This article was downloaded by: [The University of Kashmir ], [Sumira Jan]

On: 13 October 2014, At: 00:24

Publisher: Taylor & Francis
Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House,
37-41 Mortimer Street, London W1T 3JH, UK

Israel Journal of Plant Sciences

Publication details, including instructions for authors and subscription information:
http://www.tandfonline.com/loi/tips20

Variation in adaptation mechanisms of medicinal herbs

to the extreme winter conditions across the North
Western Himalaya
a

Sumira Jan , Azra N. Kamili , Rehana Hamid & Javid A. Parray

Centre of Research for Development (CORD), Kashmir University, India

Published online: 04 Sep 2014.

To cite this article: Sumira Jan, Azra N. Kamili, Rehana Hamid & Javid A. Parray (2014): Variation in adaptation mechanisms
of medicinal herbs to the extreme winter conditions across the North Western Himalaya, Israel Journal of Plant Sciences,
DOI: 10.1080/07929978.2014.939828
To link to this article: http://dx.doi.org/10.1080/07929978.2014.939828

PLEASE SCROLL DOWN FOR ARTICLE

Taylor & Francis makes every effort to ensure the accuracy of all the information (the Content) contained
in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no
representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the
Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and
are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and
should be independently verified with primary sources of information. Taylor and Francis shall not be liable for
any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever
or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of
the Content.
This article may be used for research, teaching, and private study purposes. Any substantial or systematic
reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any
form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http://
www.tandfonline.com/page/terms-and-conditions

Israel Journal of Plant Sciences, 2014

http://dx.doi.org/10.1080/07929978.2014.939828

Variation in adaptation mechanisms of medicinal herbs to the extreme winter conditions across
the North Western Himalaya
Sumira Jan*, Azra N. Kamili, Rehana Hamid and Javid A. Parray
Centre of Research for Development (CORD), Kashmir University, India

Downloaded by [The University of Kashmir ], [Sumira Jan] at 00:25 13 October 2014

(Received 21 May 2014; accepted 26 June 2014)

The North Western Himalaya region is characterized by extreme seasonal variability and local changes in microclimate.
Plants adapted to this region demonstrate marked variability to the extreme winter conditions. This region, designated as a
biodiversity hotspot, is rich in medicinal herbs, which exhibit diverse growth and adaptation mechanisms to the harsh
environment. In the present study, physiological mechanisms of adaptation to the winter season conditions were compared
in five medicinal herbs. Components of the photosynthetic machinery, and osmoregulation which are of importance for
secondary metabolism is not well known in these alpine herbs and was studied. The medicinal herbs Rumex dentatus,
Atropa accuminata, Lupinus polyphyllus, Hyoscyamus niger, and Lavandula officinalis were collected before snowfall in
September
November to evaluate variability in metabolic and physiological responses to the varied seasonal regimes.
Plants were followed over a period of 8 weeks (summer to early winter) in three sites which vary in altitude and thereby
climatic conditions: Site I, 4175 m (Pahalgam); Site II, 3748 m (Duksum Sinthan Top); and Site III, 4100 m (Mount
Apharwat). Variations in photosynthetic capacity and components of the photosynthetic machinery, a range of
antioxidants, antioxidant enzymes and total non-structural carbohydrates (TNC) were studied to test their involvement in
adaptation schemes and secondary metabolism under the harsh winter conditions. The results demonstrate a range of
adaptations to the freezing stress conditions including high photon energy use efficiency; an increase in the proportion of
Chl b, carotenoid content; specific activity of the antioxidant enzymes superoxide dismutase, catalase, glutathione
reductase, ascorbate peroxidase activity, and TNCs.
Keywords: Western Himalaya; aromatic and medicinal herbs; snowfall; antioxidative enzymes; carotenoid

1. Introduction
Kashmir, located at the northwestern region of the
Indian subcontinent, contains the northwestern Himalayan ranges which are characterized by marked geographical and climatic variation, accompanied by a
large biodiversity. Snow-capped summits, drainage valleys, complex geological structure and rich temperate
flora and fauna characterize this diverse region, which
is therefore noted to be a combination of microclimates
(Mittermeier et al. 2005). Valley climate displays
marked seasonality typical of inner continental temperate latitudes. As the altitude rises towards the meadow
slopes (margs) of the surrounding mountains, the temperature decreases, from 24 C at Srinagar (altitude
1600 m) to 10 C at an elevation of 3600 m. The average
valley temperature ranges from 31 C to 15 C in July to
4 C to 4 C in January. High-altitude ecosystems are
considered to be hotspots of medicinal plant diversity,
but are neglected areas for research due to their inaccessibility and harsh climatic conditions. Many environmental factors such as variation in altitude, duration of
snow cover, temperature extremes, length of the

Corresponding author. Email: sumira.sam@gmail.com

2014 Taylor & Francis

vegetation seasons, and radiation intensity affects the

plant growth, physiology and metabolic profile (Neuner
et al. 1999). It is well known that the growth response
of plants to extreme climatic conditions depends on
their prior exposure and characteristic ecophysiological
traits that largely differ among populations thriving at
different altitudes (Hacker & Neuner 2006).
Plants under snowfall are exposed to diffuse light or
darkness and moderate or constant temperatures around
0 C depending on the thickness and uniformity of the
snow cover. As a result, plants are exposed to rapid temporal changes in microclimate (Larcher & Bauer 1981).
Kappen (1993) illustrated the photosynthetic activity in
evergreen woody and herbaceous species from high
mountainous habitats at below freezing temperatures
between 2 C and 9 C. Ice formation in photosynthetic tissues can hamper photosynthetic activity
(Kutsch & Kappen 1997). High mountain plants such as
Rannunculus glacialis exhibit normal photosynthetic
activity even under 11 C (Tieszen et al. 1981). During
the winter, snow protects the photosynthetic tissue of
green plants from the stressful combination of

Downloaded by [The University of Kashmir ], [Sumira Jan] at 00:25 13 October 2014

