicki RM (eds) Postharvest
handling and storage of cut flowers, florist’s green and potted
plants. Timber Press, Portland
8. Gratani L, Covone F, Larcher W (2006) Leaf plasticity in response
to light of three evergreen species of the Mediterranean maquis.
Trees 20:549–558
9. Gregory PJ, Palta JA, Batts GR (1997) Root systems and root:
mass ratio—carbon allocation under current and projected atmos-
pheric conditions in arable crops. Plant Soil 187:221–228
10. Kozlowski TT, Pallardy SG (1997) Growth control in woody
plants. Academic Press, San Diego
11. Mielke MS, Schaffer B (2010) Photosynthetic and growth
responses of Eugenia uniflora L. seedlings to soil flooding
and light intensity. Environ Exp Bot 68:113–121. https​://doi.
org/10.1016/j.envex​pbot.2009.11.007
12. Osakabe Y, Osakabe K, Shinozaki K, Tran L-SP (2014) Response
of plants to water stress. Front Plant Sci 5:86
13. Figueiredo AC, Barroso JG, Pedro LG, Scheffer JJC (2008) Fac-
tors affecting secondary metabolite production in plants: volatile
components and essential oils. Flavour Fragr J 23:213–226. https​
://doi.org/10.1002/ffj.1875
14. Franz C, Novak J (2009) In: Başer KHC, Buchbauer G (eds)
Handbook of essential oils: science, technology, and applications.
CRC Press, Boca Raton
15. Loreto F, Ciccioli P, Cecinato A et al (1996) Influence of envi-
ronmental factors and air composition on the emission of [alpha]-
pinene from quercus ilex leaves. Plant Physiol 110:267–275
16. Li Y, Craker LE, Potter T (1996) Effect of light level on the essen-
tial oil production of sage (Salvia officinalis) and thyme (Thymus
vulgaris). Acta Hortic 426:419–426
17. Tibaldi G, Fontana E, Nicola S (2011) Growing conditions and
postharvest management can affect the essential oil of Origanum
1522. https​://doi.org/10.1016/j.indcr​op.2011.05.008
18. Chang X, Alderson PG, Wright CJ (2008) Solar irradiance level
alters the growth of basil (Ocimum basilicum L.) and its con-
tent of volatile oils. Environ Exp Bot 63:216–223. https​://doi.
org/10.1016/j.envex​pbot.2007.10.017
19. Rios-Estepa R, Turner GW, Lee JM et al (2008) A systems biol-
ogy approach identifies the biochemical mechanisms regulating
monoterpenoid essential oil composition in peppermint. Proc Natl
Acad Sci 105:2818–2823. https​://doi.org/10.1073/pnas.07123​
14105​
20. Mousavinik SM, Asgharipour MR, Sardashti S (2016) Manure
and light intensity affect growth characteristics and essential oil
of peppermint (Mentha piperita L.). J Essent Oil Bear Plants
19:2029–2036. https​://doi.org/10.1080/09720​60X.2016.12424​35
European Food Research and Technology (2020) 246:167–177 177
21. Degani AV, Dudai N, Bechar A, Vaknin Y (2016) Shade effects on
leaf production and essential oil content and composition of the
novel herb Eucalyptus citriodora hook. J Essent Oil Bear Plants
19:410–420. https​://doi.org/10.1080/09720​60X.2014.89008​0
22. Selmar D, Kleinwächter M (2013) Influencing the product qual-
ity by deliberately applying drought stress during the cultivation
of medicinal plants. Ind Crops Prod 42:558–566. https​://doi.
org/10.1016/j.indcr​op.2012.06.020
23. Chrysargyris A, Laoutari S, Litskas VD et al (2016) Effects of
water stress on lavender and sage biomass production, essential oil
composition and biocidal properties against Tetranychus urticae
(Koch). Sci Hortic 213:96–103. https​://doi.org/10.1016/j.scien​
ta.2016.10.024
24. Delfine S, Loreto F, Pinelli P et al (2005) Isoprenoids content
and photosynthetic limitations in rosemary and spearmint plants
under water stress. Agric Ecosyst Environ 106:243–252. https​://
doi.org/10.1016/j.agee.2004.10.012
25. Li G, Cervelli C, Ruffoni B et al (2016) Volatile diversity in wild
populations of rosemary (Rosmarinus officinalis L.) from the Tyr-
rhenian Sea vicinity cultivated under homogeneous environmental
conditions. Ind Crops Prod 84:381–390. https:​ //doi.org/10.1016/j.
