INTRODUCTION:

Among metabolic disorders, diabetes mellitus is biggest concern of modern society. Both type 1 and type 2 diabetes are associated with rise in blood glucose concentration, either due to abnormal secretion or hindered insulin sensitivity^1,2. Insulin plays an imperative role in the regulation of blood glucose. Pancreatic hormones and metabolic disturbances collaborates to form the platform for prevalence of this heterogeneous disorder³. Main factors includes abnormality of insulin receptors, signal transducer system, effecter enzymes or genes⁴.

Although diabetes mellitus have become one of the major concern for today’s generation, yet it cannot be considered as a contemporary disorder as evidence of its historical existence was reported in Egyptian manuscript three thousand years ago⁵. It is a matter of concern throughout the world, as its cases are exponentially increasing^6,7. Its expansion is a great burden not only for patients, but for health sector and society also⁸. According to International Diabetes Federation (IDF), it has been accounted that total number of diabetic patients are continuously rising, in fact in year 2000 there are 151 million cases. However, in year 2015 it increases up to 415 million. Moreover, it is expected that in year 2040 this count will reach up to 642⁶. In year 2011, South East Asia has 71.4 million cases of diabetes patients and it is assumed that it will cross 120.9 million up to year 2030 which is nearly 70 percent rise in diabetic patients^9,10. Effective medicinal treatment of diabetes mellitus has always been a challenge, no therapeutic approach is capable to treat this disorder completely without side effects^11,12. Hypoglycemic potential of numerous plants, used in diabetes treatment, has been proved already¹³. Lesser toxicity and side effects is major advantage of herbal formulations in contrast to synthetic drugs¹⁴. Furthermore, taking care of WHO recommendation on diabetes mellitus, it become necessary to investigate hypoglycemic potentials of medicinal plants^15,16. Therefore, research on medicinal plants and their constituents becoming a trend to find new mode of treatment¹⁷.

Typha elephantina Roxb., (Figure 1) is a perennial aquatic plant. It belongs to genus Typha Linn., commonly known as ‘Cattails’¹⁸. First description of Typha elephantina was given by Roxburgh (1832)¹⁹. It appears like grasses that may attain height between two - five meters²⁰. Leaves are thick and broad with flattened leaf blades and consist aerechymatous spongy tissue²¹.

Geographically, Typha elephantina is widely distributed among several continents except Antarctica²², and southern equatorial region of Africa¹⁹. Moreover, It is native to North Africa, India, Nepal, Pakistan and Iran ²³. In north India, Typha elephantina is common in swamps^24,25. Typha elephantina has been used traditionally to cure various disorders like burning syndrome, different blood disorders, bacterial infections like erysipelas, blood clotting disorders, Cystitis, burning during micturition, calculus, swelling, oligospermia, and ulcers²⁶. Looking into its traditional capabilities in treatment of various diseases and hypoglycemic effects of other plants of typha genus, current study is designed to utilize in vivo and in vitro approaches to estimate its antidiabetic potentials.

MATERIAL AND METHODS:

Animals:

Wistar rats weighing 180 ± 20g were incorporated into study. Animals were kept under proper sanitation and hygiene. Normal conservation conditions provided in clean polypropylene cages at 23±1ºC, equal hours light: dark period and relative humidity was maintained at 60±4%. Sufficient supply of feed and water was maintained. The animals had been acclimatized to the laboratory environment prior to the start of the experimentation. Animal care and experimentation during whole study was carried as per guidelines provided²⁷. The experiment protocol was accepted by the IAEC at M.M. College of Pharmacy, M.M.U Mullana (IAEC/19/20).

Plant collection and authentication:

Plant material was collected from roadside marshy areas of Kurukshetra, Haryana during July to September. Its authentication was done by Dr. B. D. Vashishta, Botany department, Kurukshetra University, Haryana, India.

Extract preparation:

After shade drying, plant material was coarsely powdered and then successively extracted using different solvents with increasing polarity in Soxhlet apparatus. Initially pet-ether (60-70°C) was used to remove fats from the plant material, as extracts containing lipid content cause hindrance in spectroscopic evaluation of isolated compound, followed by extraction with chloroform and methanol. The whole extraction procedure was carried at same temperature. After collection of the extracts rotary evaporator was then employed to concentrate them so that a concentrated crude mass can be formed for further study.

