Research Article Effect of the Combination of Gelam Honey ...downloads.hindawi.com/journals/bmri/2014/160695.pdf · Academic Editor: Paul W. Huber ... BIOKIMIA//YASMIN/-SEPTEMBER

Research ArticleEffect of the Combination of Gelam Honey and Ginger onOxidative Stress and Metabolic Profile inStreptozotocin-Induced Diabetic Sprague-Dawley Rats

Nur Fathiah Abdul Sani1 Levin Kesu Belani2

Chong Pui Sin2 Siti Nor Amilah Abdul Rahman2 Srijit Das3 Thent Zar Chi3

Suzana Makpol1 and Yasmin Anum Mohd Yusof1

1 Department of Biochemistry Faculty of Medicine Universiti Kebangsaan Malaysia Jalan Raja Muda Abdul Aziz53000 Kuala Lumpur Malaysia

2 Faculty of Medicine Universiti Kebangsaan Malaysia Medical Center (UKMMC) Jalan Yaacob Latif Cheras56000 Kuala Lumpur Malaysia

3 Department of Anatomy Faculty of Medicine Universiti Kebangsaan Malaysia Jalan Raja Muda Abdul Aziz53000 Kuala Lumpur Malaysia

Correspondence should be addressed to Yasmin AnumMohd Yusof rahmatyasminyahoocom

Received 26 February 2014 Accepted 25 March 2014 Published 16 April 2014

Academic Editor Paul W Huber

Copyright copy 2014 Nur Fathiah Abdul Sani et alThis is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in anymedium provided the originalwork is properly cited

Diabetic complications occur as a result of increased reactive oxygen species (ROS) due to long term hyperglycaemia Honey andginger have been shown to exhibit antioxidant activity which can scavenge ROS The main aim of this study was to evaluate theantioxidant and antidiabetic effects of gelam honey ginger and their combination Sprague-Dawley rats were divided into 2 majorgroups which consisted of diabetic and nondiabetic rats Diabetes was induced with streptozotocin intramuscularly (55mgkg bodyweight) Each group was further divided into 4 smaller groups according to the supplements administered distilled water honey(2 gkg body weight) ginger (60mgkg body weight) and honey + ginger Body weight and glucose levels were recorded weeklywhile blood from the orbital sinus was obtained after 3 weeks of supplementation for the estimation of metabolic profile glucosetriglyceride (TG) superoxide dismutase (SOD) catalase (CAT) glutathione peroxidase (GPx) reduced glutathione (GSH) oxidizedglutathione (GSSG) and malondialdehyde (MDA) The combination of gelam honey and ginger did not show hypoglycaemicpotential however the combination treatment reduced significantly (119875 lt 005) SOD and CAT activities as well as MDA levelwhile GSH level and GSHGSSG ratio were significantly elevated (119875 lt 005) in STZ-induced diabetic rats compared to diabeticcontrol rats

1 Introduction

Diabetes mellitus (DM) is characterized by chronic hypergly-caemia caused by defects in insulin secretion insulin actionor both resulting in impaired function in carbohydrate lipidand protein metabolism The total number of people withdiabetes globally is expected to rise from 171 million in 2000to 366 million by the year 2030 [1] In Malaysia the FirstNational Health and Morbidity Survey (NHMS I) conductedin 1986 reported a prevalence of diabetes of 63 and in theSecond National Health andMorbidity Survey (NHMS II) in

1996 this had risen to 83 [2] According to the third survey(NHMS III) the overall prevalence of diabetes is 116 inindividuals above 18 years and 149 in those above 30 years[3] Alarmingly the fourth survey conducted in 2011 (NHMSIV) showed that the prevalence of diabetes in individuals of18 years and above had risen to 152 [4]

According to American Diabetes Association diabetesmellitus (DM) is a group of metabolic diseases characterizedby hyperglycemia resulting from defects in insulin secretioninsulin action or both Type I DM or juvenile-onset diabetesis caused by an autoimmune destruction of beta cells of

Hindawi Publishing CorporationBioMed Research InternationalVolume 2014 Article ID 160695 9 pageshttpdxdoiorg1011552014160695

2 BioMed Research International

the pancreas causing no production of insulin Type II DMor adult onset diabetes involves a combination of reducedinsulin production with increased cell insulin resistance Thethird type of DM is known as gestational DM which occursduring pregnancy [5] Although the exact mechanism of howDM may cause oxidative stress is not known it is highlyprobable that long termhyperglycaemia causes autooxidationof glucose which will eventually lead to microvascular andmacrovascular complications [6] Oxidative stress resultsfrom the inability of endogenous antioxidants to neutralizethe abnormally high level of reactive oxygen species (ROS)which can react with and damage complex cellular moleculessuch as lipids proteins and DNA [6] This results in changesin the activity of endogenous antioxidant enzymes such assuperoxide dismutase (SOD) glutathione peroxidase (GPx)and malondialdehyde level (MDA) as well as DNA damagein DM patients [7 8] Antioxidant enzymes and dietarycompounds such as vitamins C E and flavonoids may havean effect in reducing the increased serum glucose levelin DM and secondary complications of neuropathy andnephropathy aswell asmacrovascular complications of strokeand cardiovascular problems in people with longstandinguncontrolled DM [9]

