Pharmacognostical, Phytochemical and In Vitro antiglycation studies on the leaf extracts of Hibiscus micranthus Linn.


Vani M1*, Padma Latha K2, Amukta Malyada A1, Harathi P1, Anusha M1, Beulah J1, Soujanya R1, Naga Bindu P1

1Department of Pharmacognosy and Phyto Chemistry, Vijaya Institute of  Pharmaceutical Sciences for Women, Enikepadu, Viayawada, Pin: 521108, Krishna District, Andhra Pradesh, India.

2Department of Pharmacology, Vijaya Institute of  Pharmaceutical Sciences for Women, Enikepadu, Viayawada, Pin: 521108, Krishna District, Andhra Pradesh, India.

*Corresponding Author E-mail:  vanimamillapalli@yahoo.co.in

 

ABSTRACT:

The aim of the present study was to carry out powder analysis, fluorescence analysis, microscopic study, physico chemical analysis, total flavonoid and phenolic content and in vitro antiglycation studies for the leaf aqueous and alcoholic extracts of Hibiscus micranthus Linn. by serum albumin assay. The results of powder analysis indicated that the powder with ferric chloride solution showed mustard green colour, under UV powder with ferric chloride showed brown colour.The total ash value 75%w/w, acid insoluble ash 25%w/w, water soluble ash 35%w/w and sulphated ash 15%w/w respectively. The water soluble extractive value 2.2%w/w, alcohol soluble extractive 19.3%w/w and  the result of loss on drying were found to be 9.33%w/w.The microscopic study observations were presence of stellate 3-4 armed trichomes, anisocytic stomata, mucilaginous parenchyma, lignified libiform xylem vessels and Ca.oxalate druse crystals. The leaf constant values include stomatal index of upper and lower epidermis to be 4.66, 4.15, palisade ratio 9, vein islet number 12, veinlet termination number to be 9.The length and width of xylem vessels to be 15.75µ, 117.62 µ. The length and width of trichomes was found to be 30.6 µ and 2.87 µ. The results of preliminary phytochemical screening showed the presence of alkaloids, tannins, phenolics, flavonoids, steroids, cardenolides, coumarin glycosides, reducing sugars, acidic compounds, inulin, waxes and reserpine.The total flavonoid content was more in methanolic extract 104±0.36 mg/g equivalents of rutin. The phenolic content was also more in methanolic extract 52.76±0.67 mg/g equivalents of rutin. The methanolic extract exhibited significant in vitro antiglycation activity at 1000 µg/ml  95.6%±0.0025 ** equivalent to standard drug rutin. Therefore the individual phytoconstituents from the extracrs can be further explored and studied for the activity.

 

KEYWORDS: Hibiscus micranthus, physico chemical, flavonoid content, antiglycation.

 

 


