The Effect of Hydro Alcoholic Nettle (Urtica dioica) Extract on Oxidative Stress in Patients with Type 2 Diabetes: A Randomized Double-blind Clinical Trial
Diabetes type 2 is a metabolic disorder that characterized
by hyperglycemia and insulin resistance. Hyperglycemia and impairment of oxidant/antioxidant
balance, can increase oxidative stress and increase risk of cardiovascular disease.
In the present study, Effects of hydro alcoholic extract of Nettle on oxidative
stress in type 2 diabetes were evaluated. Fifty patients (27 men, 23 women)
with type 2 diabetes patients were studied. They received 100 mg kg-1
of nettle extract of body weight hydro alcoholic for 8 weeks. At the baseline
and end of 8th weeks of intervention blood levels of oxidative stress markers
were measured. Data was analyzed by SPSS version 18, p<0.05 was considered
significant for all variables. After 8 weeks, Total Antioxidant Capacity (TAC)
and Superoxidant Dismutase (SOD) showed a significant increase in the intervention
group compared to the control group (p<0.05). The findings showed that the
hydro alcoholic extract of nettle has increasing effects on TAC and SOD in patients
with type 2 diabetes without no changes in Malondialdehyde (MDA) and Glutathione
Peroxides (GPX) after eight weeks intervention.
Received: November 23, 2011;
Accepted: November 28, 2011;
Published: February 16, 2012
Free radicals are atoms or molecules with unpaired electron, so they are active
and can damage different tissues in the body. In healthy person there is a balance
between antioxidant enzymes and free radical species in the body. The imbalance
causes oxidative stress (Atalay and Laaksonen, 2002).
Diabetes is one of the common diseases that are associated with increased oxidative
stress (Abdel-Hamid et al., 2008). Hyperglycemia
is one of the predisposing factor for oxidative stress (Rasheed
et al., 2008), increasing oxidative stress can increase risk factors
of cardiovascular incidence and other complications in patients with diabetes
mellitus (Vincent et al., 2004; Malekirad
et al., 2011), so every intervention with minimum side effect that
reduce glucose level and oxidative stress markers can be effective for diabetes
complication prevention (Rains and Jain, 2011).
In response to high interest of patients with diabetes for using alternative
medicine, studies on antidiabetic herbs are increasing (Egede
et al., 2002; Karim et al., 2011).
Urtica dioica (Nettle) is one of the medical herbs. Several studies showed
beneficial effects of nettle in different disease such as rheumatoid arthritis
(Nourmohammadi et al., 2010), diabetes (Namazi
et al., 2011a), atherosclerosis (Chrubasika
et al., 2007; Namazi et al., 2011b) stomachache.
Some studies have shown antioxidant effects of nettle (Ozen
et al., 2003; Kanter et al., 2005;
Yener et al., 2009; Golalipour
and Khori, 2007; Bitiren et al., 2010; Mahmoud
et al., 2006; Odukoya et al., 2007).
It seems that the effects of Nettle on oxidative stress markers in patients
with type 2 diabetes have not studied, yet. So, the aim of this study was to
investigate the effects of hydro alcoholic extract of Nettle on oxidative stress
in patients with type 2 diabetes.
MATERIALS AND METHODS
A Randomized Double-blinded clinical trial was done on 50 patients (27 men,
23 women) with type 2 diabetes (T2DM) in Clinic of Diabetes in Sina Hospital
of Tabriz (Tabriz is one of the city in North-west of Iran). The inclusion criteria
for the trial were as follows: Both genders over the age of 30 years old, HbA1C
levels equal or less than 10%, common diabetes drugs usage (Metformin and Glibenclamide),
patients with triglyceride levels less than 400 mg dL-1. The exclusion
criteria included patients with cardiovascular, renal, liver or thyroid diseases,
infections, allergies, angina and the regular use of NSAIDs (Non-Steroid Anti
Inflammatory Drugs), warfarin, alcohol, herbal tea, dietary supplements and
Patients were informed about purpose of the study, each patient that is satisfied with participate in the study, signed an informed consent form, they were advised to continue their diet and physical activity habits without any changes during intervention.