S. Jan et al.

low-temperature extremes and high irradiation. Snow

ameliorates temperature extremes by supplying the environment with moisture and allowing light transmission
that is sufficient to facilitate moderate photosynthetic
activity (Kappen & Breuer 1991). However, photosynthetic pigments are the most susceptible to low-temperature injury. Knowledge concerning the photosynthesis
of green alpine plants during the period of snow coverage is still scarce and only few studies have focused on
the effects of the changing environmental conditions
during snow melt on photosynthesis.
The herbs considered in this study belong to three
different families: Rumex dentatus (Polyganeace);
Atropa accuminata, Lupinus polyphyllus and Hyoscyamus niger (Solanaceae) and Lavandula officinalis
(Lamiaceace). The selected plants for the study differ in
growth pattern and acclimation mechanism to the winter
conditions. Atropa accuminata reaches a maximum
height of 1.6 m and requires snow coverage for protection in winter. Rumex reach 0.8 m in height, and is
adapted for growth also at wind-exposed ridges, indicating that it belongs to the most typical climate-resistant
alpine herbs. The short vegetation phase of alpine plants
may be effectively utilized if green plants are able to
start photosynthesis even under a thin snow cover (Wildi
& L
utz 1996). The regular increase in irradiation after
the plants emergence from the snow exposes the herbs
to the melting water and the accompanying temperature
fluctuations. This condition can be damaging for
unadapted tissues (L
utz 1996). Investigations concerning the temperature dependency of reproductive development in mountain plants are lacking. At high alpine or
polar sites, low temperature may retard the processes of
flower development and fruit ripening. Early onset of
winter conditions may therefore threaten reproductive
success. Alpine plants have various morphological,
physiological and metabolic means of adaptation against
extreme environmental conditions. Despite the high ecological and economic importance of Himalayan medicinal plants, the effects of harsh climate on primary and
secondary metabolites are still poorly known.

In this study, we will compare photosynthetic performance during snow coverage and after snow melt in the
medicinal herbs Rumex dentata, Atropa accuminata,
Lupinus polyphyllus, Hyoscyamus niger, and Lavandula
officinalis. These herbs may not be protected by snow
throughout the winter period. With the aim of increasing
our understanding of UV resistance and physiological
adaptations to the extreme environmental conditions, we
investigated changes in photosynthetic stress parameters:
efficiency of photosystem II, changes in chloroplast pigments, ascorbic acid and a-tocopherol, and total flavonoids. The results contribute to our understanding of plant
tolerance to the harsh environmental conditions.
2. Material and methods
A total of three sites from a series of temperate and
sub-alpine areas were selected for the study (Table 1).
The selected sites are characterized by dense populations of herbs, and represent different altitudes of the
Western Himalayas with varying snowfall. These sites
were recognized by Kaul (1997) for their luxuriant
herbal charm.
2.1.

Survey and collection of plant material

at various altitudes
To study variation among the different sites, leaves from
the above-selected herbs at respective sites were collected to evaluate variability in metabolic and physiological responses. These herbs grow in open meadows and
sub-alpine mountain hills. Rumex dentatus is one of the
most prevalent temperate and sub-alpine herbs. The
sampling sites were selected as per luxuriant growth of
herbs, dense snow cover belts and represent different
acclimation stages. Samples were taken between late
September and early November 2012 (i.e. 26 September;
4 October; 17 October; 10 November) and put into liquid
N2 after fresh weight determination. Plants were followed over a period of about 8 weeks (summer to early
winter).

Table 1. Sites selected for the study, in the Western Himalayan ranges.

Character
Altitude (m above sea level)
Forest range
Climatic zones
Direction
Latitude, longitude
Snowfall (2012)

Pisu top (Pahalgam)

Site I

Daksum Sinthan top

Site II

Mount Apharwat (Gulmarg)

Site III

4175
Upper Dachigam
Sub-alpine
Southeast
34 003600 N, 75 1102400 E
244 cm Dense

3748
Anantnag
Sub-alpine
Southeast
34
20 N, 75
20 E
221 cm Less dense

4100
Pir Panjal
Temperate
sub-alpine
Northwest
34 030N, 74 230E/34.05; 74.38
308 cm Dense

Israel Journal of Plant Sciences

2.2.3. Enzyme assays

Superoxide dismutase (SOD) activity was measured
according to Mishra et al. (1993). Catalase (CAT) activity
was analyzed according to Aebi (1984). Ascorbate peroxidase (APX) activity was assayed according to Nakano and
Asada (1981). Glutathione reductase (GR) activity was
assayed according to Sen Gupta et al. (1993).

photosynthetic apparatus. The analysis was carried out in

late September to early November, and leaf temperature
recorded dramatic changes in microclimate (Table 2).
Levels of chlorophyll a and b, and the ratio of chlorophyll
a:b, differed significantly between Sites I and III during
the summer. However, in Site II the ratio of chlorophyll a:
b was significantly lower in plants compared with Sites I
and III (P D 0.005). This decline appeared to be caused
by a decrease in chlorophyll a, as sunny leaves exhibited
an average of 10% less chlorophyll a than shade leaves,
but only 2% less chlorophyll b (Table 2). Chlorophyll a
and b levels were both significantly lower in Site I compared to Sites II and III leaves (P D 0.002 and P D 0.058,
respectively). Similar comparison of TNCs showed that
starch was significantly higher in Site III leaves than Sites
I and II leaves (P < 0.0001), although starch levels did
not differ significantly between the two sites. In all herbs,
Sites II and III leaves exhibited significantly higher soluble sugar content compared to Site I, with a nearly twofold
increase (P < 0.0001). TNCs did not differ significantly
between sites, although TNCs were twofold higher in Site
I leaves compared with Site III (P < 0.0001) (Table 3).
Carotenoids play an important role in radiation damage and free radical scavenging. Significant (P < 0.05)
increase in carotenoids content was observed in leaves of
Atropa acuminata in Site I (75%); however, Sites II and
III did not show any significant difference (Table 2). The
lowest carotenoid content was recorded in Hyoscyamus
niger, with no significant differences between sites.
Furthermore, plants adapt to environmental constrains via
high photon energy use efficiency by increasing the
proportion of Chl b.

2.2.4.

3.2.

2.2.
2.2.1.

Downloaded by [The University of Kashmir ], [Sumira Jan] at 00:25 13 October 2014

Biochemical analysis
Analyses of plastid pigments, total non-structural
carbohydrates, ascorbic acid, soluble sugars and
starch

Chlorophyll content was determined using tissue from

three leaf discs derived from the same leaves as described
by Porra (2002). Total non-structural carbohydrates
(TNCs) were analyzed as described by Porra (2002). Isolation and ascorbic acid determination by ascorbate oxidase and DTT (dithiothreitol) were performed according
to Luwe et al. (1993) and quantification of soluble sugars
and determination of starch concentration were performed
following Volenec et al. (1991).
2.2.2.

Extraction and quantification of a-tocopherol

Tocopherol was estimated by the Emmerie
Engel reaction as reported by Rosenberg (1992). The concentration
of tocopherol in the sample was calculated by the formula
of Equation (1):
Tocopherol mg Sample A520

A460 0:290:15
Standard A520
(1)

Thiobarbituric acid reactive substances

Thiobarbituric acid reactive substances, or TBARS, considered as oxidative damage products, were determined in
leaf samples following Heath and Packer (1968) with
slight modifications.
2.2.5.