In the past decade rigorous research had been performedto find out the best way to overcome the complications asobserved in DM either by modern or alternative medicineIn Malaysia research on herbal medicine in alleviating DMusing animal and human studies has gained widemomentumin the past few years [10] A formulated polyherbal drugconsisting of extract of root of M paradisiaca seed of Tindica seed of E jambolana and leaf of C indica was foundto have a potent antidiabetogenic efficacy in STZ-induceddiabetic rats by reducing oxidative stress and carbohydratemetabolic profile and enzymes activities such glucose-6-phosphatase lactate dehydrogenase and hexokinase as wellas glucose-6-phosphate dehydrogenase in liver skeletal mus-cle and cardiac muscle towards the normal range [11] Aizzatet al [12] reported a reduction in lipid peroxidation asmeasured by MDA levels and reduction in DNA damageevaluated by Comet assay in STZ-induced diabetic rats whensupplemented with 150mgkg body weightChlorella vulgaris

For centuries honey has been used as a substitute forsugar as well as providing medicinal benefit In the pastdecade numerous biological and nutritional effects of honeyhave been reported such as antimicrobial antioxidant antivi-ral antiparasitic anti-inflammatory antimutagenic anti-cancer and immunosuppressive activities [13 14] In addi-tion Yao et al [15] showed that higher concentration of gelamhoney (5 gkg body weight) reduced oxidative damage betterthan lower concentration (25 gkg body weight) in youngand middle aged rats by modulating antioxidant enzymeactivities Ginger which is an underground rhizome of plantZingiber officinale belonging to the family Zingiberaceaewas reported to have hypoglycaemic anti-inflammatoryantitumorigenic and antioxidant effects in in vitro and in vivostudies [16ndash20] However to the best of our knowledge noprevious studies have explored the antidiabetic and antioxi-dant properties of the combination of gelamhoney and gingeron the oxidative stress and metabolic profile of STZ-induced

diabetic rats although a study by Patel et al [21] reportedthe synergistic effect of the combination of honey and gingeron the antibacterial property on isolates of extracted cariousteeth during orthodontic treatment Thus we embarked onthis project to evaluate whether the combination effect ofgelam honey and ginger would synergize in their actions inreducing oxidative stress and alleviating the complications ofdiabetes

2 Materials and Methods

21 Animals and Diet A total of 60 male Sprague-Dawleyrats were obtained from by Animal Care Unit UniversitiKebangsaan Malaysia (Bangi Malaysia) The studywas approved by the Animal Ethics Committee of theFaculty of Medicine Universiti Kebangsaan Malaysia (FPBIOKIMIA2012YASMIN26-SEPTEMBER463-SEPTEM-BER-2012-AUGUST-2013) Prior to the experiment ratsweighing 200ndash250 gm were randomly chosen They werehoused individually in polycarbonate cages at a temperatureof 22∘C with a 12-hour light-12-hour dark-cycle and wasquarantined for one week and fed with a standard labchow and water ad libitum The rats were divided into twogroups which were control (28 rats) and diabetic groups(32 rats) The control group was further divided into fourgroups (i) control group fed with normal lab chow (ii)control group fed with gelam honey (20 gkg body weight)(iii) control group fed with ginger (60mgkg body weight)and (iv) control group fed with gelam honey and ginger(20 gkg bodyweight honey + 60mgkg bodyweight ginger)Diabetes was induced by a single intramuscular injectionof streptozotocin (STZ) (55mgkg body weight) STZ wasprepared by dissolving it in cold 09 NaCl It was injectedinto overnight fasting rats through the thigh muscle bulkThe rats were observed closely for symptoms of DM such aspolydipsia polyuria and weight loss Blood samples wereobtained from ratsrsquo tail vein 48 hours after STZ injection Ifblood glucose level was gt14mmolL rats were considereddiabeticThe diabetic rats were then divided into four groupsthat is (i) STZ-induced diabetic rats fed with normal chowfood (ii) STZ-induced diabetic rats fed with gelam honey(20 gkg body weight) (iii) STZ-induced diabetic rats fedwith ginger (60mgkg body weight) and (iv) STZ-induceddiabetic rats fed with gelam honey and ginger The bloodglucose level was taken once weekly starting from week 0 toweek 3 After 3 weeks blood samples were obtained fromperiorbital sinus vein for the evaluation of metabolic andoxidative markers such as blood glucose triglyceride (TG)superoxide dismutase (SOD) glutathione peroxidase (GPx)catalase (CAT) activities glutathione peroxidase (GPx)reduced glutathione (GSH) oxidized glutathione (GSSG)and malondialdehyde (MDA)

22 Ginger Extract by Hot Compressed Water Extraction(HCW) An amount of 75 gm of ground ginger was weighedbefore being loaded into a covered stainless steel meshcylinder and placed into the extraction cell of fabricated hot

BioMed Research International 3

compressed water extraction (HCW) equipment Approxi-mately 700mLof distilledwaterwas added into the extractioncell and was covered securely with stainless steel lid N

2gas

was then used to purge out air and dissolved oxygen gas fromthe cell by letting the gas passing through the cell and outusing release valve The temperature was set up according tothe design of experiment from 100 to 200∘C by increasing10∘C at each experiment in 30min of extraction time Allexperiments were carried out at constant pressure of 35MPaOnce the extraction process was completed the extractantwas immediately transferred into the cooling cell Gingerextract was collected and stored at 4∘C

