Phytosociological and biological studies on scrozenera alexandrina boiss

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The present study was designed to investigate richness and importance values of Scrozenera alexandrina growing in a number of community types along a transect in El-Hammam region, NW coast, extending for a distance of 20 km long from the sea shore in the north to the inland ridge in the south. Furthermore, the biological effect; as anti-oxidant and reducing agents for raised blood sugar, the secondary products of this plant was attempted. The results revealed that owing to the surface micro relief differences in the soil there was a variations in vegetation pattern along the transect. Scrozenera alexandrina is well represented in five community types, namely; Arthrocnemum macrostachyium, Plantago albicans, Thymelaea hirsuta, Anabasis articulatus and Scrozenera alexandrina, respectively from north to south. The plant grows vigorously in soils of loamy sand texture with moderate salinity and high amounts of organic matter, CaCO3 and fine soil ingredients. The plant, apparently, consider an indicator for rain falls in the study area.

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Key words: Scrozenera alexandrina, phytosociological and Biological studies,

Introduction

The genus Scrozenera encompasses about 160 species and being Ancient Mediterranean by origin, it is widely spread in arid regions of Eurasia and Africa. The first thorough arrangement of the genus Scrozenera was given by DE Candolle (1805). For the European flora, the genus Scrozenera is represented by 28 species (Chater, 1976). This genus is a member of the family Asteraceae which is a known source of numerous classes of bioactive natural products (Evans, 1996). Previous chemical investigations of this genus yielded dihydroisocoumarins, flavonoids, lignans, phenolic acids, sesquiterpene, sesquiterpene lacton, triterpenes and a new class of bibenzyl derivatives (Zidorn et al., 2003 & 2005, Li et al., 2004 and San et al., 2007). The genus name derives from the Spanish "escorza nera" meaning black bark. The reference to snakes in its common names serpent root on viper's grass comes from the Spanish word "scurzo" meaning viper (Stephans, 2003). In addition to utilizing the fresh leaves as an ingredient in green salad, its sub aerial parts considered by some to be a potent tonic and continue to be used in traditional medicine as analgesic, antirheumatic (in the form of plaster), anthelmintic, curing fever, carbuncle, mastitis and treatement of fertility, also its is used as a diabetic diet for its containing inulin and laevulin (Baytop, 1984).

A sesquiterpene glucoside has been isolated from a tissue culture of Scorzonera hispanica, and its structure has been established by mass spectrometry and two dimensional NMR as 6,9-dihydroxy-4,10,14,15-tetradehydroguaian-6,12-olide 9-O- β -D-glucopyranoside (Bryanskii et al., 1992)1 .

A novel tyrolobibenzyl derivative, 1→6-β-d-apiosyl- β -d-glucopyranosyl 4-[2-(4-hydroxyphenyl) ethyl]benzofuran-2-carboxylate (tyrolobibenzyl D) was isolated from Scorzonera humilis L. The biological activities of the new compound and related tyrolobibenzyls and the semi-synthetic peracetyl derivatives of tyrolobibenzyls B were assessed. The results revealed no cytotoxic activity against P388 cells and neither anti-bacterial activity against Bacillus subtilis nor antifungal activity against Candida albicans and Trichophyton mentagrophytes for any of the investigated compounds.(Zidorn et al., 2002)2.

A new guaianolide was isolated from the roots of Scorzonera austriaca. The structure was assigned as 3β, 11α-dihydroxy-4β- methyl-guaia-10 (14)-en-12, 6α-oli- de ( Li et al, 2004)3. Two triterpene fatty esters, 3b-tetradecanoyl moradiol and 3b-dodecanoyl moradiol were isolated from Scorzonera mongolica. Their structures were elucidated as 3b-tetradecanoyloxy-28-hydroxylolean-18-ene and 3b-dodecanoyl 28-hydroxylolean- 18-ene on the basis of IR, MS, 1D NMR and extensive 2DNMR spectroscopic analyses ( Wang et al., 2007)4.

From the subaerial parts of Scorzonera tomentosa L. (Asteraceae), two new dihydroisocoumarins, compounds (3RS)-3,4-dihydro-3-(4-hydroxyphenyl)-8-methoxy-1H-2-benzopyran-1-one, and named (+)-scorzotomentosin and (-)-scorzotomentosin 4'-O-b-glucoside were isolated, together with four known compounds, (+)-hydrangenol, (-)-hydrangenol 4'-O-b-glucoside, (+)-hydramacrophyllol A , and (+)-hydramacrophyllol B ( Sari et al., 2007)5.