Proline, phenols, flavonols, alkaloids and

anthocyanins
Proline content in leaf samples was estimated by the
method of Bates et al. (1973). Total phenolics was
assayed following Wolfe et al. (2003), flavonoids and flavonols following Ordonez et al. (2006), proanthocyanidins according to Sun et al. (1998), and total alkaloid
fraction as described in Wolfe et al. (2003).
3. Results
3.1.

Plastid pigments and total non-structural

carbohydrates
Results of pigment analyses reveal that in the spring under
the snow the plants have a fully developed, well-protected

Variation in total ascorbic acid and

a-tocopherol content
a-tocopherol and ascorbic acid represent the main antioxidant systems of the lipophilic and hydrophilic compartments, respectively, in plastids. Tocopherols are
lipophilic antioxidants that are synthesized in photosynthetic organisms. In higher plants, ratios of a- and
g-tocopherol is under spatial and temporal control.
a-tocopherol accumulates mainly in photosynthetic tissue
and was therefore suggested to contribute in the detoxification of ROS (reactive oxygen species) mutually with
the hydrophilic antioxidants glutathione and ascorbate
(Foyer & Noctor 2003). Based on inhibitor studies, it has
been shown that tocopherol acts as singlet oxygen scavenger in photosynthesis II of Chlamydomonas reinhardtii
(Trebst et al. 2002). The average content of a-tocopherol
was highest in Rumex dentatus at Site I and lowest in
Atropa acuminata at Site III. Antioxidant activities of Site
I leaves were significantly higher compared to Sites II and
III leaves (P < 0.0001). a-tocopherol content was positively correlated with antioxidant activity (P D 0.0005),

0.60.008f
(0.04)
0.70.013c
(0.05)
0.40.015a
(0.05)
0.20.015d
(0.043)
0.50.007b
(0.065)
Values are means from five samples ( SE). Means within columns not followed by the same letter are different at P < 0.06. Means within a column followed by different letters are different at P < 0.06.

2.47a
(0.19)
2.63a
(0.16)
2.45a
(0.046)
2.24b
(0.019)
2.70a
(0.11)
52.0a
(4.3)
34.9a
(0.98)
38.9b
(1.1)
35.9b
(1.7)
47.3a
(3.2)
15.6a
(2.1)
9.8a
(0.39)
11.3b
(0.46)
11.1b
(0.52)
12.9a
(1.2)
36.4a
(2.4)
25.8a
(0.47)
27.6b
(0.69)
24.8b
(1.2)
34.4a
(2.1)
0.70.001i
(0.076)
0.70.014g
(0.09)
0.40.013e
(0.04)
0.20.015d
(0.02)
0.50.013f
(0.013)
2.39a
(1.02)
2.91a
(0.11)
2.05a
(0.046)
1.90b
(0.013)
2.79a
(0.13)
68.0a
(5.9)
26.9a
(0.62)
45.0b
(1.4)
39.5b
(1.9)
59.3a
(3.6)
20.6a
(2.4)
6.8a
(0.23)
14.7b
(0.46)
13.6b
(0.72)
15.9a
(1.2)
49.4a
(2.9)
19.8a
(0.30)
29.6b
(0.99)
25.9b
(1.56)
44.5a
(2.1)
2.97a 0.640.001g
(1.5)
(0.098)
2.59a
0.80.002k
(0.19)
(0.05)
2.15a
0.50.001i
(0.046)
(0.07)
2.04b
0.20.02l
(0.109)
(0.02)
2.74a
0.60.001i
(0.18)
(0.067)
73.0a
(5.1)
44.9a
(1.02)
48.9b
(1.1)
44.9b
(2.03)
67.4a
(4.6)
19.6a
(4.6)
11.8a
(0.45)
15.9b
(0.46)
14.6b
(0.87)
18.0a
(2.6)
57.9a
(6.9)
30.8a
(0.67)
34.0b
(1.04)
29.6b
(1.78)
49.4a
(3.5)

Chl a:b
Chl a
Chl b
Total Chl
(mg cm2) (mg cm2) (mg cm2)

Site II

Total
carotenoid
(mg g1)
Chl a
Chl b
Total Chl
(mg cm2) (mg cm2) (mg cm2) Chl a:b
Medicinal
herbs

Rumex
dentatus
Atropa
accuminata
Lupinus
polyphyllus
Hyoscyamus
niger
Lavandula
officinalis

Chl a
Chl b
Total Chl
(mg cm2) (mg cm2) (mg cm2)
Total
carotenoid
(mg g
1)

Site III

Chl a:b

Total
carotenoid
(mg g
1)

S. Jan et al.

Site I

Table 2. Variation in levels of pigments in leaves of Rumex dentatus, Atropa accuminata, Lupinus polyphyllus, Hyoscyamus niger, and Lavandula officinalis in the field under different altitude clines.

Downloaded by [The University of Kashmir ], [Sumira Jan] at 00:25 13 October 2014

with leaves of the highest a-tocopherol concentrations

tending to exhibit the highest antioxidant activities; the r2
value for this relationship was 0.32 (Table 4). Irrespective
of site variations and higher average leaf temperatures,
the a-tocopherol contents of Hyoscyamus niger leaves
displayed but small variations. Lupinus polyphyllus exhibited a moderate antioxidant profile under all field conditions. Among the studied herbs, Atropa acuminata
accumulated comparatively larger amount of total ascorbate, exhibiting maximum value of 10.55 nmol mg1 protein h1 in Site I (altitude 4175 m, snow cover 244 cm;
Table 4). Leaves of Rumex dentatus demonstrated different response to increasing altitudes, exhibiting a maximum value of 7.55 nmol mg1 protein h1 in Site III with
an altitude of 410 m and snow cover of 308 cm.
3.3.

Change in antioxidant enzyme kinetics

Regardless of the varied environment, the specific activity

of the anti-oxidative enzymes SOD, CAT, GR and APX
was generally higher in Site III leaves compared to Sites I
and II counterparts (Tables 5). The only exception was
CAT in plants inhabiting the forest understory. The specific activity of this enzyme gradually decreased from Site
I with an altitude of 4175 m to Site III with an altitude of
4100 m. However, over the entire vegetation season, the
average activity of SOD and CAT was consistently greater
in plants growing under the leaf canopy of Site II (with
altitude 3748 m) than in those inhabiting the exposed
dune under Site III (altitude 4100 m). An inverse pattern
appeared for CAT-specific activity, which appeared to be
less active in full sunlight (Site III) than under the reduced
light level in the vegetation shade (Sites I and Site II). The
two-way ANOVA revealed a highly significant effect of
altitude on the variation in CAT and GR enzymatic activity (P < 0.0001), as well as a marginally significant altitude effect on SOD activity (P < 0.0516; Table 5). The
results suggest a plastic adjustment to temporal fluctuation
in external environmental conditions. The statistically significant F-value for the altitude-by-population interaction
indicates that the pattern of plastic responses to altitudinal
changes in the abiotic environmental conditions was
unique for each of these herbs. Further, the effect of altitude observed clearly indicates that the mean specific
activity of the antioxidative enzymes SOD, CAT, GR and
APX in all plants was significantly different between the
population experiencing full sunlight under field conditions of Site III and the population growing under reduced
light intensity in the vegetation shade (Sites I
and II).
Antioxidant activities of herbs were similar in all leaf
samples except that Atropa acuminata from Site II had
lower SOD activity than at Sites III and I. Rumex dentatus
exhibited the highest antioxidant activities (Tables 4
5).
Rumex dentatus had higher activities of GR than other