In vitro:

α-amylase activity:

α-amylase enzymatic starch digestion was used to investigate potential of all the three extracts of Typha elephantina for α-amylase inhibition, method followed as per Ali et al. (2006) with some alterations. In brief 30μl of varying concentrations of plant extracts that is 8, 15, 30, 60 and 125µg/ml, 200μl alpha-amylase was added, and then incubation was done for 20 minutes at 37°C. Then addition of 100μl of the (1%) starch solution was done and again subjected to incubation for 10 minutes at 37ºC. To stop the reaction, 200μl DNSA was added. Readings were taken at 540nm wavelength. Acarbose was taken as standard. To enhance the reproducibility and resolution of the results, experiments were repeated thrice using the same protocol^28,29.

α-glucosidase inhibition assay:

In α-glucosidase assay 95μl of the phosphate buffer (100mM) was added to 96 well micro plate. 25μl of the Alpha-glucosidase (.5U/ml) was added, and then 30μl of plant extracts (8, 15, 30, 60 and 125µg/ml) and Acarbose in same concentrations was added. Acarbose was taken as standard for the experiment. The above reaction mixtures were subjected to incubation for 20 minutes at 37ºC. After incubation, 50μl of 5mM p-nitrophenyl -D-glucopyranoside (p-NPG) was added and subjected to incubation for 10 minutes at 37ºC. 2ml of Na₂CO₃ (0.1M) was added for termination of the reaction. Absorbance was taken at 415 nm using iMark Micro plate Reader. Quantity of alpha-nitrophenol released from p-NPG was used to estimate percent inhibition^30,31.

% Inhibition = 100 × absorbance of control - absorbance of sample/Absorbance of control.

Antioxidant activity:

DPPH antioxidant assay:

DPPH stock solution was made by mixing 3.3mg of DPPH in 100ml of methanol. 1ml of different concentrations of the test solution (8, 15, 30, 60 and 125 µg/ml) was put in 2.5ml of stock solution. After keeping for half an hour, readings were taken at 517nm in a UV spectrophotometer in contrast to the standard at varying concentrations (2, 4, 6, 8 and 10µg/ml). Ascorbic acid was taken as reference^32,33. Calculation was carried using formula given below.

% Inhibition = 100 × absorbance of control - absorbance of sample/Absorbance of control

Hydrogen peroxide antioxidant activity:

For the evaluation, a solution was made by mixing hydrogen peroxide (40mM) and phosphate buffer with pH 7.4. To 0.6ml of above mixture, varying concentrations (25, 50, 100, 200 and 400µg/ml) of samples and (10, 20, 40, 60, 80µg/ml) of standard was added. After ten minutes, readings of hydrogen peroxide were taken at 230 nm. Blank contained only buffer solution^34,35. Same formula used in DPPH assay was employed to find percentage inhibition of hydrogen peroxide in samples as well as standard compounds.

In vivo

Acute toxicity studies:

For the acute toxicity evaluation, the guidelines of the Organization of Economy and Cooperation development (OECD 423) were followed. 8-12 weeks old albino mice weighing 20 to 30g were included for toxicity studies. Animals were fasted for 3-4 hours prior to dosing. After that, weighing of the animals was done followed by administration of the extracts using gastric intubation. Even after administration of the extracts, animals were not provided any food for next couple of hours^36,37. Dose of 2000mg/kg was given to 6 animals initially. For initial 30 minutes, animals were monitored independently upon dosing and periodically over 24 hours, out of which first four hours were considered crucial, hence monitored vigilantly and regularly afterwards for duration of 14 days³⁸.

Oral glucose tolerance test (OGTT):

Four groups of rats were made on random basis, having six animals each. all animals were fasted for twelve hours before experimentation. Normal control G-I animals were only administered with vehicle; G-II animals was treated with 2.5mg/kg of standard drug Glibenclamide; Group III and IV received Typha elephantina methanol extract (TEME) (250 and 500mg/kg) and served as test groups. Glucose at dose 2000mg/kg was fed, 30 minutes prior to the oral administration of test and standard compounds, to all selected animals³⁹. After 30, 60, and 120 min of oral glucose, blood was withdrawn from tail on to the strip of glucometer and concentration of glucose in the blood was traced using Accu-Chek, Roche Diagnostics, USA⁴⁰.