In Egypt, the genus Scrozenera is represented by five species, namely; S. pseudolanata Grossheim, S. mollis Bieb, S. schweinfurthii Boiss, S. drarii Tackh and S. alexanderina Boiss. The first four species are very rare in the Egyptian deserts while the last species is very common and endemic to the North-Western Coastal strip of Egypt. Scrozenera alexanderina is a small perennial herbaceous plant with tuberous ediable root. The flowers pale mauve, deep purple at the center emitting a nice scent of vanilla. It appears only on rainy seasons. No ecological, phytochemical and biological studies have been reported for S. alexandrina. Therefore, the present study was designed to investigate richness and importance values of Scrozonera alexandrina growing in a number of community types under different habitat conditions. This was carried out along a transect in El-Hammam region, the NW coast, extending from the sea shore to the inland ridge (Fig.1). This area characterized by long hot summer and short rainy winter where the average annual rainfall ranges between 100 and 150 mm (UNESCO, 1979 and Ayyad & Ghabbour, 1986). Furthermore, the biological activity (as antioxidant and reducing agent for raised blood sugar in diabetes mellitus) of secondary products of this plant was attempted.

Material and methods

Vegetation studies:-

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Five major habitats; 1-transition between the salt marsh and the ridge, 2- transition between the saline depression and the plain, 3- the plain, 4- transition between the non-saline depression and the inland ridge and 5- the inland ridge, were selected along a transect (20 km long) perpendicular to the sea shore to carry out the present investigation. Twenty four stands of an area of 50x50 meters were set up in the different major habitats. In each stand, 25 (2x2 m) randomly distributed quadrates were located according to Braun-Blanquet (1964). Within each quadrate a list of all species (perennials and annuals) was recorded, relative frequency, density and relative cover for perennial species were determined and summed to give the importance values. Nomenclature was done according to Boulous (1995). Community richness is considered as the count of genera in each major habitat.

Phytochemical and Biological studies:-

Extraction.

About 250 g of the air dried powdered of Scorzonera alexandrina (aerial parts and roots) were extracted by aqueous ethanol the combined ethanolic extracts were concentrated under reduced pressure at a temperature not exceeding 35 oC and successively extracted with diethyl ether, chloroform, ethyl acetate, and n-butanol respectively. Each extract was dried over anhydrous sodium sulphate, and solvent in each extract was concentrated.

Total Phenolics determination:

Total phenolics content of the extracts were executed with the Folin-Ciocalteu method (Liu et al., 2002)6. Two hundred microliters of sample and 0.5 mL of Folin-Ciocalteu reagents (Merck) were added to 10 mL of distilled water in a 25 mL flask. After 3 min, 1 mL of saturated Na2CO3 was added, and the volume was made up to 25 mL. The samples were left for 1 h in the dark, and then the absorbance was measured at 725 nm against a blank. A calibration curve with caffeic acid was established, where 25-200 μg of caffeic acid in 100 μL of distilled was used instead of the sample, and the total phenolic content was expressed in milligrams of caffeic acid per gram of dried plant material.

Total Flavonoid determination:

Total flavonoid content was determined by a colorimetric method (Zhisen et al., 1999) 1 mL of phytochemical extract was diluted with 5 mL of distilled water. Then 0.3 mL of a 5% NaNO2 solution was added to the mixture. After 6 min, 0.6 mL of a 10% AlCl3 ·6H2O solution was added. After 5 min, 2 mL of 1 M NaOH was added, and the total was made up to 10 mL with distilled water. The solution was mixed, and the absorbance was measured immediately against the prepared blank at 510 nm with a Hitachi U-2000 spectrophotometer. The flavonoid content was calculated using a quercetin calibration curve. The results were expressed as micrograms of quercetin equivalents (QE) per milligram of extract.

Experimental animals

The hypoglycaemic activity of the total extract of Scorzonera alexandrina was studied in adult male rats weighing from 210 to 240 g. The animals were housed under standard environmental conditions and maintained with free access to water and ad libitum standard laboratory diet.