Israel Journal of Plant Sciences

Table 3. Variation in levels of starch, soluble sugars and total non-carbohydrates in leaves of Rumex dentatus, Atropa accuminata,
Lupinus polyphyllus, Hyoscyamus niger, and Lavandula officinalis in the field under different altitude clines.

Downloaded by [The University of Kashmir ], [Sumira Jan] at 00:25 13 October 2014

Site I

Rumex
dentatus
Atropa
accuminata
Lupinus
polyphyllus
Hyoscyamus
niger
Lavandula
officinalis

Site II

Site III

Starch
(mg g1)

Soluble sugars
(mg g1)

TNC
(mg g1)

Starch
(mg g1)

Soluble sugars
(mg g1)

TNC
(mg g1)

Starch
(mg g1)

Soluble sugars
(mg g1)

TNC
(mg g1)

0.49b
(0.44)
0.118 b
(0.17)
32.45a
(1.23)
23.78bc
(1.89)
39.8a
(2.98)

83.9a
(1.02)
87.90a
(7.6)
45.89a
(3.45)
37.2 ab
(2.03)
67.80b
(4.56)

65.89c
(2.09)
83.89b
(4.3)
50.89a
(5.87)
39.08a
(4.05)
66.98a
(5.67)

0.350b
(0.42)
0.109c
(0.13)
27.0c
(3.8)
19.03c
(1.98)
31.6a
(4.02)

81.4a
(0.79)
67.5a
(2.5)
62.1c
(1.3)
43.7a
(1.78)
59.6b
(2.08)

74.9a
1.0)
71.9a
(3.8)
32.7b
(1.2)
29.8ab
(2.5)
79.0a
(4.9)

0.300b
(0.30)
0.100c
(0.10)
22.03c
(2.03)
19.03c
(1.98)
25.3a
(3.1)

74.6a
(0.8)
71.8a
(3.8)
52.7b
(1.2)
43.7a
(1.78)
45.7b
(1.2)

74.9a
1.0)
71.9a
(3.8)
32.7b
(1.2)
29.8ab
(2.5)
71.0a
(3.6)

Values are means from five samples (SE). Means within columns not followed by the same letter are different at P < 0.06. Means within a column
followed by different letters are different at P < 0.06.

herbs followed by Lavandula officinalis under all different

altitudinal clines. Activities of APX were similar for all
herbs and under different altitudinal clines. However, catalase activity was significantly higher at Site III compared
to the other two sites. The highest catalase activity was
exhibited by Rumex dentatus and Lavandula officinalis,
respectively.
Malanodialdehyde (MDA) and proline content exhibited similar trends. The concentration of TBARS was significantly enhanced in plants under the field conditions of
Site II (Tables 5
6). The maximum increment in MDA
(98.93%) and (88.27%) in proline content was recorded in
Atropa acuminata and Lavandula officinalis in Site I
plants followed by those at Site II (56.79%) MDA and
(69.69%) in Site III plants, respectively. Similarly, there
was a strong positive correlation between proline levels
and altitude: proline accumulation was highest at Site III,
indicating plant exposure to stress.

3.4.

Quantitative analysis of total flavonoids, total

flavonols, alkaloids, phenolic acids, polyphenols,
anthocyanins, and proanthocyanidins

Phenolics, total flavonoids, total flavonols, alkaloids,

polyphenols, anthocyanins and proanthocyanidins of
leaves were also compared in selected herbs. Figures 1
and 2 show the changes in methanol-soluble compounds
extracted from leaves of herbs thriving under different
environment conditions. All herbs accumulated the
maximum level of antioxidants, i.e. phenolic acids, in
Site I. Site III exhibited the lowest levels of metabolites.
The UV protection in Rumex dentatus seems to be relatively high under all altitudinal regimes. Similarly,
Lavandula officinalis and Lupinus polyphyllus leaves
always appeared well-equipped with soluble flavonoids,

compared to Atropa acuminata and Hyoscyamus niger

which accumulated alkaloids and anthocyanins especially under the intense sunlight of Site III. However,
flavonoids and anthocyanins exhibited significant difference among herbs at different sites. These metabolites
accumulated to higher levels, while alkaloids exhibited
a minor response to increasing temperatures. Lupinus
polyphyllus accumulated higher levels of alkaloids especially under Site I conditions. The data obtained confirm
that alkaloid compounds are positively correlated with
altitude of the growing site and have profound and
reproducible effects on the quantitative composition of
profiles of certain secondary metabolites in all herbs as
illustrated in Figure 1.

4. Discussion
Plants from temperate climatic regions, especially northwestern Himalaya, are exposed to extreme variations in
environmental conditions. To cope with the variable climatic factors such as intense light flux and low temperature, these plants undergo various chemical and
physiological changes (Janska et al. 2010; Sanghera et al.
2011). The increase in light intensity under cold conditions enhances the susceptibility of herbs to photo-oxidative damage (Neill & Gould 2003; Hughes et al. 2005).
The main hypothesis guiding the work plan of the present
study was the inverse relation in secondary metabolism
versus primary product output and antioxidant defense
protecting the underlying mesophyll cells by absorbing
blue
green light. The absorbance of blue
green light by
accumulation of secondary metabolites, especially proanthocyanidins and flavonols, seems to result from alterations in chlorophyll ratios observed in Rumex sp. and

9.47 0.13a 10.55 0.24a

1.50 0.17d

Atropa
accuminata

4.55 0.26b

2.45 0.05c

0.9 0.76bc

6.85 0.97b
6.98 0.20c 1.23 0.09a

1.02 0.11b

8.08 0.23b

0.4 0.06a

1.64 0.16a

a-Tocopherol
(mg g1)

4.4 0.13b

2.3 0.31b

3.7 0.3a

1.4 0.14d

3.9 0.20a

3.6 0.20c

5.5 0.11a

3.3 0.15b

7.5 0.09a

1.5 0.44d

Reduced
Oxidised
ascorbic acid ascorbic acid
(n mol mg1 (n mol mg1
protein h1) protein h1)

