Variability of Stomatal Index and Chlorophyll Content in four

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Variability of Stomatal Index and Chlorophyll Content in four

Transcript Of Variability of Stomatal Index and Chlorophyll Content in four

International Research Journal of Biological Sciences ___________________________________ ISSN 2278-3202

Vol. 4(2), 16-20, February (2015)

Int. Res. J. Biological Sci.

Variability of Stomatal Index and Chlorophyll Content in four species of
Solanaceae Members
Ajayan K.V.1, Babu R.L.2 and Patil Bayakka B1 1Department of P.G Studies and Research in Botany, Karnataka State Women’s University, Bijapur, 586108, Karnataka, INDIA 2Department of P.G Studies and Research in Bioinformatics and Biotechnology, Karnataka State Women’s University, Bijapur, 586108,
Karnataka, INDIA

Abstract

Available online at: www.isca.in, www.isca.me Received 30th September 2014, revised 14th November 2014, accepted 18th January 2015

The present investigation focuses on stomatal index and chlorophyll contents of species like Datura innoxia L., Capsicum annum L., Lycopersicon esculentum M., Solanum nigrum L. and their comparative account on taxonomic similarity. There were no relation between stomatal size and growth habitat. Our study revealed that Lycopersicon significantly showing reduced in stomatal size and more in number at the lower epidermis, in addition to size of the stomata and subsidiary cells of these species were closely allied. All the investigated species has shown anomocytic stomata. The anatomical structure of subsidiary cells shows irregular in shape, size, and numbers. Irrespective of leaf surface they have peculiar types of anomocytic stomata, stomatal type, and stomatal index. Quantification study of chlorophyll pigment revealed that there is no significant difference among the four species in terms of quantity and taxonomical variations in these species.

Keyword: Stomata, chlorophyll, Solanaceae.
Introduction
The present work was carried in four members of solanaceae family, in order to know the taxonomic similarities in terms of stomatal index, types and quantity of chlorophyll in the selected candidate species. The solanaceae family belongs to herbs. Interestingly most of the members of solanaceae are herbs while some species are climbing habit. The family solanaceae consists of about 90 genera and nearly 3000 species1,2. The morphological and leaf epidermal features has been found to be key element in the taxonomical studies. One of the excellent review studies highlights the application of morphological features in plant systematic studies2-4. The introduced the term stomatal index to express stomatal frequency independent to the size of the epidermal cells4. He points out that the stomatal index is a useful taxonomic character in leaf area studies5. The taxa collected from different localities showed more or less constant stomatal index values and can be used in distinguishing different taxa. Systematic studies on the development and morphology of stomatal types may be expected to give evidence for various evolutionary trends among the angiosperm families and may aid in assigning taxa of uncertain to proper positions6. Different taxonomic groups differ in the type of stomata, their distribution and frequency, thus analysis of stomatal characters gives the taxonomic as well as phylogenetic significance.
The plant morphologists, physiologists and taxonomist have long been studied the stomatogenesis. The morphology and ontogenies of taxa are important in intrageneric systematics. Diversity in stomata types, even on the same surface of an organ, indicates the limitation for using stomata as a taxonomic

character7. In addition to diversity, the most repetitively stomata type can also be used as a taxonomic character for the morphological and phylogenetic studies8. Therefore the present work has been undertaken, to elucidate the diagnostic significance of stomata. Many groups highlighted the importance of folio epidermal characters are important in systematic botany, the use of modern techniques and chemical composition4, 9.
Material and Methods
The four candidate species with four different genera were collected at Karnataka State Women’s University Campus Bijapur (Torvi). The selected members were Solanum nigrum. L .Datura innoxia L, Lycopersicon esculentum. M and Capsicum annum. L. The fresh leaves are collected and used immediately for the study. The collected plants were healthy and have no any physiological deforms on leaves. All candidate species were collected from open spaces.
Macroscopic and Surface Study: The collected leaves are washed thoroughly with water and both upper and lower epidermises were peeled off by freehand and the 1mm square section of peeled leaves taken for the microscopic examination. Stomatal Index and number of stomata of the both surface of leaves were carefully observed and recorded for further analysis as per API standard.
Determination of Stomatal Density and Stomatal Index: The stomatal density was determined as the number of stomata per square millimeter of leaf. The index was determined as the

International Science Congress Association

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International Research Journal of Biological Sciences _____________________________________________ISSN 2278-3202

Vol. 4(2), 16-20, February (2015)

Int. Res. J. Biological Sci.