STZ-induced Diabetic study:

Single dose of streptozotocin (55mg/kg)⁴¹ was administered intraperitoneally to the overnight fasted rats⁴². Dose preparation was carried spontaneously using very cold citrate buffer with pH 4.5. G-I animals were given no treatment or STZ throughout the study. For the next twelve days animals were kept under observation with proper feed and water, and on the twelfth day animals were screened for blood glucose. Animals were believed to have diabetes, if blood glucose levels were found to be 200 mg/dl or higher and incorporated into the study^43,44.

Experimental design:

5 groups (G) of rats were assigned containing six rats (n=6) each.

G-I Normal healthy rat’s administered 0.9 percent sodium chloride (NaCl).

G-II Diabetic rats were given only 0.9 percent NaCl.

G-III Diabetic rats + glibenclamide (2.5mg/kg)

G-IV Diabetic rats + Typha elephantina methanol extract (250 mg/kg).

G-V Diabetic rats + Typha elephantina methanol extract (500 mg/kg).

Dose of methanol extract of Typha elephantina and Glibenclamide were made in vehicle solution (0.9% NaCl) at room temperature and administered orally for 21 days. Animal were weighed on first day of study and after 21 days. Liver and pancreas of all animals were taken out and weighed after completion of the study.

Biochemical estimations:

For tracing blood glucose levels, one touch glucometer (Accuchek, Roche Diagnostics) was used at weekly intervals. After completion of the complete study, blood sample were procured in EDTA tubes, by cardiac puncture as well as retro-orbital plexus method, from all animals. After that, for 20 minutes at 3000, rpm centrifugation of blood samples was done. Serum separation and storage at -20 degree Celsius was done till tests were performed⁴⁵. Complete lipid profile including Cholesterol, HDL, LDL, VLDL, triglycerides, Urea, Creatinine analysis was done from collected serum samples with the help of auto-analyzer using ERBA diagnostic kits.

Statistical analysis:

For tracing significant difference (P < 0.05), One-way ANOVA was applied and comparative analysis was done with Dunnet’s t-test in vivo studies, for instance, blood glucose, body weight, biochemical parameters and in vitro inhibition assays. For representation of Results, mean± standard error of mean, pattern was employed.

RESULTS:

In vitro:

α- Amylase activity:

Different concentrations 8, 15, 30, 60, 125µg/ml of Typha elephantina petroleum ether, chloroform and methanol extract were selected. Methanol extract of Typha elephantina (TEME) exhibited 57.48±1.42 in comparison to 66.7±0.94 percent inhibition of standard Acarbose shown in Figure 1. Percent inhibition of chloroform and petroleum ether extract of Typha elephantina was found to be insignificant in comparison to that of standard.

α- Glucosidase Inhibition Assay:

Typha elephantina extracts were studied at 8, 15, 30, 60, 125µg/ml concentration and TEME exhibited significant inhibition in comparison to standard Acarbose while TECE and TEPE showed minimum inhibition (Figure 2). With increase in concentration, percentage inhibition was found to be increased. The maximum enzyme inhibition with standard drug i.e. Acarbose was 70.31±1.25 at 125µg/ml, while TEME exhibited 53.64±0.92 percent inhibition.

Antioxidant activity:

DPPH assay for radical scavenging activity:

In DPPH assay, TEME showed 56.44±1.64 percent inhibition at 125µg/ml, while TEPE and TECE exhibit only 19.16±0.36 and 38.16±0.62 respectively. TEME exhibits comparable inhibition to that of standard ascorbic acid (Table 1).

H₂O₂ free radical scavenging activity:

TEME at 25, 50, 100, 200 and 400µg/ml, exhibited efficient results in hydrogen peroxide radical scavenging activity in contrast to ascorbic acid taken as standard, which showed 79.91 inhibition at 400µg/ml concentration details given in Table 2.