3.92 0.16a

1.13 0.09a

a-Tocopherol
(mg g1)

5.77 0.64a
6.35 0.54b 0.59 0.48ac

9.76b 0.26

6.98 0.78b 0.18 0.03b

7.55c 0.11

Total
ascorbic acid
(n mol mg1
protein h1)

2.38 0.11c 6.34b 0.36c 0.43 0.01b

4.37 0.09a

0.78 0.6bc

1.98 0.56b

6.78 0.19a

2.32d 0.06

Oxidised
ascorbic acid
(n mol mg1
protein h1)

6.34b 0.07

0.98 0.09a

5.22a 0.14

Reduced
ascorbic acid
(n mol mg1
protein h1)

Site III

6.9 0.98ab 3.36c 0.20

0.29 0.09a

1.36 0.19a

a-Tocopherol
(mg g1)

8.02 0.28c 0.98 0.03b

7.92a 0.30

7.18ab 0.25

8.99a 0.23

5.87c 0.31

Total
ascorbic acid
(n mol mg1
protein h1)

Site II

SOD

APX
2.76a
0.008
2.37d
0.011
2.65b
0.017
2.46c
0.021
2.69b
0.015

GR
2.76a
0.008
2.37d
0.011
2.65b
0.017
2.46c
0.021
2.69b
0.015

CAT
2.84a
0.038
1.70e
0.007
2.48c
0.009
1.93d
0.010
2.74b
0.011

MDA
61.88a
2.22
46.21c
0.90
50.80b
2.69
58.90a
1.29
53.10b
1.54

SOD
2.57a
0.010
1.90e
0.007
2.43c
0.006
2.37d
0.012
2.49b
0.010

APX

2.91a
0.008
2.57e
0.012
2.86b
0.011
2.67d
0.012
2.81c
0.027

GR

Site II

2.91a
0.008
2.57e
0.012
2.86b
0.011
2.67d
0.012
2.81c
0.027

CAT

2.56a
0.016
1.74e
0.007
2.19c
0.007
1.96d
0.014
2.35b
0.019

MDA

61.20a
0.73
48.82c
0.90
53.66b
0.01
59.88a
0.58
55.06b
1.11

SOD

2.04a
0.009
1.74e
0.009
1.90c
0.004
1.82d
0.010
1.94b
0.007

APX

2.86a
0.014
2.44e
0.012
2.80b
0.008
2.58d
0.014
2.67c
0.013

GR

Site III
MDA
2.04a 1.90a
0.009
0.009
1.74e 1.63d
0.009
0.010
1.90c 1.80bc
0.004
0.008
1.82d 1.76c
0.010
0.008
1.94b 1.84b
0.007
0.008

CAT

Values are means from five samples (SE). Means within columns not followed by the same letter are different at P < 0.06. Means within a column followed by different letters are different at P < 0.06.
SOD, superoxide dismutase activity; AP, ascorbate peroxidase activity; GR, glutathione reductase activity; CAT, catalase activity; MDA, lipid peroxidation.

Rumex
63.61a 2.53a
dentatus
0.23
0.009
Atropa
47.51ab 1.87e
accuminata
0.16
0.008
Lupinus
54.07a 2.39c
polyphyllus
0.01
0.010
Hyoscyamus niger
49.57a 2.25d
12.08
0.019
Lavandula
56.20a 2.45b
officinalis
0.02
0.009

Plants

Site I

Table 5. Variation in levels of superoxide dismutase activity (EU mg
1 protein h
1), ascorbate peroxidase activity (mmol mg
1 protein h
1), glutathione reductase activity (m mol
mg
1 protein h
1), catalase activity (mmol mg
1 protein h
1), lipid peroxidation [(MDA) nmol g
1 fw] in leaves of different medicinal herbs in the field under different altitude clines.

Values are means from five samples ( SE). Means within columns not followed by the same letter are different at P < 0.06. Means within a column followed by different letters are different at P < 0.06.

Lavandula
officinalis

Lupinus
3.44 0.20c 4.65 0.16b
polyphyllus
Hyoscyamus 2.98 0.17ab 3.87 1.02a
niger

6.30 0.21c

1.57 0.10d

4.72 0.28a

Rumex
dentatus

Medicinal
herbs

Total
ascorbic acid
(n mol mg1
protein h1)

Oxidised
ascorbic acid
(n mol mg1
protein h1)

Reduced
ascorbic acid
(n mol mg1
protein h1)

Site I

Table 4. Variation in levels of total ascorbate [n mol mg1 protein h1] and alpha tocopherol (mg g1) in leaves of Rumex dentatus, Atropa accuminata, Lupinus polyphyllus,
Hyoscyamus niger, and Lavandula officinalis in the field under different altitude clines.

Downloaded by [The University of Kashmir ], [Sumira Jan] at 00:25 13 October 2014

6
S. Jan et al.

Israel Journal of Plant Sciences

Table 6. Variation in proline content [ng g
1 fw] in leaves of Rumex dentatus, Atropa accuminata, Lupinus polyphyllus, Hyoscyamus
niger, and Lavandula officinalis in the field under different altitude clines.
Selected herbs
Rumex dentatus
Atropa accuminata
Lupinus polyphyllus
Hyoscyamus niger
Lavandula officinalis
LSD at 5%

Site I

Site II

Site III

44.18e 0.015
48.72a 0.018
47.03d 0.019
47.10c 0.012
48.72a 0.018
0.048

45.66e 0.014
58.73c 0.007
58.73c 0.007
61.77b 0.007
65.97a 0.007
0.054

43.87e 0.020
53.89c 0.006
53.89c 0.006
56.87b 0.012
57.69a 0.010
0.043

Downloaded by [The University of Kashmir ], [Sumira Jan] at 00:25 13 October 2014

Values are means from five samples (SE). Means within columns not followed by the same letter are different at P < 0.06. Means within a column followed by different letters are different at P < 0.06.