After adjusting the patients by age, sex and duration of diabetes, they were randomly divided into intervention and control groups, they received 100 mg kg-1 of nettle extract or placebo in 3 portions a day, They dissolved each portion in 1 glass of water and drank after each 3 main meals for 8 weeks. Patients were contacted every week with telephone, they were asked for any compliance about nettle extract usage. Each two weeks, patients were asked to return any used bottles of extract and received new bottles. Biochemical, dietary record and physical activity were assessed at the beginning and end of the study. Eventually, forty five patients completed the study.
This research was approved by the Ethics committee and Human Studies review board of Iran-Tabriz University of Medical Sciences.
Extract specifications: Stinging Nettle certified by the Pharmacogenosy department in Iran-Gorgan University of Medical Science. Aerial parts of Nettle dried and powdered, extract was prepared with percolation method and ethanol (60°) was used. Final hydro alcoholic extract of Nettle contained 45% ethanol, 55% water and 2.7 g of dry matter in 1 L of extract. Water and alcohol percent in placebo was equal to Water and alcohol percent in Nettle extract, chlorophyll color added to placebo. There was not any difference in color between extract and placebo.
Participants were instructed to complete 24 h dietary recall for three days
(2 week days and 1 weekend day) at baseline and the end of study. These records
were used to calculate the habitual dietary energy and nutrient intake. International
Physical Activity (IPA) questionnaires (Hagstromera et
al., 2006) were filled out by researcher with face to face interview
at the baseline and the end of study. Weight and height were measured by standard
method, then BMI (Body Mass Index) was calculated by divided weight (kg) to
square of height (m).
Biochemical measurements: Five mililiter of blood was taken from forearm
vein after an overnight fast (12-14 h) at the beginning and end of the study.
Two mililiter of blood was collected in heparinized tubes for measurement of
GPX activity in erythrocyte and TAC. TAC measured by Ferric Reducing Antioxidant
Capacity (FRAP) method (Benzie and Strain, 1999).
Rest of blood sample was collected in tube containing EDTA, for measurement
of erythrocytes SOD. SOD and GPX were measured by commercially kit (Ransel and
Randox, UK) and by colorimetric method on an Autoanalyser. MDA level was used
as the (Idonije et al., 2011) indicator of lipid
peroxidation and were determined via reaction with thiobarbituric acid (TBA)
(Ahmed et al., 2006).
Statistical analysis: Data are analysed as Mean±Standard Deviation (SD). Kolmogorov-Smirnov test was used to determine normality of the data. Data with Abnormal distribution were converted to normal distribution by calculating logarithmic ratio. Then data at the end of study were compared to their own baseline values by Paired t-test. Comparison quantitative and qualitative variables between two groups was performed by Students t-test and Exact fisher test, respectively. SPSS version 18 (IBM Inc, USA) was used for data statistical analyses.assessment of dietary intake was done with Nutritionist IV software. The p<0.05 was considered significance for all variables.
The mean values of sex, age and duration of diabetes at the baseline, did not
show any statistical significant differences because of adjusting before dividing
patients into two groups. physical activity level showed no significant differences
in two groups at the baseline (p<0.05) (Table 1).
Comparison of two groups by BMI index showed that in case and control groups 9 and 11% of patient had normal BMI, 66 and 51% were overweight, respectively (Fig. 1). At the baseline, there were not any significant differences between groups. Also, there were not observed any statistical significant changes in BMI during the intervention.
||Characteristic of patients in intervention and control groups
|*: Mean±SD, : Independent samples t-test for
comparison of two groups at the baseline, ◊ : Exact fisher test
|| Comparison of Dietary Intake in two groups at the baseline
and the end
|*: Mean±SD, : Independent samples t-test for
comparison of two groups. p<0.05 considered as significant difference
between two groups
|| Comparison of antioxidant status and MDA in two groups at
the baseline and the end
|*: Mean±SD, : ANCOVA. p<0.05 considered as
Significant difference between two groups
||Comparison of patients with Body Mass Index (BMI) in two groups
at the baseline
Dietary intake showed significant difference in vitamin A and C levels between two groups at the baseline. For neutralizing their confounder effects, laboratory data were compared after adjusting two groups in vitamin A and C levels (Table 2).