number of stomata per square millimeter divided by the number of stomata plus number of epidermal cells per square millimeter multiplied by 100. The lengths of stomata were measured to determine the stomatal size. The stomatal index (SI) was intended using the equation described by Salisbury, that is:
Where, S = donates the number of stomata per unit area and E = the number of epidermal cells in the same unit area.
Chlorophyll quantification: Collected fresh healthy leaves of selected four genera of solanaceae members and washed thoroughly with running tap water then allowed to dry it at room temperature. Taken 5gm of each leaf sample in mortar and homogenised it with 90% acetone by using pestle. The homogenised extracts were subjected for centrifugation at 5000 rpm at 15 minutes at room temperature, then filtered by whatsman’s filter paper and collected the supernatant in clean test tubes and mixed with equal amount of supernatant and 90% acetone then measured the absorbance of solutions by using spectrophotometer (shimadzu) at 663nm and 645nm wave lengths and 90% acetone taken as standard blank solution. The calculation was done as per Arnon’s method.
Results and Discussion
Microscopic studies of the stomatal density of adaxial surface leaf shows that the maximum numbers of stomata were found in L.esculentum 172 mm2 and minimum stomatal density recorded in 19 mm2. The size of stomata are very minute, because of the minute size density of stomata occurrence and their distribution more on Lycopersicon in other hand the C. annum has sparsely distribution and their density less at adaxial surface as compared with abaxial side (table-1). Another interesting taxonomic feature out of four species only L. esculentum has more density of stomata on adaxial surface as compared to abaxial surface. Many plant systematics and morphologists were revealed that the distribution and frequency of stomata are useful in solving several problems of plant systematic11-13. He was the first to report that the frequency of stomata is high when the size of epidermal cells is low and the frequency is low when the epidermal cells are large14.
Abaxial surface of leaf shown an interesting features of stomatal distribution in C.annum it has three times more than adaxial surface but in case of other three species comparatively less even though Lycopersicon showing less number of stomatal density 86.81 mm2 table-1. Similar kinds of observation noticed15, the quantity of stomata per square millimetre and stomatal index were found to be useful in deciphering the individual species, among the genus Cinnamomum.
The type of stomata is also of considerable significance in delimiting the taxa, both at the species and generic levels. In L.

esculentum, C. annum and S.nigram have shown typical anomocytic stomata and D. innoxia has both anomocytic and anisotypic stomata on their upper and lower surface. The most of species of solanaceae shown anomocytic and anisocytic types of stomata it indicates that variability of stomatal types in family and generic and species level. Similar work was conducted16,17. In some cases, the dominant type of stomata is anomocytic or anisocytic in the leaves of Wedelia urticifolia DC and Wedelia trilobata (L.)AS.Hith. In addition to these two types, polocytic and hemiparacytic stomata are observed in Wedelia chinensis (Osbeck) Merr.The different species of Spilanthes jacq possess diacytic stomata as the most frequent type.

Table-1

Stomatal index of selected species of solanaceae members

S. Plant species No

Adaxial surface 10X

No of

Stomatal

stomata

index (%)

1 Datura innoxia, L.

67.00

25%

2 Lycopersicon esculentum M.

172.00

19%

3 Capsicum annum L.

19.00

13%

4 Solanum nigrum L.

30.00

21%

Abaxial surface

1 Datura innoxia, L.

71.00

25%

2 Lycopersicon esculentum M.

87.00

55%

3 Capsicum annum L.

59.00

16%

4 Solanum nigrum L.

46.00

20%

Stomatal Index: The comparative microscopic study of stomatal index in four species of solanaceae emphasised that the least stomatal index (13%) of C.annum at abaxial surface Interestingly, the highest stomatal index D. innoxia shown at adaxial surface has 25% and same index were at abaxial side also it indicates that there is no variability in both surfaces; on the other abaxial surface L.esculentum has maximum stomatal index (55%) but has least stomatal index of C. annum (16 %) in table-1. The comparative microscopic study of stomatal index in four species of solanaceae emphasised that the least stomatal index (13%) of C.annum at abaxial surface Interestingly, the highest stomatal index D.innoxia shown at adaxial surface has 25% and same index were at abaxial side also it indicates that there is no variability in both surfaces; on the other abaxial surface L. esculentum has maximum stomatal index (55%) but has least stomatal index of C.annum (16 %) shown in table-1. The stomatal index of the individual species within a genus is different, except in few cases, which makes it a useful diagnostic feature. These studies revealed that stomatal index of various genus of the same family significantly varied. The presence or absence of stomata on the epidermis of leaves is extremely useful in delineating taxa both at the species and generic levels. Other studies on stomata revealed the application of the character at various taxonomic levels18,19. The term stomatal index was first introduced to express stomatal

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International Research Journal of Biological Sciences _____________________________________________ISSN 2278-3202

Vol. 4(2), 16-20, February (2015)

Int. Res. J. Biological Sci.

occurrence independent of the size of epidermal cells10,14. He
points out that the stomatal index is a useful taxonomic character when comparable leaf areas are used5. Stomatal index is one of the extensively used features for pharmacognostic
studies. Similar kinds of work was done in the of Cassia genus
made very careful tests of stornatal index on the genus Cassia,
Linn and inferred that stomatal index is the same for different varieties within a species20,21. In this studies of stomatal indexes
were varied in adaxial and abaxial sides of same leaf. So, the

classification of taxa based on stomatal index is deceiving actual classification of plants.
Micrometric analysis table-2 of stomata complex was presented as paired group correlation (Coph correlation 0.7459) dendrogram (cluster analysis) shown that C. annum and S. nigram + 0.64 similarity with paired group ; only + 0.32 similarity with L. esculentum and other genera D. innoxia has 0.16 least similarities among themselves figure-1.