Figure 1: Percent inhibition of α-Amylase inhibition by Typha elephantina ; (TEPE: Typha elephantina petroleum extract, TECE: Typha elephantina chloroform extract, TEME: Typha elephantina methanol extract)

Figure 2: α- glucosidase inhibition activity of Typha elephantina; (TEPE: Typha elephantina petroleum extract, TECE: Typha elephantina chloroform extract, TEME: Typha elephantina methanol extract)

Table 1: DPPH scavenging activity of extracts of Typha elephantina

Drug	8µg/ml	15µg/ml	30µg/ml	60µg/ml	125µg/ml
TEPE	4.83±1.64	8.77±2.18	11.28±1.08	15.94±0.36	19.16±0.36
TECE	14.15±1.29	17.73±0.62	27.77±0.95	35.29±0.95	38.16±0.62
TEME	20.24±0.72	26.33±0.62	40.67±0.95	50.35±0.95	56.44±1.64
	2 µg/ml	4 µg/ml	6µg/ml	8 µg/ml	10 µg/ml
Ascorbic Acid	37.09±0.62	44.25±0.36	53.22±0.62	66.12±0.62	75.08±1.29

TEPE: Typha elephantina petroleum ether extract; TECE: Typha elephantina chloroform extract; TEME: Typha elephantina methanol extract

Table 2: H₂O₂ scavenging activity of extracts of Typha elephantina

DRUG	25 µg/ml	50 µg/ml	100 µg/ml	200 µg/ml	400 µg/ml
TEPE	5.18±0.35	6.23±1.27	13.95±1.61	21.67±1.27	26.58±0.61
TECE	11.49±0.93	16.05±0.61	24.47±0.61	35.7±1.27	44.47±1.61
TEME	26.23±0.35	29.04±1.86	41.32±0.61	52.89±0.61	58.16±0.61
	10 µg/ml	20 µg/ml	40 µg/ml	60 µg/ml	80 µg/ml
Ascorbic Acid	40.61±0.7	48.33±1.27	53.6±0.93	64.12±0.93	79.91±0.35

TEPE: Typha elephantina petroleum ether extract; TECE: Typha elephantina chloroform extract; TEME: Typha elephantina methanol extract

In vivo

Toxicity study:

No mortality was observed in animals both sex. Moreover, no clinical sign were observed in any animal. The overall study showed that LD50 of oral toxicity of extract to be above 2000mg/kg body weight in mice.

OGTT:

Blood glucose concentration of Group I (normal control) enhanced significantly in comparison to the Group III and IV. The mean blood glucose level after 30 min in Group I was 145.71mg/dl (Table 3). Oral administration of extracts and standard drug caused comparable fall in blood glucose, in contrast to the result obtained from Group I. Gliblenclamide (2.5mg/kg) along with TEME (500mg/kg) exhibited reduction of blood glucose concentrations to a maximum after 30 minutes of glucose administration (121.10±1.60mg/dl and 128.89±1.68mg/dl respectively).

Effect on body weight:

G-II (diabetic control) rats were observed to have remarkable reduction in the body weight, in contrast to Group I (normal control) and treated Groups III, IV and V as given in Table 4. Mean body weight of normal control group was 178.83±1.54g at the beginning of the study that got raised to 189.56±1.63g after 21 days. Body weight of G-II rats was significantly and sequentially reduced from 187.67±1.48g to 141.81±1.03g from beginning to the end of study. The mean body weight in Group III at Day 1 was 191±2.73g, while these values were 195.55±3.08, 209.24±3.3, 206.28±2.95 g at 7th, 14th and 21st day respectively. 500 mg/kg dose of TEME showed significant rise in body weight from 186.42±1.46g to 208.79±1.64g. In addition to bodyweight, weight of liver and pancreas was also observed which reflected enhancement in weight of liver on the other hand reduction in pancreas weight details given in Table 4.