Paired t-test showed that there were no changes in dietary intake and physical activity level during the study in both groups.
The antioxidant parameters and MDA showed significant differences at the baseline in two study groups (Table 3). Independent t-test showed statistical significant differences in SOD (1200±111 at the baseline, 1500±154 U mg-1 Hb at the end; p = 0.02) and TAC (1.51±0.34 at the baseline, 0.51±0.01 U m M Fe+2 L-1 at the end; p = 0.02) in intervention group compare to control group. But no statistically significant differences between two groups in MDA and GPX were seen (p>0.05).
This study showed that hydro alcoholic extract of Nettle increased TAC and
increased SOD, in type 2 diabetes patients after 8 weeks. But it could not change
GPX level and lipid peroxidant indicator (MDA). Results suggested that hydro
alcoholic extract of Nettle can protect body against free radicals and diabetes
Golalipour and Khori (2007) concluded that the hydroalcoholic
extract of nettle, by the present antioxidant compounds scavenged free radicals,
so could regenerate pancreatic beta cells. So, they suggested that nettle has
protective effect against oxidative stress in hyperglycemia rats.
Yener et al. (2009) studied on antioxidant effects
of nettle seeds on rats, they showed that nettle has protective effect on liver
against Aflatoxin, so they declared that nettle seed may have antioxidant properties.
Gulcin et al. (2004) showed that water extract
of nettle in 50, 100 and 250 μg mL-1 have stronger antioxidant
effects compare to alpha tocopherol in linoleic acid peroxidation. All of these
reviews are in support of the results of present study.
Toldy et al. (2005) showed that 30 mg kg-1
nettle decreased ROS (Reactive Oxygen Species). Another study showed that the
hydroalcoholic extract of nettle, decreased brain peroxidation more than 50%
and had inhibition effect on xanthine oxidase about 30%.
Ozen and Korkmaz (2003) with experiments on rats concluded
that hydroalcoholic extract of nettle has significant effect on antioxidant
enzymes such as catalase, SOD, GPX and glutathione Reductase. Present study
confirmed by Ozen and Korkmaz (2003) study about decreasing
effects of nettle on SOD, On the other hand Ozen results about effects of nettle
on GPX does not confirm the present study result. Dose, duration of diabetes,
amount and solvent (Samsam-shariat, 1980) type may cause
these differences in results of studies.
Studies have shown that flavonoids and carotenoids properties, in alcohol solvent
showed more Antioxidant characteristics than water solvent (Annegowda
et al., 2010). So, 45% ethanol in the present extract may caused
suitable background in improving antioxidative status.
Poly phenol compounds, are the most important part of the flavonoids family
(Boots et al., 2008), the polyphenols that are
found in nettles can be pointed to tannin, anthocyanin, chlorogenic acid and
cafe oil malic.these compounds can play antioxidant activity. Quercitin (3,
3, 4, 5, 7-penta hydroxy flavone), is dedicated to the greatest extent of flavonoids
in nettle. These properties may be one of the factor that caused antioxidative
characteristic of nettle in patients with type 2 diabetes (Pourmorad
et al., 2006).
More studies are suggested for determination of antioxidative effects of nettle in patients with diabetes by longer time intervention and larger sample size.
Hydro alcoholic extract of Nettle increased TAC and increased SOD Levels in
type 2 diabetes patients after 8 weeks intervention. So, it seems that hydro
alcoholic extract of Nettle can play a protective role from CVD in patients
with type 2 diabetes by improving Antioxidant status.
Authors would like to thanks the Traditional Medicine Association of Iran-Eastern Azerbaijan and Giah Esanse Company for preparing the hydro alcoholic extract of Nettle and placebo. This research was financially supported by Nutrition Research Center, Tabriz University of Medical Sciences, Iran. This study is a part of results of N. Namazi MSD dissertation.
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