indcr​op.2016.02.029
26. Thakur M, Bhattacharya S, Khosla PK, Puri S (2018) Improv-
ing production of plant secondary metabolites through biotic and
abiotic elicitation. J Appl Res Med Aromat Plants. https​://doi.
org/10.1016/j.jarma​p.2018.11.004
27. Cingolani E (1973) Italian pharmacopoeia—8th edition. Farmaco
Prat 28(11):559–584
28. NIST (National Institute of Standards and Technology), NIST
chemistry WebBook, NIST standard reference database number
69, 2018, available online at: http://webbo​ok.nist.gov/chemi​stry.
Accessed on July 2019
29. Hammer O, Harper D, Ryan P (2012) PAST: paleontological sta-
tistics software package for education and data analysis. Paleontol
Electron 4(art. 4):9
30. Givnish T (1988) Adaptation to sun and shade: a whole-plant per-
spective. Funct Plant Biol 15:63. https​://doi.org/10.1071/PP988​
0063
31. Kumar R, Sharma S, Pathania V (2013) Effect of shading and
plant density on growth, yield and oil composition of clary sage
(Salvia sclarea L.) in north western Himalaya. J Essent Oil Res
25:23–32. https​://doi.org/10.1080/10412​905.2012.74246​7
32. Zervoudakis G, Salahas G, Kaspiris G, Konstantinpoulou E (2012)
Influence of light intensity on growth and physiological charac-
teristics of common sage (Salvia officinalis L.). Braz Arch Biol
Technol 55:88–95. https​://doi.org/10.1590/S1516​-89132​01200​
01000​11
33. Sánchez-Blanco JM, Ferrández T, Navarro A, Bañon S, José
Alarcón J (2004) Effects of irrigation and air humidity precon-
ditioning on water relations, growth and survival of Rosmarinus
officinalis plants during and after transplanting. J Plant Physiol
161:1133–1142. https​://doi.org/10.1016/j.jplph​.2004.01.011
34. Flamini G, Cioni PL, Morelli I et al (2002) Main agronomic—pro-
ductive characteristics of two ecotypes of Rosmarinus officinalis
L. and chemical composition of their essential oils. J Agric Food
Chem 50:3512–3517. https​://doi.org/10.1021/jf011​138j
35. Sell C (2009) In: Başer KHC, Buchbauer G (eds) Handbook of
essential oils: science, technology, and applications. CRC Press,
Boca Raton
36. Wise ML, Savage TJ, Katahira E, Croteau R (1998) Monoterpene
synthases from common sage (Salvia officinalis) cDNA isolation,
characterization, and functional expression of (+)-sabinene syn-
thase, 1, 8-cineole synthase, and (+)-bornyl diphosphate syn-
thase. J Biol Chem 273:14891–14899. https​://doi.org/10.1074/
jbc.273.24.14891​
37. Radwan A, Kleinwächter M, Selmar D (2017) Impact of drought
stress on specialised metabolism: biosynthesis and the expression
of monoterpene synthases in sage (Salvia officinalis). Phytochem-
istry 141:20–26. https:​ //doi.org/10.1016/j.phytoc​ hem.2017.05.005
38. Kamath A, Asha MR, Ravi R et al (2001) Comparative study of
odour and GC-olfactometric profiles of selected essential oils.
Flavour Fragr J 16:401–407. https​://doi.org/10.1002/ffj.1020
39. Zaouali Y, Bouzaine T, Boussaid M (2010) Essential oils com-
position in two Rosmarinus officinalis L. varieties and incidence
for antimicrobial and antioxidant activities. Food Chem Toxicol
48:3144–3152. https​://doi.org/10.1016/j.fct.2010.08.010
Publisher’s Note Springer Nature remains neutral with regard to
jurisdictional claims in published maps and institutional affiliations.
13