STZ-induced Diabetic study

Remarkable rise of blood glucose was analyzed in diabetic control (G-II) animals in contrast to the Group I, Group III, IV and V. In normal control (G-I), mean blood glucose was 103.66±3.31mg/dl on the 1st day and 109.33±2.35mg/dl on 21^stday. In Group II (diabetic control), blood glucose level on the (1st day) was 269.67 ± 3.31 mg/dl and increased to 305.83 ± 2.91mg/dl on (21st day). The mean body glucose in Group III at Day 1 was 270.17±3.02mg/dl, while these values were 121.58±1.36, 113.47±1.27, 108.07±1.21mg/dl at 7th, 14th and21st day respectively. TEME exhibits remarkable fall in blood glucose levels in contrast to the result obtained from diabetic animals. Animals administered 500mg/kg of TEME reflected fall in blood glucose from 271.67±2.98 to 141.50±3.54. Details are given in Table 5.

Biochemical parameter

There was a marked increase in all the biochemical parameters except HDL in Group II (Diabetic control). Standard drug Gliblenclamide (2.5mg/kg) effectively controlled the levels of all the biochemical parameters. 500 mg/kg dose of TEME exhibited better efficacy than TEME 250mg/kg in decreasing cholesterol levels and increasing HDL level (Table 6). 21 days of TEME Administration remarkably reduced SGOT, SGPT, creatinine and urea level in dose dependent manner, of diabetic animals.

Table 3: OGTT study results of Typha elephantina methanol extract

Groups N=6	Treatment	Glucose concentration (mg/dl)
Groups N=6	Treatment	0 min	30 min	60 min	120 min
I	Normal Control	98.83±1.77	145.71±2.73	129.83±1.53	118.33±0.87
II	Gliblenclamide (2.5 mg/kg)	100.91±1.34	121.10±1.60***	111.01±1.47***	105.96±1.40***
III	TEME 250	102.14±1.44	137.89±1.95*	120.97±2.17**	115.93±2.08
IV	TEME 500	97.25±2.08	128.89±1.68***	115.14±1.50***	105.38±1.38***

Readings represented as mean ± Standard error mean (n=6).* p < 0.05,** p < 0.01 and *** p < 0.001 In comparison to normal control animals (One-way ANNOVA with Dunnet’s t-test); TEME: Typha elephantina methanol extract

Table 4: Liver, Pancreas and body weight analysis of rats after treatment with TEME

Groups N=6	Treatment	Body weight (gm)				Liver Weight(g)	Pancreas Weight (g)
Groups N=6	Treatment	1st Day	7th Day	14th Day	21st Day	Liver Weight(g)	Pancreas Weight (g)
I	Normal Control + .1 ml vehicle	178.83±1.54*	185.37±1.76*	185.99±1.6***	189.56±1.63***	5.4±0.06***	0.84±0.03***
II	Diabetic Control+ .1 ml vehicle	187.67±1.48	175.43±1.57	157.9±1.2	141.81±1.03	4.05±0.04	0.99±0.03
III	Diabetic+ Gliblenclamide (2.5 mg/kg)	191±2.73	195.55±3.08***	209.24±3.3***	206.28±2.95***	5.45±0.04***	0.86±0.03***
IV	Diabetic + TEME (250mg/kg)	192.25±2.78	208.4±3.01***	211.48±3.06***	217.24±3.14***	5.32±0.14***	0.81±0.02***
V	Diabetic + TEME (500mg/kg)	186.42±1.46	199.47±1.57***	204.87±1.61***	208.79±1.64***	5.16±0.13***	0.78±0.02***

Readings represented as mean ± Standard error mean (n=6).* p < 0.05,** p < 0.01 and *** p < 0.001 In comparison to diabetic control animals (One-way ANNOVA with Dunnet’s t-test); TEME: Typha elephantina methanol extract

Table 5: Blood glucose levels of rats treated with TEME and standard drug

Groups N=6	Treatment	Fasting blood glucose (mg/dl)
Groups N=6	Treatment	1st Day	7th Day	14th Day	21st Day
I	Normal Control + .1 ml vehicle	103.66±3.31***	109.83±1.70***	107.16±3.02***	109.33±2.35***
II	Diabetic Control+ .1 ml vehicle	271.67 ± 3.31	285.50 ± 1.52	296.67 ± 1.74	305.83 ± 2.91
III	Diabetic + Gliblenclamide (2.5 mg/kg)	270.17±3.02	121.58±1.36***	113.47±1.27***	108.07±1.21***
IV	Diabetic + TEME (250mg/kg)	267.33±2.275*	215.47±1.82***	202.00±1.70***	203.17±3.04***
V	Diabetic + TEME (500mg/kg)	271.67±2.98	149.42±1.64***	144.83±2.83***	141.50±3.54***