1. INTRODUCTION:

The herbal remedies have been the basis for the discovery of new and novel drug molecules. Glycation is a normal ageing process which results in the formation of advanced glycation end products i.e. AGE’s. The AGE’s accumulate in more amount in the body, bring about change in the structure and function of body proteins which is nothing but ageing and lead to various pathological disorders like diabetes, cataract, retinopathy, neuropathy, atherosclerosis, nephropathy, wounds, obesity, coronary heart disease, inflammatory diseases like arthritis, alzheimer’s, cancer, gastrointestinal disorders etc. Hence, search for new antiglycating drugs from natural sources has assumed greater importance as herbal drugs are absorbed well, cost effective as well as confer least side effects. In order to find new potential antiglycating drugs the plant selected was Hibiscus micranthus Linn. belonging to the family Malvaceae as there were no scientific reports published on the leaves of the plant for antiglycating activity. Traditionally the plant is considered a valuable febrifuge, fruits and flowers as hypoglycemic agents in India, Ceylon, Saudi Arabia and tropical Africa. In India a root paste is applied to the skull to cure head ache. In Tanzania the leaves are eaten as a vegetable and the twigs are used, either peeled or unpeeled, as chewing sticks. The plant was studied for anabolizing activity 1. The ethanolic extract of aerial parts and roots showed significant antifungal, antibacterial activity and anti cancer activity2 . The ethanolic extract of aerial parts and roots showed significant haemopoeitic, antipyretic and antiinflammatory activity 3. The plant was studied for female antifertility activity and viralizing activity4. The phytochemical analysis of various plant parts of Hibiscus micranthus Linn. are as followed. Petroleum ether and benzene extracts of leaves yielded long chain alkanes, alcohols, an acid, a ketone and β-sitosterol. Pentacosane, a ketone non adecanone, docosyl alcohol, tricosyl alcohol, pentacosyl alcohol and myristic acid2.Petroleum ether and benzene extracts of stem yielded long chain alkanes, alcohols, an acid, a ketone and β-sitosterol, tetracosane, octacosane, eicosyl alcohol, tetracosanol2. A simple, precise and accurate HPTLC method development and validation for the determination of rutin in the flavonoidal fraction of stem extracts of Hibiscus micranthus Linn. The rutin content was found to be 9.93%5. Phytochemical screening of the extracts showed the presence of sterols, flavonoids, carbohydrates, phenols and tannins. Chemical profiling for the hydro alcoholic extract was performed by GC-MS analysis. HPLC quantification method has been developed for the quantification of rutin 5.Ether extract from the aerial parts yielded phenolic acids vanillic acid, proto catechuic acid, naringenin, kaempferol and quercetin2. n-hexane extracts of leaf, stem and root analyzed by GC-MS, 23 compounds were analysed 6. The seed contains 15.2% oil (dry weight basis). Fatty acids present in the seed oil include: palmitic acid 18.6%, stearic acid 3.5%, oleic acid 10.1% and linoleic acid 59.8%. The oil also contains malvalic acid (1.7%) and sterculic acid (3.1%), which are cyclo propenoid fatty acids known to cause physiological disorders in animals7. The study focuses on Pharmacognostical, physico chemical and phytochemical studies along with in vitro antiglycation activity on the aqueous and methanolic extracts of leaves of Hibiscus micranthus Linn.

2. MATERIALS AND METHODS:

The Plant material leaves of Hibiscus micranthus Linn. were collected in the month of January during afternoon from the grounds of Vijaya institute of Pharmaceutical Sciences for Women, Enikepadu, Vijayawada. Herbarium was prepared and the sample was authenticated by Dr. D.T. P. Satyanarayana Raju, Plant taxonomist, department of Botany and Microbiology, Acharya Nagarjuna University, Guntur. The photographs of the plant and leaves and powder were depicted in fig 1,2 and 3.

 

 

 

Fig 1: Hibiscus micranthus tree

Fig 2: Hibiscus micranthus  flower

 

                                   

  


 

 

 

 

 

 

Fig 3: Fresh leaves, dried leaves and leaf powder of Hibiscus micranthus

 

 


2.1 PHARMACOGNOSTIC STUDY

2.1.1Powder analysis by nacked eye and  fluorescence under UV

The powder analysis was carried out by observing with nacked eye and observed for  fluorescence under UV8.

 

Fig 4: Powder of Hibiscus micranthus showing different colours when treated with chemical reagents

The results were noted after treating with different chemical reagents in table no1, 2 and the photographs were given in fig 4 

 

2.1.2 PHYSICOCHEMICAL ANALYSIS OF THE LEAF POWDER

2.1.2.1 Determination of Total ash , acid insoluble ash, water soluble ash, sulphated  ash 9,10,11. The results and photographs were given in fig 5and table 3.

 

 

 

 

 

Fig 5: Determination of Total ash and sulphated  ash

 

 


2.1.2.2 Determination of Alcohol soluble and water soluble extractive 9,10,11 . The results and photographs were given in fig 6 and table 3.