After an overnight fast, streptozotocin at a dose of 65 mg/kg dissolved in 0.1M fresh cold citrate buffer, pH 4.5, was injected intravenously to rats into the tail vein. Stable hyperglycaemia was confirmed after 18 h of STZ injection. Only rats with post-absorptive blood glucose levels more than 16 mmol/l (300 mg/dl) were used in this study. Normal and diabetic rats were randomly assigned to two groups containing six rats each. One control group received distilled water, a second treated group received the aqueous extract of Scorzonera alexandrina at a dose of 20 mg/kg. For repeated oral administration, rats were treated once daily for two weeks and blood glucose levels were followed during this period. The rats (n = 6 in each group) were treated orally. All experiments were performed in fasted rats. Blood samples from experimental rats were collected from the tail vein. The blood glucose levels were determined by glucose oxidase method

Soil analysis:-

Soil samples associated with Scrozonera alexandrina were collected from the five major habitats at one depth (0-25 cm). Soil texture (gravel, sands and silt) and some chemical properties (EC, pH, CaCO3, organic matter (O.M), Na+, K+, Ca+2, Mg+2 and P+3) were determined according to Allen (1974) and Allen et al. (1989).

Results and Discussion

Soil characteristics:-

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The physical and chemical characteristics of the soil supporting Scrozonera alexandrina at the study area presented in Table (2). The results revealed that there are wide and narrow ranges of variations in some soil characteristics between the different study habitats. Physically, loamy sand is the dominant texture in the all selected sites. The soil associated with Scrozonera plant is soil more consolidated sand in saline and none saline depression as well as in plains and ridges. Chemically it is evident from the data presented in Table (2) that S. alexandrina grows in soils with different properties. It is noteworthy that the plant can grow under more drastic soil conditions, but the data of the present investigation show those of the soil which the plant prefers and grows better. As shown in Table (2), the plant grows vigorously as the electric conductivity increased. The ranges of E.C. were 14.7, 34.4, 26.8, 29 and 40.8 Ds/Cm-1 in SI, SII, SIII, SIV and SV, respectively. The soil reaction (pH) was approximately the same in all sites where it ranged from neutral (7.02) to very slightly alkaline (7.49). Calcium was the dominant cation followed by sodium, magnesium and potassium with few exceptions in some sites. Phosphorus was the least one among all analyzed cations. It is more interesting here to note that Site V had the highest values of silt (24.25 %), total cations (402.42 me/L), organic matter (0.45 %) and E.C. (40.8 Ds/Cm-1) followed by site II and site IV in every components with few exceptions. The results of the present work coincided with those of Abdel-Razik et al. (1984). Ahmed (1982) and (EEAA and UNEP, 1993) demonstrated that the consolidated sandy soils having high amounts of silt and CaCO3 are quite hospitable to small herbs such as S. alexandrina, and other desert shrubs.

Vegetation studies:-

Phytosociolgically, the number of perennials recorded in the selected sites was forty four species, of which nineteen species were common in all sites. Five species only dominate or share dominance in the different studied microhabitats, Table (3). It is apparent that site IV had the smallest number of genera (20 genus), while the largest number (29 genus) was recorded in site II followed by site I (26 genus) and site III&V (22 genus for each). It is evident from the results of the present investigation that S. alexandrina associated with other communities such as Arthrocnemum macrostachyium, Plantago albicans, Thymelaea hirsuta and Anabasis articulata. It is noteworthy that these communities inhabiting a number of microhabitats of different soil conditions. These differences not only restricted the distribution of the studied species, but also affect its prosperity, growth rate and density. Generally, it is found as shown in Table (2), that S. alexandrina grows in soils of E.C. ranged from 14.7 to 40.8 Ds/Cm-1, organic matter fluctuated within narrow ranges (0.27-0.45 %), CaCO3 ranged between 0.98 and 5.4 % and in soil of considerable amounts of silt (13.14-24.25 %). Also, the soil supporting the plant was distinctly slight alkaline with pH values ranging from 7.02 to 7.49. Regarding its distribution in the study area, the importance value of S. alexandrina increased from north to south until it reached the highest value (IV=81.3) in the inland part compared with all other species along the transect. The IVs of the plant recorded the values 14.17, 30.47, 27.03, 29.72 and 81.30 in SI, SII, SIII, SIV and SV, respectively, Table (3). More-over, the results clearly indicated that S. alexandrina constitute the community type of SV, while it ranked 3rd in SIV, 4th in SII, 6th in SIII and 10th in SI, respectively. The increment or decrement of the IV values of S. alexandrina may be due to the variations in soil characteristics of the study area. In this respect results of the present work were in agreement with the findings of many authors where they concluded that soil properties (topography, texture, moisture content and mineralogical composition), besides climatic conditions, play an important role in vegetation distribution. Ahmed (1982) demonstrated that S. alexandrina grows as an accidental species in the north western coastal zone of Egypt in barley fields and uncultivated desert areas, particularly rocky ridges. He also, mentioned that the horizontal variation that has been shown by natural vegetation is naturally a reflection in both physical and chemical properties of the soil. Moussa (2001), Zaghloul & Moussa (2005) and Moussa & Farida (2007) concluded that the variation in soil properties affect the floristic composition and species distribution in certain area. Ebad et al. (2006) investigated that altitudinal gradients affect some physiological and ecological aspects of Solanum incanum plant. The most interesting point here that S. alexandrina is eating by local people as a fertility and potent agent for men. The appearance of this plant is a good example indicating the heavy rainfall season. The plant species are considered better indicators of zonation than physical factors (Lewis, 1961). Girgis (1971) recognized nine types of plant indicators: indicators of soil, habitat conditions, salinity, land forms, geological formations, cultivation, ground water, grazing and frequency of dew.