Table 6: Effect of TEME biochemical parameter of diabetic rats

Parameters	Normal Control + 0.1 ml vehicle	Diabetic Control+ 0.1 ml vehicle	Diabetic + Gliblenclamide (2.5 mg/kg)	Diabetic + TEME (250 mg/kg)	Diabetic +TEME (500 mg/kg)
Total Cholesterol (mg/dl)	99.15±1.79***	237.96±4.3	112.54±2.03***	157.77±2.85***	132.97±2.4***
Triglycerides (mg/dl)	77.87±0.95***	179.09±2.19	84.87±1.04***	150.91±1.84***	105.03±1.28***
HDL (mg/dl)	47.4±0.78***	27.44±0.45	49.77±0.82***	38.28±0.63***	34.13±0.56***
LDL (mg/dl)	36.18±2.18***	174.7±4.61	45.79±2.43***	89.31±3.2***	77.84±2.69***
VLDL (mg/dl)	15.57±0.19***	35.82±0.44	16.97±0.21***	30.18±0.37***	21.01±0.26***
SGOT (IU/L)	34.97±1.24***	74.3±2.65	41.96±1.49***	60.58±2.16***	50.2±1.79***
SGPT (IU/L)	24.75±1.03***	77.97±3.23	32.18±1.33***	42.33±1.76***	42.8±1.77***
Creatinine (mg/dl)	0.47±0.05***	0.98±0.11	0.56±0.06**	0.96±0.1	0.73±0.08
Serum Urea (mg/dl)	34.55±0.55***	107.12±1.7	38.01±0.6***	78.37±1.24***	70.86±1.13***

DISCUSSION:

Whole study was focused to trace out antidiabetic properties of Typha elephantina leaves, by animal studies, α-amylase, α- glucosidase assays and prediction of the responsible constituents by molecular docking. Three different tools were employed for the determination of the goal, including in vitro and in vivo studies. In vitro α-amylase and α-glucosidase were carried to identify antidiabetic potential between Typha elephantina methanol, petroleum ether and chloroform extract. Further, in vivo study results provided the testimony of Typha elephantina leaves methanol extract to possess antidiabetic potentials. Diabetic rats administered with 250mg/kg and 500mg/kg of TEME gave comparable results to the group administered standard Gliblenclamide (2.5mg/kg), in reducing blood glucose levels. 500mg/kg of TEME gave more significant results. In addition, results of other biochemical parameters like total cholesterol, triglycerides, HDL, SGOT, SGPT, urea and creatinine found to be better in Drug treated groups in contrast to control group. Typha elephantina extracts also exhibited antioxidant properties, by DPPH and Hydrogen peroxide assays, indicates towards role of this plant in diabetes. Subsequently, more research is required to analyze the molecular mechanism involved behind the tendency of Typha elephantina in reducing and control of blood glucose.

REFERENCES:

1. El-Atat F, McFarlane SI and Sowers JR. Diabetes, hypertension, and cardiovascular derangements: pathophysiology and management. Current Hypertension Reports. 2004; 6(3): 215-23.

2. Goyal R and Jialal I. Diabetes Mellitus Type 2: Stat Pearls Publishing; 2019.

3. Organization WH. The world health report: 1999: making a difference: World Health Organization; 1999.

4. Kharroubi AT and Darwish HM. Diabetes mellitus: The epidemic of the century. World Journal of Diabetes. 2015; 6(6): 850.

5. Ahmed AM. History of diabetes mellitus. Saudi Medical Journal. 2002; 23(4): 373-8.

6. Wang Z, Zhao X, Liu X, Lu W, Jia S, Hong T, Li R, Zhang H, Peng L and Zhan X. Anti-diabetic activity evaluation of a polysaccharide extracted from Gynostemma pentaphyllum. International Journal of Biological Macromolecules. 2019; 126: 209-14.