23 Honey Malaysian monofloral gelam honey is producedby Apis mellifera bees and the major nectar and pollencollected by the bees is from the plant Melaleuca cajuputiPowell which is known locally as the ldquogelam treerdquo It wasprovided by the National Apiary Department of AgricultureBatu Pahat Johor Malaysia

24 Preparation of Blood Sample for Enzyme Assays Bloodobtained from the periorbital sinus vein was collected in testtubes with heparin to prevent blood coagulation and theplasma was separated The blood samples were then rinsedwith the same volume of 09 normal saline (NaCl) andcentrifuged at 3000 rpm for 10min at 4∘C The upper layerwas removed and the above procedure was repeated with09 NaCl until it became clear The lower layer termedhemolysate was then used for the antioxidant enzyme assaysTheupper layer termed supernatant was used for lipid profiletest

25 Determination of Body Weight The rats were weighedweekly from the start of the study (ie at the end of oneweek of acclimatization to the laboratory environment) witha laboratory scale (Harvard Trip Balance Florham Park NJUSA) to the nearest gram

26 Determination of Glucose Level Fasting blood glucoselevel was estimated by glucometer (Accuchek Instant Ger-many) using a drop of blood taken from the tail vein of ratsGlucose estimation was made weekly throughout the periodof study

27 Triglyceride (TG) Level Measurement Blood triglyceridelevel of animals was estimated by Reflotron (Boehringer-Mannheim Germany) using plasma from the processedblood The fresh whole blood taken from the periorbitalsinus of rats was centrifuged at 3000 rpm 4∘C for 10minThe plasma was then obtained from the processed blood Anamount of 30120583L of plasmawas placed on the Roche ReflotronTriglyceride strip and then placed into the Reflotronmachineto be analysed The amount of triglyceride is expressed asmmolL

28 Superoxide Dismutase (SOD) Activity MeasurementSuperoxide dismutase activity was measured according tothe method of Beyer Jr and Fridovich [22] Briefly 10mL

aliquot of a mixture containing 01mM phosphate buffer pH78 57120583M nitro blue tetrazolium (Sigma St Louis USA)99mM L-methionine (Sigma St Louis USA) and 0025Triton-X (Sigma St Louis USA) was pipetted into test tubesThen 20120583L of hemolysate and 10 120583L of a solution containing44mg100mL riboflavin (Sigma St Louis USA) were addedinto the mixture The tubes were illuminated for 7min inan aluminium foil-lined box containing two 20-W SylvaniaGroLux fluorescent lamps Absorbance was thenmeasured ata wavelength of 560 nm One unit of SOD was defined as theamount of enzyme required to inhibit nitro blue tetrazoliumreduction by 50 per min per mL hemolysate Enzymeactivity was expressed as units per mg of Hb (Umg Hb)

29 Glutathione Peroxidase (GPx) Activity MeasurementGPx was determined by the method developed by Paglia andValentine [23] The reaction mixture contained 005M phos-phate buffer pH 70 84mMNADPH (Sigma St Louis USA)1125M sodium azide (Hopkin amp William England) 5mMreduced glutathione (GSH) NADPH (Sigma St Louis USA)and 3UmL glutathione reductase (Sigma St Louis USA)The hemolysate was prepared by adding an equal volume ofdistilled water to the RBC pellet and was allowed to standfor 1 h at 4∘C Then four parts by volume of distilled waterwere added Finally double strength Drabkinrsquos reagent (EagleDiagnostics Japan) was added to yield the final hemolysateThe reaction was initiated by adding 01mL of 22mM H

2O2

(Merck Darmstadt German) The conversion of NADPH toNADP+ was followed by measuring the change in ODminat 340 nm One unit of GPx was defined as the amount ofenzyme required to oxidize 1 120583mol NADPHmin per mLhemolysate Enzyme activity was expressed as milliunits permg of Hb (mUmgHb)

210 Catalase (CAT)ActivityMeasurement CATwas assayedby the method of Aebi [24] The reaction mixture consistedof 50mM phosphate buffer pH 70 and 30mM hydrogenperoxide (H

2O2) An amount of 0025mL of haemolysate was

added with 125mL of phosphate buffer to make a solutionwith 500x dilutionThe reactionwas startedwith the additionof 10mL 30mM H

2O2to 20mL of hemolysate and the

absorbance at 240 nmwas read at room temperature against ablank containing 20mL haemolysate plus 10mL phosphatebufferOne unit of catalase enzymewas defined as the amountof enzyme which liberates half the peroxide oxygen fromH2O2solution in 30 s at room temperature Enzyme activity

was expressed as units permg ofHb (UmgHb) Hemoglobinin the hemolysate was measured by using Hemoglobin kit(Eagle Diagnostics Japan)

211 Total Glutathione (TGSH) Reduced Glutathione (GSH)Level and Oxidized Glutathione (GSSG) Level MeasurementTGSH GSH and GSSG were determined by the method ofTietze [25] and Griffith [26] with some modifications Forthe measurement of total TGSH content 30 120583L of plasmawas incubated with 30 120583L of 551015840-dithiobis(2-nitrobenzoicacid) (DTNB) (Sigma St Louis USA) phosphate buffercontaining EDTA glutathione reductase (250 unitsmL) and