 

 

Fig 6: Alcohol and water soluble extractive

 

2.1.2.3 Determination of moisture content (Loss on drying) 9,10,11

Gravimetric method: The results were given in fig 7 and table 3.

 

Fig 7: Determination of loss on drying

 

2.1.3 MICROSCOPY OF LEAF AND LEAF POWDER11,12,13,14

2.1.3.1 Transverse section of leaf, Surface preparation, Determination of leaf constants namely Stomatal number, Stomatal index, Palisade ratio, Vein-islet and veinlet termination number. Observation of powder characteristics, determination of length and width of trichomes and xylem fibres9,10,11.

The results are presented in fig  13, 14, 8, table 4, 15 and table 5.


 

Fig 8(a):Determination of stomatal number and stomatal index lower epidermis

 

Fig 8(b):Determination of stomatal number and stomatal index of  upper epidermis


 

Fig 8(c):Determination of  upper palisade ratio


 


 

Fig 8(d):Determination of vein islet and veinlet termination

 


 

2.2 PHYTOCHEMICAL STUDY 9,10,11

2.2.1 QUALITATIVE SCREENING 9,10,11,15 The aqueous and methanolic extracts of the leaves of Hibiscus micranthus were screened for the phytochemical constituents according to the standard methods as given in Khandelwal. The results were given in table 6 .

 

2.2.2  QUANTITATIVE DETERMINATION

The aqueous and methanolic extracts of the leaves of Hibiscus micranthus were screened for the total content of following phytochemical constituents according to the standard methods as follows.

 

2.2.2 (a) Flavonoids:16

Materials: Aqueous extractof the leaves ofHibiscus micranthus, methanolic extractof the leaves of Hibiscus micranthus, 2% methanolic AlCl3(Finar),rutin (Sigma-Aldrich), Centrifuge (Lab India), incubator (Bio-tech).

 

Preparation of Reagents: 2 ml of methanolic AlCl3 solution was dissolved in 100 ml of water.

 

Method: The extract (1.5 mL) was added to 1.5 ml of 2% methanolic AlCl3 solution. The mixture was vigorously shaken on Centrifuge for 5 minutes at 200 rpm and the absorbance was read at 367 nm after 10 minutes of incubation. Rutin was used as a standard for the calibration curve. The assay was carried out in triplicate (Saper B R and Kim et al 2010).The results were calculated by the given formula and were given in table 7 and the pictures were given in fig 9.

C = c.V/m

C – Total phenolic compounds mg/g of plant extract

       c –  The concentration of standard established from the calibration curve mg/ml

V – The volume of extract in ml

  m -The weight of pure plant extract

 

Fig 9: Determination of total flavonoids of  Hibiscus micrantus leaf extracts at various concentations

 

2.2.2(b) Phenolics:17

Materials: Aqueous extract of the leaves of Hibiscus micranthus, methanolic extract of the leaves of Hibiscus micranthus, 0.2 N FolinCiocalteu phenol reagent (Merck), 2% Sodium carbonate (Finar), rutin (Sigma-Aldrich).

 

Method: This was determined using Folin-Ciocalteu method. The extract (0.5 ml) was added to 10 ml deionized distilled water and 2.5 ml of 0.2 N Folin-Ciocalteu phenol reagent. The mixture was left undisturbed at room temperature for 5 minutes and then 2 ml of 2% sodium carbonate was added. The absorbance of the resulting solution was read at 780 nm and repeated three times. Rutin was used as a standard for calibration curve. This was done in triplicate. The results were calculated by the given formula and were given in table 7 and picture were given in fig 10.

 

C = c.V/m

C – Total phenolic compounds mg/gm of plant extract                                                                                                                  

c – The concentration of standard established from the calibration curve mg/ml

V – The volume of extract in ml

m -The weight of pure plant extract

 

Fig 10: Determination of total phenolic content of Hibiscus micranthus leaf extracts at different concentations

 

2.3 IN VITRO ANTIGLYCATION ASSAY 18

Materials: Bovine serum albumin(Sigma–Aldrich),glucose, phosphate buffer saline, trichloroacetic acid (Sigma–Aldrich), rutin (Sigma–Aldrich), centrifuge, incubator, UV spectrophotometer.