7. Maheshwari RA, Khatri K, Sailor G and Balaraman R. Antidiabetic activity of Dibolin (a polyherbal formulation) in streptozotocin-nicotinamide induced type 2 diabetic rats. Int J Pharm Pharm Sci. 2014; 2: 893-7.

8. Khan Y, Gupta P, Bihari B and Verma VKJAJoPR. A Review on Diabetes and Its Management. 2013; 3(1): 28-33.

9. Shrestha AD, Kosalram K and Gopichandran V. Gender difference in care of type 2 diabetes. Journal of the Nepal Medical Association. 2013; 52(189).

10. Atlas ID. Brussels, Belgium: international diabetes federation; 2013. International Diabetes Federation. 2017.

11. Verspohl E. Novel pharmacological approaches to the treatment of type 2 diabetes. Pharmacological Reviews. 2012; 64(2): 188-237.

12. Kaur HJAJoNE and Research. Effectiveness of structured teaching programme regarding self care management in relation to prevention of complications among diabetics. 2014; 4(3): 279-83.

13. Sabu M and Kuttan R. Anti-diabetic activity of medicinal plants and its relationship with their antioxidant property. Journal of Ethnopharmacology. 2002; 81(2): 155-60.

14. Sharma A, Shanker C, Tyagi LK, Singh M and Rao CV. Herbal medicine for market potential in India: an overview. Acad J Plant Sci. 2008; 1(2): 26-36.

15. Organization WH. WHO Expert Committee on Diabetes Mellitus [meeting held in Geneva from 25 September to 1 October 1979]: second report. 1980.

16. Tripathi S, Saroj BK, Khan MYJAJoP and Technology. Pharmacological Evaluation of Folk Medicinally used Plant by usin Streptozotocin Induced Rat Models. 2018; 8(4): 231-43.

17. Tiwari AK and Rao JM. Diabetes mellitus and multiple therapeutic approaches of phytochemicals: Present status and future prospects. Current Science. 2002: 30-8.

18. Ruangrungsi N, Aukkanibutra A, Phadungcharoen T and Lee M. Constituents of Typha elephantina. Scientific Society. 1987; 13: 57-62.

19. Sharma K and Gopal B. A note on the identity of Typha elephantina roxb. Aquatic Botany. 1980; 9: 381-7.

20. Moniruzzaman M, Rahman M, Aktar S and Khan M. Equilibrium and Kinetic Parameters Determination of Cr (VI) Adsorption by Hogla Leaves (Typha elephantina Roxb.). International Journal of Waste Resources. 2017; 7(301): 2.

21. Uddin M, Mukul SA, Khan M, Chowdhury M, Uddin M and Fujikawa S. Indigenous management practices of Hogla (Typha elephantina Roxb.) in local plantations of floodplain areas of Bangladesh. Subtropical Agriculture Research and Development. 2006; 4(3): 114-9.

22. Zhou B, Tu T, Kong F, Wen J and Xu X. Revised phylogeny and historical biogeography of the cosmopolitan aquatic plant genus Typha (Typhaceae). Scientific Reports. 2018; 8(1): 8813.

23. Boulos L. Flora of egypt: Al Hadara Publishing Cairo; 2005.

24. Sharma K and Gopal B. Seed germination and occurrence of seedlings of Typha species in nature. Aquatic Botany. 1978; 4: 353-8.

25. Bhatti J, Veer V and Negi B. Discovery of the natural habitat of the aquatic thysanopteran, Organothrips indicus (Terebrantia: Thripidae) in India and North America. Oriental Insects. 1998; 32(1): 259-66.

26. Khare CP. Ayurvedic pharmacopoeial plant drugs: expanded therapeutics: Routledge; 2015.

27. Council NR. Guide for the care and use of laboratory animals: National Academies Press; 2010.

28. Sudha P, Zinjarde SS, Bhargava SY and Kumar AR. Potent α-amylase inhibitory activity of Indian Ayurvedic medicinal plants. BMC Complementary and Alternative Medicine. 2011; 11(1): 5.

29. Anarthe S, Chaudhari SJRJoP and Phytochemistry. Hypoglycemic Activity of Leucas linifolia (Lamiaceae) Spreng. 2011; 3(1): 34-7.