Table ( ) Total Phenolics and total flavonoidof of S. Alexandrina extract.

Plant Name

Total

extract

Diethyl ether

extract

Chloroform

extract

Ethyl

acetate extract

Butanol

extract

S. Alexandrina

Total Phenolics

9.2+1.2

4.3+0.33

4.8+0.23

10.1+1.2

6.9+0.16

Total flavonoids

87+3.4

43+2.3

47+3.1

89+4.1

81+3.33

Results of Total Phenolics expressed as milligrams of caffeic acid per gram of dried plant material.

Results of Total flavonoids expressed as micrograms of querecetin equivalents (QE) per mg of extract.

Effect of prolonged administration:

Table 1: Effect of prolonged oral administration of ethanolic extract of Scorzonera alexandrina for 6 successive weeks on the serum activity of ALT and AST and serum levels of total bilirubin, total protein, albumin and glucose in rats, (n=6).

Groups

ALT (UL-1)

AST (UL-1)

T. bilirubin (mg dL-1)

T. protein (g dL-1)

Albumin (g dL-1)

Glucose (mg dL-1)

Control

66.50±3.56

139.38±6.44

1.51±0.10

8.0±0.34

3.5±0.17

115.83±5.84

Scorzonera alexandrina

61.70±3.82

134.27±6.80

1.53±0.11

8.5±0.38

3.5±0.11

88.63±4.75**

Table 2: Effect of prolonged oral administration of ethanolic extract of Scorzonera alexandrina for 6 successive weeks on the serum levels of urea and creatinine in rats, (n=6).

Groups

Urea (mg dL-1)

Creatinine (mg dL-1)

Control

37.7±1.12

0.35±0.02

Scorzonera alexandrina

38.4±1,47

0.38±0.01

Hepatoprotective activity:

Table 3: Effect of the ethanolic extract of Scorzonera alexandrina on the serum activity of ALT and AST and serum levels of total bilirubin, total protein, albumin and glucose in rats with CCl4 induced - hepatotoxicity.

Groups

ALT (UL-1)

AST

(UL-1)

T. bilirubin (mg dL-1)

T. protein (g dL-1)

Albumin (g dL-1)

Glucose (mg dL-1)

Control

360.7±11.7

449.2±13.9

3.8±0.12

5.5±0.21

2.0±0.13

241.5±9.22

Scorzonera alexandrina

372.5±11.9

434.6±13.4

3.3±0.11

5.7±0.19

2.1±0.14

221,7±9.06

Conclusion

In Egypt, S. alexandrina plant grows on consolidated soils, particularly in none-saline depression, in the inland plateau at the Mediterranean coastal region. The soils supporting the plant have loamy sand texture in the all studies sites. S. alexandrina grows vigorously in sites with moderate salinity and higher amounts of CaCO3 organic matter (O.M.) and fine soil fractions (silt).

S. alexandrina is well represented in five community types with different importance values ranged from 14.17 to 81.3. The increment or decrement in the importance values of the plant could be attributed to the variations in soil characteristics. The plant is associated to 44 perennial species, in the study area, besides 20 annual species. The associations, both annuals and perennials, vary in number according to micro-relives in habitat conditions along the transect from the sea shore to the inland plateau. It is evident clearly from the present investigation that S. alexandrina is active only during the rainy seasons, accordingly, the plant is consider as a good indicator for heavy rains that increase the soil moisture content. From economic point of view, planting and propagation of S. alexandrina for medicinal purposes must be taken in consideration through exploiting the newly reclaimed lands at the Egyptian country.