NADPH Optical densities of the chromophores formedweremonitored at 405 nm for 4 minutes For GSSG assay 100 120583Lof plasma is derivatized by adding 2 120583L of 2-vinylpyridine(Sigma St Louis USA) and incubated for 30min at 25∘CThe assay was performed in a similar manner as totalGSH equivalent Protein concentrationwas determined usingthe Bradford Protein Assay Kit (Bio-Rad USA) Reducedglutathione (GSH) was calculated by using the formula

TGSH equivalent = 2 GSSG + GSH (1)

212 Malondialdehyde (MDA) Level Measurement Plasmamalondialdehyde (MDA) was determined using high per-formance liquid chromatography (HPLC) with photo diodearray detector (Shimadzu Japan) as described by Pilz et al[27] with some modifications Briefly samples (50120583L) weremixed with 200120583L of 13M NaOH and incubated at 60∘Cfor 60min After cooling the mixture 100 120583L of 35 HCIO

4

was added and centrifuged at 10000 g for 10min at 4∘CThe supernatant of the samples (300 120583L) was transferred into15mL of HPLC tube 50 120583L of 5mM DNPH solution wasadded into the mixture and incubated for 30min at roomtemperature Then 40 120583L samples were injected into theHPLCThe amount of MDA is expressed as concentration ofMDA in nmol per mL plasma

213 Statistical Analysis Data was analysed using SPSS pack-age (version 21) Results refer to mean plusmn SD Statisticalevaluation was assessed using analysis of variance (ANOVA)A 119875 lt 005 was considered significant

3 Results

31 Effect of Gelam Honey Ginger and Their Combinationon Body Weight in STZ-Induced Diabetic Rats As shownin Table 1 diabetic control rats showed reduction in bodyweight as compared to normal control rats which furtherdecreased during the experimental period Treatment withgelam honey ginger and their combination did not improvethe body weight after 3 weeks

32 Effect of Gelam Honey Ginger andTheir Combination onBlood Glucose Level in STZ-Induced Diabetic Rats All ratsinjected with STZ developed severe diabetes as indicated byserum glucose concentrations gt14mmolL over the period of3 weeks The diabetic control rats had significantly elevatedblood glucose level compared to normal control rats at theend of week 3 Treatment of the diabetic rats with honeyginger or their combination did not significantly reduce theglucose levels when comparedwith the diabetic control group(Figure 1)

33 Effect of Gelam Honey Ginger and Their Combinationon Triglyceride (TG) Level in STZ-Induced Diabetic RatsThe effect of gelam honey ginger and their combination ontriglyceride (TG) level in STZ-induced diabetic rat is shownin Figure 2 There was a significantly (119875 lt 005) higher levelof TG in the diabetic group treated with single treatment of

0050

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Figure 1 Blood glucose level after 3 weeks of treatment in normaland STZ-induced diabetic rats with various treatments Data areexpressed as mean plusmn SD a significant (119875 lt 005) compared tonormal control b significant (119875 lt 005) compared to normal +gelam honey c significant (119875 lt 005) compared to normal + gingerd significant (119875 lt 005) compared to normal combination

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Figure 2 Blood triglyceride level after 3 weeks of treatment innormal and STZ-induced diabetic rats with various treatmentsData are expressed as mean plusmn SD a significant (119875 lt 005)compared to normal control b significant (119875 lt 005) comparedto normal + gelam honey c significant (119875 lt 005) compared tonormal + ginger d significant (119875 lt 005) compared to normalcombination e significant (119875 lt 005) compared to diabetic controlf significant (119875 lt 005) compared to diabetic + gelam honey

honey and ginger however combination of ginger and honeysignificantly (119875 lt 005) reduced the TG level compared tosingle treatment in diabetic rats

34 Effect of Gelam Honey Ginger andTheir Combination onSuperoxide Dismutase (SOD) Level in STZ-Induced DiabeticRats No significant change in the SOD activity was observedbetween normal control and diabetic control group Incontrast the activity of SOD enzyme decreased significantly


Table 1 Effects of honey ginger and the combination of honey and ginger on body weight (g) in STZ-induced diabetic rats

Groups Body weight in gms (mean plusmn SD)0 week 1st week 2nd week 3rd week

Normal Control 2276 plusmn 107 2702 plusmn 230a 2957 plusmn 267ab 3010 plusmn 307ab

Normal + Honey 2311 plusmn 138 2720 plusmn 141a 2911 plusmn 161ab 3063 plusmn 158ab

Normal + Ginger 2112 plusmn 203 2551 plusmn 311 2716 plusmn 371b 2908 plusmn 374abc

Normal + Combination 2261 plusmn 174 2694 plusmn 115 2876 plusmn 131ab 3042 plusmn 171bc

Diabetic Control 2248 plusmn 85 2099 plusmn 173a 2140 plusmn 296ab 2059 plusmn 525Diabetic + Honey 2309 plusmn 42 2176 plusmn 162a 2142 plusmn 220a 2139 plusmn 323a

Diabetic + Ginger 2337 plusmn 89 2030 plusmn 316a 1957 plusmn 303ab 1880 plusmn 319abc

Diabetic + Combination 2383 plusmn 113 2137 plusmn 181 2042 plusmn 273 2022 plusmn 380a

Data are expressed as mean plusmn SD aSignificant different (119875 lt 005) when compared to week 0 bsignificant different (119875 lt 005) when compared to week 1csignificant different (119875 lt 005) when compared to week 2