Table-2

Micrometery of stomatal complexes (at 10x field) of selected species

Sl. Species No

Length of subsidiary cells(µm)

Adaxial surface 10X

Breath of subsidiary Length of guard

cells(µm

cell(µm)

1

Datura innoxia, L.

40.04±10.37

30.94±12.21

35.13±28.04

2

Lycopersicon

esculentum M.

43.68±9.96

43.68±14.95

9.1±0.00

3

Capsicum annum L.

41.86±4.98

58.24±16.53

18.2±0.00

4

Solanum nigrum L.

45.5±11.14

43.68±9.96

30.94±4.98

Abaxial surface 10X

1

Datura innoxia, L.

34.58±7.61

20.02±4.07

25.48±4.07

2

Lycopersicon

esculentum M.

63.70±11.15

21.84±8.14

18.20±0.00

3

Capsicum annum L.

23.66±18.56

20.57±6.06

45.5±33.44

4

Solanum nigrum L.

38.22±7.61

34.58±18.65

36.40±0.00

Breadth of guard cells(µm) 18.2±0.00
18.2±0.00
25.48±4.07 20.02±4.07
36.40±6.43
10.92±4.07
10.74±1.00 10.92±4.07

Figure-1 Paired group correlation dendrogram of subsidiary cells and guard cell the lower epidermis of selected species

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International Research Journal of Biological Sciences _____________________________________________ISSN 2278-3202

Vol. 4(2), 16-20, February (2015)

Int. Res. J. Biological Sci.

Taxonomy Related To Chlorophyll Pigments : Chlorophyll are the principal component of pigment protein complexes which take part in a most important role in the photosynthesis, any variation in chlorophyll content is normal to bring change in photosynthetic rate. In this investigation, it found that higher total chlorophyll content in D.innoxia (8.01mg/l) and least number of total chlorophyll content in C. annum. (7.20mg/l) table-3.The diversity of chlorophylls such as chl-a, chl-b etc and total chlorophyll contents does not give any valid information for classification of higher plant because these pigments are commonly distributed all the vascular and non-vascular plant with slightly varied its concentration with species to species. The chlorophyll-a, chlorophyll-b and total chlorophyll contents of four species had shown comparatively slight differences among themselves. Even though, chlorophyll content angiosperms did not give any valid information classification of taxa. Comparative account of chlorophyll and other accessory pigments are principle role in classification of algae because of their cell contain higher concentration other than the principle pigments such as chlorophyll. If the plants growing in shade, it contain less chlorophyll a and more chlorophyll b. Replacing Mg with Fe ions gives a grey-brown chlorophyll product and the presence of Zn and Cu ions increases the stability of the natural green colour22. In this regards the chlorophyll is a principle pigments in all autotrophic green plants hence taxonomic classification of green plants on the bases of chlorophyll along impossible but its concentration will vary species to species. Even though, plant pigments are mainly used for classification lower groups of plants particularly algae.

Table-3

Estimated chlorophyll contents of selected species,

solanaceae members

S. Species No

Chl-

Chl-

a(mg/l) b(mg/l)

Total chl (mg/l)

1

Datura innoxia L.

2.70

0.59

8.01

2

Lycopersicon

esculentum M.

2.78

0.31

7.27

3

Capsicum annum L. 2.79

0.43

7.70

4

Solanum nigrum L. 2.84

0.23

7.20

Conclusion
All investigated four species of solanaceae shown a massive variation in the epidermal cells character at different level of taxonomy. Leaf epidermal anatomy with reference to stomata type, density, stomatal indexes and chlorophyll contents very constructive way for taxonomic point. Among four species D.innoxia shown higher variability in all aspects except the stomatal type; it show a typical anomocytic stomata. . Further molecular level study enhances understanding of stomatal properties. This study may provide a platform for future researcher to make out a distinguishing feature among the four species.

Acknowledgment
Author wish to express sincere gratitude to Karnataka State Women’s University, Bijapur for providing lab facility and also acknowledge to Prof. K.P Sreenath Plant Taxonomist, Bangalore University, for identification of candidate species and Prof. Srinath Department of Botany Gulbarga University .We extend our sincere thanks to post graduate student Miss Priyanka P. Koti, Miss. Anupama S. Nayak for collective support to complete this research work.
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SpeciesStomataChlorophyllTaxaSignificance