 

Method: Antiglycation activity was determined using the bovine serum albumin assay with slight modification. In all experiments, the final reaction volume was 1.0 ml and carried out in 1.5 ml Eppendorf tubes. Bovine Serum Albumin 500 µl (1 mg/ml concentration) was incubated with glucose 400 µl (500 mM final concentration) and 100 µl sample, 100 µl phosphate buffer saline was used as the sample control and 100 µl rutin (Sigma–Aldrich) as the reference standard. A negative control was carried out at the same time with BSA 500 μl (1 mg/ml concentration), 400 μl phosphate buffer saline and 100 μl sample incubated under same conditions (Fig 11).

 


 

 

Fig 11: Hibiscus micranthus leaf extracts incubated for AGE formation


The reaction was allowed to proceed at 60 °C for 24 hours and terminated by adding 10 µl of 100% (W/V) trichloroacetic acid (TCA). The TCA added mixture was kept at 4 °C for 10 minutes and centrifuged 4 minutes at 13000 rpm. The precipitate was redissolved with alkaline phosphate buffer saline (pH 10) and was quantified for the relative amount of glycated BSA based on fluoresncee intensity by UV spectrophotometer.

 

Fig 12: AGE formation inhibited in Hibiscus micranthus leaf extracts after incubation at various concentrations

 

 

The excitation and emission wavelength used were at 370 nm and 440 nm respectively. Each sample was analyzed in five concentrations and in triplicate. Percentage of inhibition was calculated using the formula given below and the sample concentration required for the 50% of inhibition was calculated using the formula given below.The results are given in table 8 and picture given in fig 11, 12 .

 

% of Inhibition = OD blank – (OD sample – OD sample negative)/ OD blank x 10.

 

3. RESULTS AND DISCUSSION:

3.1.1POWDER ANALYSIS BY NACKED EYE AND  FLUORESCENCE UNDER UV

The results of powder analysis by nacked eye are produced in table 1.

 

Table 1: Powder analysis with chemical reagents for the leaves of Hibiscus micranthus

S.No

 Reagents

Colour observed

1.

Powder as such

Light green

2.

Powder + Concentrated HCl

Brownish green

3.

Powder + Concentrated HNO3

Yellow

4.

Powder + Concentrated H2SO4

Dark brown

5.

Powder + 5% NaOH Solution

Light green

6.

Powder + 5% KOH solution

Light green

7.

Powder + 5% FeCl3

Mustard green

8.

Powder + Picric acid

Yellowish green

9.

Powder + Ammonia

Light yellowish green

10.

Powder +MethanolicNaOH

Dark green

11.

Powder + Aq. NaOH

Green

12

Pet.ether

Light green

 

The powder as such showed light green colour,  on treatment with  5% FeCl3, ammonia, Conc. HCl, Conc. HNO3 and Conc. H2SO4 showed mustard green, light yellowish green, brownish  green, yellow and dark brown colours respectively.

 

Fluorescence Analysis of the leaf powder:

The fluorescence analysis for the leaf powder of Hibiscus micranthus on treatment with chemical reagents showed the following different colours when observed under UV light.

 

Table 2: Fluorescence analysis of Hibiscus micranthus leaf powder:

S.no

Treatment with Chemical Reagents

Fluorescence  Observed

1.

Powder as such

Light green

 2.

Powder + 1N NaOH in methanol

Dark green

3.

Powder + 1N NaoH in water

Light green

4.

Powder + 50% HCL

Dark green

5.

Powder + 50% H2SO4

Dark green

 6.

Powder + 50% HNO3

Brown

 7.

Powder + Petroleum ether

Green

 8.

Powder + NH3

Light green

 9.

Powder + Picric acid

Yellowish  green

10.

Powder + FeCl3 solution

Brown

11.