(119875 lt 005) in the diabetic rats treated with ginger andcombination of gelam honey and ginger as compared tonormal rats and diabetic control group (Figure 3)

35 Effect of Gelam Honey Ginger andTheir Combination onCatalase (CAT) Level in STZ-Induced Diabetic Rats Diabeticrats had significantly (119875 lt 005) higher levels of CAT activitycompared to the nondiabetic rats with treatment Howeverthe diabetic group treated with gelam honey ginger andtheir combination managed to show a significant (119875 lt 005)reduction in CAT activity when compared with the diabeticcontrol and normal control group (Figure 4)

36 Effect of Gelam Honey Ginger andTheir Combination onGlutathione Peroxidase (GPx) Level in STZ-Induced DiabeticRats Diabetic rats showed a significantly (119875 lt 005)higher GPx activity when compared to the nondiabetic ratsTreatment of diabetic rats with gelam honey ginger and theircombination did not produce any significant effects on GPxactivity compared to normal control and diabetic controlgroup (Figure 5)

37 Effect of Gelam Honey Ginger and Their Combinationon Levels of Reduced Glutathione (GSH) and on GSHGSSGRatio in STZ-Induced Diabetic Rats Diabetic rats showed asignificantly (119875 lt 005) lower GSH content and GSHGSSGratio as compared to the nondiabetic rats Combination treat-ment of gelam honey and ginger increased GSH content andGSHGSSG ratio significantly (119875 lt 005) when compared tothe diabetic control group (Figure 6)

38 Effect of Gelam Honey Ginger andTheir Combination onMalondialdehyde (MDA) Level in STZ-Induced Diabetic RatsThe result demonstrated that MDA level is significantly (119875 lt005) increased in diabetic rats as compared to nondiabeticrats However the diabetic rats that received a combinationof gelam honey and ginger exhibited a significant (119875 lt 005)decrease inMDA concentration compared to diabetic controlrats (Figure 7)

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Figure 3 Blood superoxide dismutase (SOD) level after 3 weeksof treatment in normal and STZ-induced diabetic rats with varioustreatments Data are expressed as mean plusmn SD a significant (119875 lt005) compared to normal control e significant (119875 lt 005)compared to diabetic control

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Figure 4 Blood catalase (CAT) level after 3 weeks of treatment innormal and STZ-induced diabetic rats with various treatments Dataare expressed as mean plusmn SD a significant (119875 lt 005) comparedto normal control b significant (119875 lt 005) compared to normal+ gelam honey c significant (119875 lt 005) compared to normal +ginger d significant (119875 lt 005) compared to normal combinatione significant (119875 lt 005) compared to diabetic control


002040608

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Figure 5 Blood glutathione peroxidase (GPx) level after 3 weeksof treatment in normal and STZ-induced diabetic rats with varioustreatments Data are expressed as mean plusmn SD a significant (119875 lt005) compared to normal control b significant (119875 lt 005)compared to normal + gelam honey c significant (119875 lt 005)compared to normal + ginger d significant (119875 lt 005) comparedto normal combination

4 Discussion

The role of oxidative stress in the pathogenesis and com-plications of diabetes mellitus is well recognized [28ndash31]Several studies have reported that persistent hyperglycaemiacan cause high production of ROS which may lead tocellular oxidative damage including DNA lipids and protein[32ndash35] Thus reducing the blood glucose level in DMis important to decrease the production of ROS as wellas to alleviate complications of DM as a result of tissuesdamage Omotayo et al [36] showed that honey exerts ahypoglycemic effect and ameliorates the oxidative stress inkidneys of STZ induced diabetic rats after following fourweeks of treatment while Ojewole [37] found that gingerexhibits similar hypoglycaemic properties under normal anddiabetic condition The ability of ginger to reduce the bloodglucose level in STZ-induced DM rats was also reported byMorakinyo et al [38] after six weeks of treatment Howeverin the present study we did not observe any hypoglycaemiceffect of honey ginger and their combination in diabeticrats over the three weeks of treatment This may be due tothe shorter duration of treatment adopted in this study Itwas also noted that TG levels were high in diabetic controland diabetic treatment groups showing that neither honeynor ginger had hypolipidemic effect on diabetic rats Ourresults are contradictory to the study by Akhani et al [16]and Al-Amin et al [39] who reported that ginger extract wassignificantly effective in lowering glucose cholesterol andTGlevels in STZ-induced diabetic rats compared with controldiabetic rats Perhaps this can be explained by excessivelyhigh glucose levels that are known to convert to TG duringcarbohydrate metabolism [40]

Reactive oxygen species (ROS) such as hydroxyl radi-cal (OH∙) superoxide anion (O

2

∙) and hydrogen peroxide(H2O2) are naturally generated in the body during normal

metabolism They are neutralized by the endogenous antiox-idant enzymes such as SOD CAT and GPx to preventoxidative stress [41] However overproduction of ROS inDM leads to imbalance between oxidants and antioxidantsresulting in the development of complications of DM suchas diabetic neuropathy and nephropathy [8] Exogenousantioxidants such as ginger honeyChlorella vulgaris vitaminE and vitamin Cmay help in eradicating the large amount ofROS in DM [13 21 37 42]