Lavandula sp. (Table 2). Accumulation of anthocyanins

and a decline in chlorophyll a:b ratio immediately before
snowfall in the alpine herbs was reported in the present
study. These findings are interesting, as the decline in
chlorophyll a:b ratio and the increase in anthocyanin content suggests a shift from carbon fixation to light capture
(Grace & Logan 1996; Demmig-Adams 1998). Photosynthetic pigments are the most susceptible site for low-temperature injury. Proteomic analysis of cold-hardy plants
displayed significant changes in photosynthetic enzymes
(Janmohammadi 2012). Carotenoids aid in protection of
the photosynthetic pigments, and therefore their

accumulation in the cells under low temperatures is

considered to stabilize thylakoid membranes against
photo-oxidative stress (Ivanov et al. 2006; Laugier et al.
2010). The highest accumulation of carotenoids was
observed in Atropa accuminata, reaching about 0.8 mg g
1
FW in leaves. However, alkaloid concentration decreased
with the increase of altitude, suggesting that the change in
carotenoids may be a part of an acclimation mechanism to
a cold season at high altitude.
Huner et al. (1993) demonstrated that carbohydrate
accumulation is involve in plant cold tolerance. Such
accumulation necessitates exclusion of photosynthates

Figure 1. Quantitative analysis of (a) phenolic acids (GAE g
1 of extract), (b) flavonoids and (c) total flavonols [quercetin equivalent
(mg g
1)] in leaves of Rumex dentatus, Atropa accuminata, Lupinus polyphyllus, Hyoscyamus niger, and Lavandula officinalis under
different altitudes. GAE, gallic acid equivalents.

Downloaded by [The University of Kashmir ], [Sumira Jan] at 00:25 13 October 2014

S. Jan et al.

Figure 2. Quantitative analysis of (d) proanthocyanidins, (e) total alkaloids, and (f) anthocyanins (catechin equivalent (mg g
1)] in leaves
of Rumex dentatus, Atropa accuminata, Lupinus polyphyllus, Hyoscyamus niger, and Lavandula officinalis under different altitudes.

from the plastid to the sink organs, entailing that transport

processes are adapted to lower temperatures. High-mountain plants synthesize carbohydrates following snowmelt
during springtime; however, underground organs and
some aerial parts continue to be exposed to cold temperatures due to snow melting (Lutz 1996; Lutz et al. 2005).
In the present study, total non-structural carbohydrate
content did not differ significantly between sites, although
TNCs were twofold higher in Site I leaves compared with
Site III (P < 0.0001; Table 3). Accumulation of carbohydrates is part of an adaptation process of high land plants
to cold stress. Leaf carbohydrate analysis suggested that
the induced change in antioxidants is not aimed at ameliorating source:sink imbalance in high-altitude leaves.
Accumulation of carbohydrates evolved as a new metabolic approach under low-temperature stress (Nagele
et al. 2012). It is recently assumed that under high light
(red) leaves fix more carbon than green leaves; and that
carbon sinks are reduced during the winter due to the termination of growth, inhibition of translocation caused by
soil freezing, and/or reduced metabolic rates. This could
result in accumulation of carbon in alternative sinks (i.e.
the phenylpropanoid pathway), leading to the assembly of
anthocynanins and flavonoids (Hughes et al. 2005). Concurrent to our study, Kontunen et al. (2002) suggested that
low temperature results in increment of soluble sugars

that are produced due to conversion of carbohydrates to

sugars. Cold tolerance of high-land plants has often been
correlated with fructan accumulation under cold acclimation (Livingston et al. 2009; Sui et al. 2012). a-tocopherol, ascorbate (AsA) and dehydroascorbate (dAsA) were
previously identified frost-tolerant tree species (Larsson
1988; Polle & Rennenberg 1994). The adjustment of
plants to the unusual climatic conditions of the studied
regions is apparent by the varied metabolic changes leading to regulation of antioxidative enzymes and the
increase in antioxidants. Dormant needles demonstrated
increased content of both AsA and dAsA, although the
ratio of AsA:dAsA was significantly lower (Wingsle &
Moritz 1997). These findings indicate that AsA metabolism is involved in low-temperature-induced oxidative
stress. In the present study, the highest a-tocopherol content was found in Rumex dentatus in Site I and the lowest
in Atropa acuminata leaves in Site III plants. a-tocopherol, AsA and dAsA of Site I leaves were significantly
higher than Sites II and III leaves (P < 0.0001). The
increase of the lipophilic antioxidant a-tocopherol in
these herbs can explain their protective function (i.e. inactivation of free radicals such as singlet oxygen and
superoxide radicals; Lutz et al. 2005). The hydrophilic
antioxidant ascorbic acid exhibited same variation as
a-tocopherol in our study. Wildi and Lutz (1996)

Downloaded by [The University of Kashmir ], [Sumira Jan] at 00:25 13 October 2014

Israel Journal of Plant Sciences

investigated numerous alpine plants and concluded that
there exists no regular stratagem but species-specific
accretion of diverse antioxidants. Streb et al. (1997) also
illustrated that ascorbate contents in plastids of Homogyne, Soldanella alpina, and Ranunculus glacialis were
relatively higher than lowland species, depicting the possible role of intense irradiation at higher altitudes.
A statistically significant effect of altitude was
observed, indicating that the specific activity of the antioxidative enzymes SOD, CAT, GR, and APX in all plants
was varied between the populations experiencing full sunlight under the conditions of Site III and reduced light
intensity in the shade vegetation (Sites I and II). Significant disparity in antioxidative activity in high mountain
plants was considered to result from antioxidative defense
and biochemical adjustment (Oncel et al. 2004). Obata
et al. (1996) suggested as well that an enzyme may exhibit
a diverse level of resistance to an array of climatic factors
in varied species leading to decline or amplification of
enzymatic activity. Our results for catalase, peroxidase,
and total antioxidant content in the leaves of medicinal
plants from different altitudes of western Himalaya suggests that acclimation of plants to specific local climatic
environments involves regulation of antioxidative
enzymes along with accumulation of protective proteins.
Variation of enzymatic activity is characteristic of plants
exposed to the extreme conditions of high mountains
(Chkhubianishvili et al. 2011). In the herbs studied in the
present project, a decline of the activity of one enzyme
was accompanied by activation of another.
In all herbs the highest levels of antioxidants, phenolic
acid, was found in Site I (up to a twofold increase
compared Sites II and III). Site III exhibited the lowest
metabolites (Figures 1 and 2). The UV protection in
Rumex dentatus seems to be relatively high under all altitudes. Antioxidant enzymes including SOD, APX and
CAT, SOD radicals reduce H2O2 by utilizing diverse substrates such as phenolic compounds, lignin precursors,
auxin, and secondary metabolites (De Gara 2004; Passardi
et al. 2005). In recent years, the protective role of anthocyanins
the water-soluble flavonoid pigment
against
photo-oxidative stress in vegetative tissues has been
demonstrated (Neill & Gould 2003; Grace 2005).
Anthocyanins are also known to reduce ROS production,
enhance ROS scavenging and are thereby involved in
oxidative defense under biotic or abiotic stress (Hatier &
Gould 2008). Several in vitro studies reported that foliar
anthocyanins act in both light scavenging and radical
scavenging (Hatier & Gould 2008; Hughes et al. 2005). In
support to our findings, a positive correlation between the
altitude of the growing site and the contents of flavonoids
and phenolic acids was demonstrated in flowering heads
collected from different altitudes ranging from 180 to
1060 m (C. capillaris), from 190 to 1290 m (H. pilosella),
and from 20 to 1290 m (H. radicata) (Zidorn et al. 2005;