30. Rani A, Kumar S and Khar RK. In Vitro Antidiabetic and Hypolipidemic Activity of Selected Medicinal Plants.

31. Kumar DL, Rao K, Bindu M, Kumar DS, David BJRJoP and Pharmacodynamics. Alpha-Glucosidase Inhibitory Activities of Wrightia tinctoria Roxb and Schrebera swietenoides Roxb Bark Extract. 2011; 3(1): 22-4.

32. Tiwari P and Patel RKJAJoPA. Estimation of total phenolics and flavonoids and antioxidant potential of Ashwagandharishta prepared by Traditional and Modern Methods. 2013; 3(4): 147-52.

33. Krishna KV, Karuppuraj V and Perumal KJAJoPA. Antioxidant activity and Folic acid content in indigenous isolates of Ganoderma lucidum. 2016; 6(4): 213-5.

34. Nabavi S, Ebrahimzadeh M, Nabavi S, Hamidinia A and Bekhradnia A. Determination of antioxidant activity, phenol and flavonoids content of Parrotia persica Mey. Pharmacologyonline. 2008; 2(9): 560-7.

35. Potbhare M, Phendarkar N and Khobragade DJAJoRiPS. In vitro Evaluation of Antioxidant Potential of N-Acetyl-D-Glucosamine. 2017; 7(2): 120-2.

36. Verma A, Jana GK, Sen S, Chakraborty R, Sachan S and Mishra A. Pharmacological evaluation of Saraca indica leaves for central nervous system depressant activity in mice. J Pharm Sci Res. 2010; 2(6): 338-43.

37. Pandit R, Phadke A and Jagtap A. Antidiabetic effect of Ficus religiosa extract in streptozotocin-induced diabetic rats. Journal of Ethnopharmacology. 2010; 128(2): 462-6.

38. Thippeswamy B, Mishra B, Veerapur V and Gupta G. Anxiolytic activity of Nymphaea alba Linn. in mice as experimental models of anxiety. Indian Journal of Pharmacology. 2011; 43(1): 50.

39. Udia P, Ogbonna O, Antai A, Mbatutung I, Eyo SJRJoP and Phytochemistry. Oral glucose tolerance test and some haematological effects of aqueous leaf extract of Rothmannia hispida (K. Schunn) fargel on normoglycaemic albino rats. 2013; 5(6): 300-5.

40. Kaushik P, Kaushik D, Khokra SL and Sharma A. Antidiabetic activity of the plant Abutilon indicum in streptozotocin-induced experimental diabetes in rats. Int J Pharmacog Phytochem Res. 2010; 2: 45-9.

41. Aslan M, Orhan DD, Orhan N, Sezik E and Yesilada E. In vivo antidiabetic and antioxidant potential of Helichrysum plicatum ssp. plicatum capitulums in streptozotocin-induced-diabetic rats. Journal of Ethnopharmacology. 2007; 109(1): 54-9.

42. Muruganandan S, Srinivasan K, Gupta S, Gupta P and Lal J. Effect of mangiferin on hyperglycemia and atherogenicity in streptozotocin diabetic rats. Journal of Ethnopharmacology. 2005; 97(3): 497-501.

43. Nayak Y, Hillemane V, Daroji VK, Jayashree B and Unnikrishnan M. Antidiabetic activity of benzopyrone analogues in nicotinamide-Streptozotocin induced Type 2 diabetes in rats. The Scientific World Journal. 2014; 2014.

44. Raj SM, Mohammed S, Kumar SV, Debnath SJRJoP and Phytochemistry. Antidiabetic effect of Luffa acutangula fruits and histology of organs in streptozotocin induced diabetic in rats. 2012; 4(2): 64-9.

45. Saifi A, Chauhan R and Dwivedi JJAJoPR. Development of a polyherbal formulation FMST and evaluation for antidiabetic activity in alloxan induced diabetic rats. 2017; 7(1): 1-7.

Received on 20.06.2020 Modified on 27.07.2020

Research J. Pharm. and Tech. 2021; 14(6):3150-3156.

DOI: 10.52711/0974-360X.2021.00549