Powder + 5% KOH

Dark green

 

The powder as such showed light green colour, with 1N NaOH in methanol dark green colour, with 1N NaOH in water light green colour, dark green colour with 50% HCL, 50% H2SO4, brown with 50%HNO3, with petroleum ether green, with ammonia light green, with picric acid yellowish green, with FeCl3 mustard green colour and with 5% KOH showed dark green colours respectively.

 

3.1.2 PHYSICO CHEMICAL ANALYSIS OF THE LEAF POWDER

The results of Physico chemical analysis were represented as below.

 

Table 3: Physico chemical Parameters of Hibiscus micranthus leaf powder

S.No

 Parameter

Values % w/w

1.

Total ash value

75% w/w

2.

Acid insoluble ash

25% w/w

3.

Water soluble ash

35% w/w

4.

Sulphated ash

15 %w/w

5.

Water soluble extractive

2.2% w/w

6.

Alcohol soluble extractive

19.3 %  w/w

7.

Loss on drying

79.33% w/w

 

The various Physico chemical parameters studied for the leaf powder of Hibiscus micrantus include total ash value, acid insoluble ash value, water soluble ash value, sulfated ash value, alcohol soluble extractive value, and water soluble extractive value, loss on drying or moisture content. The results were as followed 75% w/w, 25% w/w, 35% w/w, 15%w/w, 2.2% w/w, 19.3% w/w, 79.33% w/w respectively.

 

 

 

2.1.3 MICROSCOPY OF LEAF AND POWDER :

2.1.3.1 Transverse section of leaves of Hibiscus micranthus:

Transverse section of leaves of Hibiscus micranthus showed following characters.

 

Fig13(a):T.S. of Hibiscus micranthus leaf showing upper, lower epidermis and vascular bundle

 

Fig 13(b):T.S. of Hibiscus micranthus leaf showing upper and lower collenchyma and midrib

 

Fig 13(c):T.S ofHibiscus micranthus leaf showing mucilagenous cells, xylem vessels and trichomes

 

Fig 12(d):T.S of Hibiscus micranthus showing Stellate trichomes in the lamina

 

Fig 13(e):T.S of Hibiscus micranthus showing lignified Stellatecovering and glandular trichomes

 

Fig 13(f):T.S of Hibiscus micranthus showing xylem vessels in the lamina

 

Anatomical description of leaves of hibiscus micranthus:

Lamina:

Epidermis: The outermost single layer  covered with simple stellate lignified covering and glandular trichomes that are 3-4 armed, 1-celled, filiform, basal cells prominent and short and a few stomata are seen  on both upper and lower epidermis.

 

Mesophyll :It is dorsiventral leaf contains single layeed palisade cells present beneath the uper epidermis. spongy  mucilagenous parenchymatous cells and calcium oxalate druse crystals that are granular were observed.

 

Midrib: It contains collenchymatous cells below the upper epidermis and beneaththe lower epidermis cortical parenchyma. A well developed vascular bundle is seen in the centre of midrib. Xylem vessels are lignified, libiform type with very thick wall, pointed tips, narrower and have simple pits. They contain perforation plates having short pointed tails.They are longer than the tracheids of plant.

 

3.4.2 Surface preparation of leaves of Hibiscus micranthus:

Surface preparation shows anisocytic stomata and stellate trichomes.


                                    

 

 

Fig 14(a): Surface preparation of Hibiscus micranthus leaf

 


 

Fig 14(b): Surface preparation of Hibiscus micranthus leafupper epidermis showinganiocytic stomata and palisade cells

Fig 14(c): Surface preparation of lower epidermis showing anisocytic stomata and epidermal cells

 

Fig 14(d): Surface preparation showing veinislet and veinlet termination

 

Determination of leaf constants :

The results for the leaf constant value determination are represented as below.