The antioxidant defence system enzymes SOD CAT andGPx are increased in DM because of excessive levels ofglucose in mitochondria resulting in overdrive of electrontransport chain producing excess free radicals [43] Severalstudies have shown the ability of honey and ginger tomodulate antioxidant enzymes activities in DM Omotayo etal [36] showed that SOD activity was decreased in the STZ-induced diabetic rats treated with honey In the present studyboth honey and ginger were able to reduce the SOD and CATactivities significantly in STZ-induced diabetic rats whencompared to diabetic rats without treatment Interestinglywe found that the diabetic rats treated with combination ofhoney and ginger showed lower SOD and CAT activitieswhen compared to diabetic group treated with honey andginger aloneThe ability of honey to prevent oxidative damagein diabetic rats might be due to its phenolic antioxidantscontent or through the action of CAT activity found in honeyin reducing hydrogen peroxide Ginger extract possessesantioxidative characteristic by scavenging superoxide anionand hydroxyl radicals [44]We also observed a nonsignificantreduction in SOD activity in diabetic group compared tonormal control group

Glutathione is the major endogenous antioxidant pro-duced by the cells participating directly in the neutralizationof free radicals and reactive oxygen compounds [45] Inhealthy cells and tissues most of the total glutathione arein the reduced form (GSH) and less than 10 exists inthe oxidized form (GSSG) The ratio of GSH to GSSG isa sensitive indicator of oxidative stress or overall health[46] The results of the present study showed that GSHlevel and GSHGSSG ratio were decreased in diabetic ratsThis significant depletion of GSH level and GSHGSSGratio in diabetic rats would indicate its increased utilizationagainst reactive oxygen species generated in diabetic rats [47]Although treatment with gelam honey alone did not produceany effect on GSH level or GSHGSSG ratio ginger extracthowever managed to increase GSH level and GSHGSSGratio back to normal level while the combination treatmentshowed significant increase above normal level Thus ourfindings suggest that combination of honey and ginger mayoffer better antioxidant effect by scavenging free radicalsand restoring the imbalance between oxidantantioxidanthomeostasis developed during diabetic condition

Malondialdehyde (MDA) is generated from the degrada-tion of polyunsaturated lipids by ROS It is one of the mostfrequently used indicators of lipid peroxidation Omotayo etal [36] had demonstrated that elevated levels of MDA inSTZ-induced diabetic rats were reduced after the treatmentwith honey Similarly our study showed that the diabeticrats which received gelam honey in combination with ginger


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betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(b)

Figure 6 (a) Plasma GSH level and (b) ratio of GSHGSSG after 3 weeks of treatment in normal and STZ-induced diabetic rats with varioustreatments Data are expressed as mean plusmn SD a significant (119875 lt 005) compared to normal control b significant (119875 lt 005) compared tonormal + gelam honey c significant (119875 lt 005) compared to normal + ginger d significant (119875 lt 005) compared to normal combinatione significant (119875 lt 005) compared to diabetic control f significant (119875 lt 005) compared to diabetic + gelam honey g significant (119875 lt 005)compared to diabetic + ginger

0

10

20

30

40

50

60

Mal

onde

alde

hyde

elev

el (n

mol

L)

Malondialdehyde (MDA) abca

bcabcde

abcd

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

Figure 7 Plasma malondialdehyde (MDA) level after 3 weeks oftreatment in normal and STZ-induced diabetic rats with varioustreatments Data are expressed as mean plusmn SD a significant (119875 lt005) compared to normal control b significant (119875 lt 005)compared to normal + gelam honey c significant (119875 lt 005)compared to normal + ginger d significant (119875 lt 005) compared tonormal combination e significant (119875 lt 005) compared to diabeticcontrol

showed significant reduction in lipid peroxidation (MDA)However gelam honey and ginger alone may not offerprotection against lipid peroxidative damage in diabetic rats

5 Conclusion

DM is a common chronic disease which can cause multiplecomplications if the glucose level is uncontrolled This is

due to the oxidative stress which can cause changes in theendogenous antioxidants Our present study showed that thecombination of gelam honey and ginger provided a betterantioxidant effect as compared to gelam honey or gingeralone as evidenced by the significantly reduced SOD andCAT activities depletedMDA level increasedGSH level andincreased GSHGSSG ratio in diabetic rats Further researchwith longer duration of treatment with ginger and honeywould perhaps give better insight to the changes in thebiochemical profile related to diabetic complications Thecorrect combination of gelam honey and ginger may offer apotential adjuvant to antidiabetic medications to reduce theoxidative stress and complications of DM

Conflict of Interests

The authors declare that they have no conflict of interests

Authorsrsquo Contribution

Yasmin Anum Mohd Yusof Suzana Makpol Srijit Das andThent Zar Chi were responsible for the study conceptionand design Nur Fathiah Abdul Sani Levin Kesu BelaniChong Pui Sin and Siti Nor Amilah Abdul Rahman wereresponsible for acquisition of data Nur Fathiah Abdul SaniLevinKesu Belani Chong Pui Sin and Siti NorAmilahAbdulRahman were responsible for analysis and interpretation ofdata Drafting of manuscript was by Nur Fathiah Abdul SaniLevinKesu Belani Chong Pui Sin and Siti NorAmilahAbdulRahman Critical revision was by YasminAnumMohdYusofSuzana Makpol and Srijit Das