Spitaler et al. 2008). An increase in phenolic compounds

and carotenoids with increasing altitude was suggested to
result increased UV radiation (Korner 1999). Bilger et al.
(2007) confirmed that low temperatures during the growth
period increase rates of phenolic biosynthesis in diverse
plant species even under high levels of UV-B radiation.
Phenolic compounds display an imperative function in the
hydrogen peroxide scavenging system of plants, which
further involves peroxidase, ascorbic acid, and glutathione
(Takahama & Oniki 1997). Although flavonoids and
anthocyanins exhibited significant difference among herbs
at the different sites, alkaloids showed relatively little
response to increasing temperature. However, Lupinus
polyphyllus accumulated higher levels of alkaloids, especially under Site I conditions. A similar response has been
documented before for Lupinus argenteus, exhibiting a
decline in quinolizidine alkaloid with increasing elevation
(Carey & Wink, 1994). Declining levels of alkaloids at
higher altitudes may be due to lower herbivory, as most
alkaloids are induced under intense light and herbivore
damage (Wink 1993).
Acknowledgments
This work is part of the DBT-PDF RA fellowship wide sanction
no.: BT/RA-PDF-5525/PBD/16/567/2012 Dtd. 22/05/2012. The
first author thanks the Director, CORD, University of Kashmir
Srinagar J&K, India for providing the necessary support.

References
Aebi H. 1984. Catalase in vitro. Meth Enzymol. 105:121
126.
Bates L, Waldren PP, Teare JD. 1973. Rapid determination of
free proline of water stress studies. Plant Soil. 39:205
207
Bilger W, Rolland M, Nybakken L. 2007. UV screening in
higher plants induced by low temperature in the absence of
UV-B radiation. Photoch Photobio Sci. 6:190
195
Carey DB, Wink M. 1994. Elevational variation of quinolizidine
alkaloid contents in a lupine (Lupinus argenteus) of the
Rocky Mountains. J Chem Eco. 20:849
857
Chkhubianishvili E, Kacharava N, Badridze G, Chanishvili S,
Kurdadze T. 2011. Activity of peroxidase, catalase and content of total proteins in leaves of some herbaceous plants of
high mountains of the Caucasus. Bull of the Georg Natl
Acad Sci. 5:96
100.
De Gara L. 2004. Class III peroxidases and ascorbate metabolism in plants. Phytochem Rev. 3:195
205.
Demmig-Adams B. 1998. Survey of thermal energy dissipation
and pigment composition in sun and shade leaves. Plant Cell
Physiol. 39:474
482.
Foyer CH, Graham, N. 2003. Redox sensing and signalling associated with reactive oxygen in chloroplasts, peroxisomes
and mitochondria. Physiologia Plantarum. 119(3):355
364.
Grace SC. 2005. Phenolics as antioxidants. In: Smirnoff N, editor. Reactive oxygen species in plants. Oxford: Blackwell
Publishing Ltd; p. 141
68
Grace SC, Logan BA. 1996. Acclimation of foliar antioxidant
systems to growth irradiance in three broad-leaved evergreen species. Plant Physio. 112:1631
1640.
Hatier JHB, Gould KS. 2008. Foliar anthocyanins as modulators
of stress signals. J Theor Biol. 253:625
7.

Downloaded by [The University of Kashmir ], [Sumira Jan] at 00:25 13 October 2014

10

S. Jan et al.

Hacker J, Neuner G. 2006. Photosynthetic capacity and PS II

efficiency of the evergreen alpine cushion plant Saxifraga
paniculata during winter at different altitudes. Arct Antarct
Alp Res. 38:198
205.
Heath RL, Packer L. 1968. Photoperoxidation in isolated chloroplasts. I. Kinetics and stoichiometry of fatty acid peroxidation. Arch Biochem Biophys. 125:189
198.
Hughes NM, Neufeld HS, Burkey KO. 2005. Functional role of
anthocyanins in high-light winter leaves of the evergreen
herb Galax urceolata. New Phytol. 168:575
83.
Huner NPA, Oquist G, Hurry VM, Krol M, Falk S, Griffith M.
1993. Photosynthesis, photoinhibition and low temperature
in cold tolerant plants. Photosynth Res. 37:19
39.
Ivanov AG, Sane PV, Krol M, Gray GR, Balseris A, Savitch LV,
Oquist G, Huner NP. 2006. Acclimation to temperature and
irradiance modulates PSII charge recombination. FEBS
Lett. 580:2797
2802.
Janmohammadi. 2012. Cold acclimation and vegetative/reproductive transition in winter cereals. Albanian J Agr Sci.
11:199
205
Janska A, Marsk P, Zelenkova S, Ovesna J. 2010. Cold stress
and acclimation-What is important for metabolic adjustment. Plant Biol. 12: 395
405.
Kappen L. 1993. Plant activity under snow and ice, with particular reference to lichens. Arctic. 46:297
302.
Kappen L, Breuer M. 1991. Ecological and physiological investigations in continental Antarctic cryptogams. II. Moisture
relations and photosynthesis of lichens near Casey Station,
Wilkes Land. Ant Science 3:273
278.
Kaul MK. 1997. Medicinal plants of Kashmir and Ladakh. New
Delhi: Indus publishing Co.
Kontunen-Soppela S, Lankila J, Lahdesmaki P, Laine K. 2002.
Response of proteins and carbohydrate metabolism of Scots
pine seedlings to low temperature. J Plant Physiol. 159:
175
180.
Korner C. 1999. Alpine plant life. Functional plant ecology of
high mountain ecosystems. 2nd ed. Berlin: Springer.
Kutsch WL, Kappen L. 1997. Aspects of carbon and nitrogen
cycling in soils of the Bornhoved Lake district II. Modelling
the influence of temperature increase on soil respiration and
organic carbon content in arable soils under different managements. Biogeochem 39(2):207
224.
Larcher W, Bauer H. 1981. Ecological significance of resistance
to low temperature. In: Lange OL, Nobel PS, Osmond CB,
Ziegler H, editors. Physiological plant ecology I: responses
to the physical environment. Berlin, Heidelberg: Springer;
p. 403
437.
Larsson RA. 1988. The antioxidants of higher plants. Phytochem. 27:969
978.
Laugier E, Tarrago L, Vieira Dos Santos C, Eymery F, Havaux
M, Rey P. 2010. Arabidopsis thaliana plastidial methionine
sulfoxide reductases B, MSRBs, account for most leaf peptide MSR activity and are essential for growth under environmental constraints through a role in the preservation of
photosystem antennae. Plant J. 61:271
282.
Livingston DP, Hincha DK, Heyer AG. 2009. Fructan and its
relationship to abiotic stress tolerance in plants. Cell Mole
Life Sci. 66:2007
2023
Lutz C. 1996. Avoidance of photoinhibition and examples of
photodestruction in high alpine Eriophorum. J Plant Physiol.
148:120
128.
Lutz C, Schonauer E, Neuner G. 2005. Physiological adaptation
before and after snow melt in green overwintering leaves of
some alpine plants. Phyton. 45:139
156.