 

Table  4: Determination of Leaf constants for Hibiscus micranthus leaves

Sl.no

Leaf constants

Value

1

Stomatal number of Upperand lower epidermis

3

2

Stomatal index

Upper epidermis

Lower epidemis

 

4.66

4.15

3

Palisade ratio

9

4

Vein islet number

12

5

Vein let termination number

9

 

The value for number of stomata was3in both upper and lower epidermis ofHibiscus micranthus leaves.The values of stomatal index for upper epidermis include 4.66 and for lower epidermis 4.15. The value for palisade ratio was 9. The value for veinlet and veinislet termination was 12 and 9 respectively.

 

Powder microscopy of leaves of Hibiscus micranthus:

The Powder characteristics of Hibiscus micranthus leaves showed epidermal cells, druse crystals of calcium oxalate, lignified xylem vessels, trachieids, lignified stellate 2-3-4 armed glandular and covering trichomes.

 

Fig 15(a): Powder microscopy –Thick walled xylem vessel

 

Fig15(b): Powder microscopy – lignified xylem vessel, lignified trichomes, mesophyllcells

 

Fig 15(c): Powder analysis - Libiform xylem vessels with short pointed tails, Stellate trichomes.

 

Fig 15(d): Powder analysis showing libiform xylem tracheid vessel with short tail

 

Fig 15(e): Powderanalysis showing glandular trichomes and covering trichomes

 

Fig 15(f): Powder analysis showing long, elongated, narrower libiform vessel

 

Fig 15(g): Powder analysis showing lignified long, elongated, narrowerlibiform vessel, xylem tracheid

 

Fig 15(h): Powder analysis showing 2 armed trichomes and calcium oxalate druses

 

Fig 15(i): Powder analysis showing libiform xylem vessel and trichome with basal cell, epidermal and mesophyll cells

Determination of length and width of xylem fibers and trichomes

The results of length and width of xylem vessels and trichomes were given as below.

 

Table 5: Determination of length and width of trichomes and xylem fibres:

Sl .No

Parameter

Measurement

Length                              Width

1

Xylem vessels

15.75µ                              117.62µ

2

Trichomes

30.6  µ                              2.87 µ

 

 

The average value for the determination of length and width of xylem vessels was found to be 15.75 and 30.6 µ and for trichomes in the leaf powder of Hibiscus micranthuswas found to be 117.62and2.87 µ.

 

3.2 PHYTOCHEMICAL STUDY

3.2.1 The results of qualitative phytochemical screening are given in table 6.

 

Table 6: Qualitative phytochemical screening of of Hibiscus micranthus leaves

S.No

Phytochemical

AQELHM

MELHM

1.

Alkaloids

+

+

2.

Tannins

+

+

3.

Phenolics

+

+

4.

Steroids

+

+

5.

Flavonoids

+

+

6.

Cardenolides

+

+

7.

Reducing sugar test

+

+

8.

Reserpine test

+

+

9.

Waxes

+

+

10.

Acid compounds

+

+

11.

Inulin test

+

+

12

Comarin glycosides

+

+

AQLHM, MELHM- aqueous extract of leaves of Hibiscus micranthus, methanolic extract of leaves of Hibiscus micranthus.

 

 

The Qualitative preliminary phytochemical screening of aqueous and alcoholic leaf extracts of Hibiscus micranthus revealed the presence of alkaloids, tropane alkaloids, opium alkaloids, purine alkaloids, tannins, phenolics, steroids, flavonoids, cardenolides, reducing sugars, reserpine, waxes, acid compounds, inulin, mucilage and coumarine glycosides were present.

 

3.2.2 Quantitative determination

Table 7: Quantitative Phytochemical Screeningof aqueous and alcoholic extracts of Hibiscus micranthusleaves

S.no

Phytochemical

AQLHM

MELHM

1.

Flavanoids (mg/g)

72±0.20

104±0.36

2.

Phenolics (mg/g)

25.4±0.40

52.76±0.67

Values represented mean ± S.D. of three parallel measurements. AQLHM, MELHM- aqueous extract of Hibiscus micranthus, methanolic extract of leaves of Hibiscus micranthus.