Acknowledgments

The authors would like to express sincere gratitude to MohdRazman Mohd Noor and Ahmad Fais Abd Rasid for theirexpertise in collecting blood samples from periorbital sinusvein Fazaine Zakaria for her assistance in enzyme assayand Siti Nor Asyikin Zakaria for her technical assistance inusing HPLC This study was financially supported by specialstudy Grant sanctioned by the Faculty of Medicine Univer-siti Kebangsaan Malaysia (FF-073-2013) and the UniversityResearch Grant (GUP-2012-016)

References

[1] S Wild G Roglic A Green R Sicree and H King ldquoGlobalPrevalence of Diabetes estimates for the year 2000 and projec-tions for 2030rdquoDiabetes Care vol 27 no 5 pp 1047ndash1053 2004

[2] B M Mafauzy ldquoDiabetes mellitus in MalaysiardquoMedical Journalof Malaysia vol 61 no 4 pp 397ndash398 2006

[3] G R LetchumanWMWanNazaimoonW BWanMohamadet al ldquoPrevalence of diabetes in the Malaysian National HealthMorbidity Survey III 2006rdquo The Medical Journal of Malaysiavol 65 no 3 pp 180ndash186 2010

[4] ldquoNational Health and Morbidity Surveyrdquo Fact Sheet 2011[5] ldquoDiagnosis and Classification of Diabetes Mellitus American

Diabetes Associationrdquo Diabetes Care supplement 1 no 272004

[6] F A Matough S B Budin Z A Hamid N Alwahaibi and JMohamed ldquoThe role of antioxidants in diabetic complicationsrdquoSQU Medical Journal vol 12 no 1 pp 5ndash18 2012

[7] P Martın-Gallan A Carrascosa M Gussinye and CDomınguez ldquoBiomarkers of diabetes-associated oxidativestress and antioxidant status in young diabetic patients with orwithout subclinical complicationsrdquo Free Radical Biology andMedicine vol 34 no 12 pp 1563ndash1574 2003

[8] J S Johansen A KHarris D J Rychly andA Ergul ldquoOxidativestress and the use of antioxidants in diabetes linking basicscience to clinical praticerdquo Cardiovascular Diabetology vol 4article 5 2005

[9] D Bonnefont-Rousselot ldquoThe role of antioxidant micronutri-ents in the prevention of diabetic complicationsrdquo Treatments inEndocrinology vol 3 no 1 pp 41ndash52 2004

[10] P O Cheow C L Seng and A H Tengku ldquoA review ofclinical trials in emerging botanical interventions for type 2diabetes mellitusrdquo in Topics in the Prevention Treatment andComplications of Type II Diabetes Mellitus 2011

[11] K Chatterjee K M Ali C Mallick and D Ghosh ldquoAntihyper-glycaemic antioxidative activities of a formulated polyherbaldrugMTEC (modified) in streptozotocin-induced diabetic ratrdquoJournal of Medicinal Plant Research vol 3 no 6 pp 468ndash4802009

[12] O Aizzat S W Yap H Sopiah et al ldquoModulation of oxidativestress by Chlorella vulgaris in streptozotocin (STZ) induceddiabetic Sprague-Dawley ratsrdquo Advances in Medical Sciencesvol 55 no 2 pp 281ndash288 2010

[13] S Bogdanov T Jurendic R Sieber and P Gallmann ldquoHoney fornutrition and health a reviewrdquo Journal of the American Collegeof Nutrition vol 27 no 6 pp 677ndash689 2008

[14] S Z Hussein K M Yusoff S Makpol and Y A M YusofldquoGelam honey attenuates carrageenan-induced rat paw inflam-mation via NF-120581B pathwayrdquo PloS ONE vol 8 no 8 Article IDe72365 2013

[15] L K Yao S L A RazakN Ismail et al ldquoMalaysian gelamhoneyreduces oxidative damage and modulates antioxidant enzymeactivities in young and middle aged ratsrdquo Journal of MedicinalPlant Research vol 5 no 23 pp 5618ndash5625 2011

[16] S P Akhani S L Vishwakarma and R K Goyal ldquoAnti-diabeticactivity of Zingiber officinale in streptozotocin-induced type Idiabetic ratsrdquo Journal of Pharmacy and Pharmacology vol 56no 1 pp 101ndash105 2004

[17] N Ahmad S Sulaiman N A Mukti N A Murad N A AHamid and Y A M Yusof ldquoEffects of ginger extract (Zingiberofficinale Roscoe) on antioxidant status of hepatocarcinomainduced ratsrdquoMalaysian Journal of Biochemistry and MolecularBiology vol 14 pp 7ndash12 2006

[18] S H M Habib S Makpol N A A Hamid S Das W Z WNgah and Y A M Yusof ldquoGinger extract (Zingiber officinale)has anti-cancer and anti-inflammatory effects on ethionine-induced hepatoma ratsrdquoClinics vol 63 no 6 pp 807ndash813 2008

[19] S Abdullah S A Z Abidin N A Murad S Makpol W Z WNgah and Y A M Yusof ldquoGinger extract (Zingiber officinale)triggers apoptosis and G0G1 cells arrest in HCT 116 and HT29 colon cancer cell linesrdquo African Journal of BiochemistryResearch vol 4 pp 134ndash142 2010