Luwe MWF, Takahama U, Heber U. 1993. Role of ascorbate in

detoxifying ozone in the apoplast of spinach (Spinacia oleracea) leaves. Plant Physiol. 101: 969.
Mishra NP, Mishra RK, Singhal GS. 1993. Changes in the activities of anti-oxidant enzymes during exposure of intact wheat
leaves to strong visible light at different temperatures in the
presence of protein synthesis inhibitors. Plant Physiol.
102:903
10.
Mittermeier RA, Gil PR, Hoffman M, Pilgrim J, Brooks T, Mittermeier CG, Lamoreux J, da Fonseca GAB. 2005. Hotspots
revisited: earths biologically richest and most endangered
terrestrial ecoregions. Washington, DC: Conservation
International.
Nagele T, Stutz S, Hormiller II, Heyer AG. 2012. Identification
of a metabolic bottleneck for cold acclimation in Arabidopsis thaliana. Plant J. 72:102
114.
Nakano Y, Asada K. 1981. Hydrogen peroxide is scavenged by
ascorbate-specific peroxidase in spinach chloroplasts. Plant
Cell Physiol. 22:867
880.
Neill SO, Gould KS. 2003. Anthocyanins in leaves: light attenuators or antioxidants. Func Plant Bio. 30:865
873.
Neuner G, Ambach D, Aichner K. 1999. Impact of snow cover
on photoinhibition and winter desiccation in evergreen
Rhododendron ferrugineum leaves during subalpine winter.
Tree Physiol. 19:725
732.
Obata N, Inouc N, Umebayashi M. 1996. J Soil Sci Plant Nutr.
42:363
366.
Oncel I, Jurdakulol E, Kele J et al. 2004. Role of antioxidant
defense system and biochemical adaptation on stress tolerance of high mountain and steppe plants. Acta Oecologia.
26(3):211
218.
Ordonez AAL, Gomez JD, Vattuone MA, Isla MI. 2006. Antioxidant activities of Sechium edule (Jacq.) Swart extracts.
Food Chem. 97:452
458.
Passardi F, Cosio C, Penel C et al. 2005. Peroxidases have more
functions than a Swiss army knife. Plant Cell Rep. 24:255
65.
Polle A, Rennenberg H. 1994. In: Foyer CH. and Mullineaux
PM, editors. Causes of photooxidative stress and amelioration of defense systems in plants. Boca Raton, FL: CRC
Press; p. 199
218.
Porra RJ. 2002. The chequered history of the development and
use of simultaneous equations for the accurate determination
of chlorophylls a and b. Photosynth Res. 73:149
156.
Rosenberg HR. 1992. Chemistry and physiology of the vitamins.
New York; Interscience Publishers; p.452
453.
Sanghera GS, Wani SH, Hussain W, Singh NB. 2011. Engineering cold stress tolerance in crop plants. Curr Genomics.
12:30
43.
Sen Gupta A, Heinen JL, Holaday AS. 1993. Increased resistance to oxidative stress in transgenic plants that overexpress chloroplastic Cu/Zn superoxide dismutase. Proc
Natl Acad Sci USA. 90:1629
33.
Spitaler R, Winkler A, Lins I, Yanar S, Stuppner H, Zidorn C.
2008. Altitudinal variation of phenolic contents in flowering
heads of Arnica montana cv. ARBO: a 3-year comparison. J
Chem Ecol. 34:369
375.
Streb P, Feierabend J, Bligny R.1997. Resistance to photoinhibition of photosystem II and catalase and antioxidative protection in high mountain plants. Plant, Cell and Environ.
20:1030
1040.
Sui X, Meng F, Wang H, Wei Y, Li R, Wang Z, Hu L, Wang S,
Zhang Z. 2012. Molecular cloning, characteristics, low temperature response of raffinose synthase gene in Cucumis
sativa L. J Plant Phys. 169:1883
1891.

Downloaded by [The University of Kashmir ], [Sumira Jan] at 00:25 13 October 2014

Israel Journal of Plant Sciences

Sun JS, Tsuang YW, Chen IJ, Huang WC, Hang YS, Lu FJ.
1998. An ultra-weak chemiluminescence study on oxidative
stress in rabbits following acute thermal injury. Burns.
24:225
231.
Takahama U, Oniki T. 1997. A peroxidase/phenolics/ascorbate
system can scavenge hydrogen peroxide in plant cells. Physiol Plant. 101:845
852.
Tieszen LL, Lewis MC, Miller PC, Mayo J, Chapin III FS, Oechel
W. 1981. An analysis of processes of primary production in
tundra growth forms. In: Bliss LC, Heal OW, Moore IJ, editors. Tundra ecology: a comparative analysis. Cambridge,
UK: Cambridge University Press; p. 285
356.
Trebst A, Depka B, Hollander-Czytko H. 2002. A specific role
for tocopherol and of chemical singlet oxygen quenchers in
the maintenance of photosystem II structure and function in
Chlamydomonas reinhardtii. FEBS Lett. 516(1):156
160.
Volenec JJ, Boyce PJ, Hendershot KL. 1991. Carbohydrate
metabolism in taproots of Medicago sativa L. during

11

winter adaptation and spring regrowth. Plant Physiol.

96:786
793.
Wildi B, Lutz C. 1996. Antioxidant composition of selected high
alpine plant species from different altitudes. Plant Cell
Environ. 19:138
146.
Wingsle G, Moritz T. 1997. Analysis of ascorbate and dehydroascorbate in plant extracts by high-resolution selected
ion monitoring gas chromatography mass spectrometry. J of
Chromatogr A. 782:95
103.
Wink M. 1993. Quinolizidine alkaloids, pp. 197
239, In: Waterman P, editor. Methods in plant biochemistry, alkaloids and
sulphur compounds. Vol. 8, London: Academic Press.
Wolfe K, Wu X, Liu RH. 2003. Antioxidant activity of apple
peels. J Agr Food Chem. 51:609
614.
Zidorn C, Schubert B, Stuppner H. 2005. Altitudinal differences in
the contents of phenolics in flowering heads of three members
of the tribe Lactuceae (Asteraceae) occurring as introduced
species in New Zealand. Biochem Syst Ecol. 33:855
872.