 

Fig 16: Standard calibration curve of Rutin for flavonoids

 

Fig 17: Standard calibration curve of Rutin for phenolics

 

The quantitative preliminary phytochemical screening of the total flavonoid content was 72 mg/g equivalent of standard drug rutin and the phenolic content was 25.4 mg/g equivalents of standard drug rutin in the aqueous extract of leaves of Hibiscus micranthus. The methanolic extract of leaves of Hibiscus micranthus shows flavanoid content 104 mg/g equivalent of standard drug rutin and phenolic content was 52.76mg/g equivalent of standard drug rutin respectively.

 

The results indicated that flavonoids and phenolics were present in more amount in the methanolic extract of the leaves of Hibiscus micranthus when compared to the aqueous extract.

 

3.3 IN VITRO ANTIGLYCATION ACTIVITY

In vitro antiglycation activity was determined as percentage inhibition of AGE’S by bovine serum albumin assay method in both aqueous and methanolic extracts of leaves of Hibiscus micranthus at various concentrations (200, 400, 600, 800, 1000  µg/ml) was as followed.




 

Antiglycation by Bovine serum albumin assay method

Table 8:  Antiglycation effect of different concentrations of aqueous and methanolic extracts of Hibiscus micranthusleaves by bovine serum albumin assay method

S.No.

Concentration (µg/ml )

AQLHM

       MELHM

STD

1.

200µg/ml

48.33%±0.0029**

56.25%±0.0012**

69.88%±0.0016*

2.

400µg/ml

50.32%±0.001**

68.33%±0.0013*

73.42%±0.0012*

3.

600µg/ml

56.08%±0.078**

76.25%±0.044*

79.99%±0.0042*

4.

800µg/ml               

61.66%±0.0018**

80.39%±0.0028**

87.32%±0.004*

5.

1000µg/ml

91.25%±0.008**

95.06%±0.0025**

96.02%±0.0011**

The values are expressed as Mean ±SEM, n= 6. The values are significant,* p< 0.05;**p < 0.01 when compared with standardAqELHM, MELHM, STD– aqueous extract of leaves of Hibiscus micranthus, methanolic extract of leaves of Hibiscus micranthus, Standard.

 

 


Fig 18: Bovine serum albumin assay on the leaf extracts of Hibiscus micranthus

 

The percentage inhibition of aqueous extract of leaf extracts of Hibiscus micranthus by bovine serum albumin assay at various concentrations (200,400, 600, 800, 1000 µg/ml) was 48.33% ± 0.0029, 50.32% ± 0.00127, 56.08% ± 0.0786, 61.66%±0.0018, 91.25% ± 0.008 respectively. The percentage inhibition of methanolic extract of leaf extracts of Hibiscus micranthus by bovine serum albuminassay at various concentrations (200, 400, 600, 800, 1000 µg/ml) was 56.25%±0.0012, 68.33%±0.0013*, 76.25%±0.044*, 80.39%±0.0028**, 95.66%±0.0025**. The percentage inhibition of standard rutin by nucleation assay at various concentrations (200, 400, 600, 800, 1000µg/ml) was 69.88%±0.0016*, 73.42%±0.0012*, 79.99%± 0.0042*, 87.32%±0.004* 96.02%±0.0011** respectively.

 

The alcoholic extract of leaves of Hibiscus micranthus was showing significant antiglycation activity at a concentration of 1000µg/ml when compared with standard as more amount of phenolics and flavonoids were present in the alcoholic extract.

 

4. ACKNOWLEDGEMENT:

The authors are thankful to Viaya institute of Pharmaceutical Sciences for Women for providing facilities to carry out the research work.

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Received on 13.11.2016             Modified on 22.12.2016

Accepted on 15.01.2017           © RJPT All right reserved

Research J. Pharm. and Tech. 2017; 10(3): 663-673.

DOI: 10.5958/0974-360X.2017.00124.X

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