[20] HHarliansyahNAMuradWZWNgah andYAMYusofldquoAntiproliferative antioxidant and apoptosis effects of ZingiberOfficenale and 6-Gingerol on HepG2 cellsrdquo Asian Journal ofBiochemistry vol 2 no 6 pp 421ndash426 2009

[21] R V Patel V T Thaker and V K Patel ldquoAntimicrobial activityof ginger and honey on isolates of extracted carious teethduring orthodontic treatmentrdquo Asian Pacific Journal of TropicalBiomedicine vol 1 no 1 pp S58ndashS61 2011

[22] W F Beyer Jr and I Fridovich ldquoAssaying for superoxidedismutase activity some large consequences of minor changesin conditionsrdquo Analytical Biochemistry vol 161 no 2 pp 559ndash566 1987

[23] D E Paglia and W N Valentine ldquoStudies on the quantitativeand qualitative characterization of erythrocyte glutathione per-oxidaserdquo The Journal of Laboratory and Clinical Medicine vol70 no 1 pp 158ndash169 1967

[24] H Aebi ldquoCatalase in vitrordquoMethods in Enzymology vol 105 pp121ndash126 1984

[25] F Tietze ldquoEnzymic method for quantitative determination ofnanogram amounts of total and oxidized glutathione appli-cations to mammalian blood and other tissuesrdquo AnalyticalBiochemistry vol 27 no 3 pp 502ndash522 1969

[26] O W Griffith ldquoDetermination of glutathione and glutathionedisulfide using glutathione reductase and 2-vinylpyridinerdquoAnalytical Biochemistry vol 106 no 1 pp 207ndash212 1980

[27] J Pilz I Meineke and C H Gleiter ldquoMeasurement of free andbound malondialdehyde in plasma by high-performance liquidchromatography as the 24-dinitrophenylhydrazine derivativerdquoJournal of Chromatography B Biomedical Sciences and Applica-tions vol 742 no 2 pp 315ndash325 2000

[28] D Giugliano A Ceriello and G Paolisso ldquoOxidative stress anddiabetic vascular complicationsrdquo Diabetes Care vol 19 no 3pp 257ndash267 1996

[29] J W Baynes and S R Thorpe ldquoThe role of oxidative stress indiabetic complicationsrdquo Current Opinion in Endocrinology andDiabetes vol 3 no 4 pp 277ndash284 1996


[30] R A Simmons ldquoDevelopmental origins of diabetes the role ofoxidative stressrdquo Free Radical Biology and Medicine vol 40 no6 pp 917ndash922 2006

[31] A C Maritim R A Sanders and J B Watkins III ldquoDiabetesoxidative stress and antioxidants a reviewrdquo Journal of Biochem-ical and Molecular Toxicology vol 17 no 1 pp 24ndash38 2003

[32] M R Bennett ldquoReactive oxygen species and death oxidativeDNA damage in atherosclerosisrdquo Circulation Research vol 88no 7 pp 648ndash650 2001

[33] D A Slatter C H Bolton and A J Bailey ldquoThe importance oflipid-derived malondialdehyde in diabetes mellitusrdquoDiabetolo-gia vol 43 no 5 pp 550ndash557 2000

[34] J Cederberg S Basu and U J Eriksson ldquoIncreased rate oflipid peroxidation and protein carbonylation in experimentaldiabetic pregnancyrdquo Diabetologia vol 44 no 6 pp 766ndash7742001

[35] J Blasiak M Arabski R Krupa et al ldquoDNA damage and repairin type 2 diabetes mellitusrdquo Mutation Research Fundamentaland Molecular Mechanisms of Mutagenesis vol 554 no 1-2 pp297ndash304 2004

[36] E O Omotayo S Gurtu S A Sulaiman M S A WahabK N S Sirajudeen and M S M Salleh ldquoHypoglycemic andantioxidant effects of honey supplementation in streptozotocin-induced diabetic ratsrdquo International Journal for Vitamin andNutrition Research vol 80 no 1 pp 74ndash82 2010

[37] J A O Ojewole ldquoAnalgesic antiinflammatory and hypogly-caemic effects of ethanol extract of Zingiber officinale (Roscoe)rhizomes (Zingiberaceae) in mice and ratsrdquo PhytotherapyResearch vol 20 no 9 pp 764ndash772 2006

[38] A O Morakinyo A J Akindele and Z Ahmed ldquoModulationof antioxidant enzymes and inflammatory cytokines possiblemechanism of anti-diabetic effect of ginger extractsrdquo AfricanJournal of Biomedical Research vol 14 pp 195ndash202 2011

[39] Z M Al-Amin M Thomson K K Al-Qattan and M AlildquoAnti-diabetic and hypolipidaemic properties of ginger (Zin-giber officinale) in streptozotocin-induced diabetic ratsrdquo BritishJournal of Nutrition vol 96 no 4 pp 660ndash666 2006

[40] M A Lieberman and A MarksMarkrsquos Basic Medical Biochem-istry Linppincott Williams ampWilkins 4th edition 2012

[41] D J Betteridge ldquoWhat is oxidative stressrdquoMetabolism Clinicaland Experimental vol 49 no 2 pp 3ndash8 2000

[42] J Cederberg C M Siman and U J Eriksson ldquoCombinedtreatment with vitamin E and vitamin C decreases oxidativestress and improves fetal outcome in experimental diabeticpregnancyrdquo Pediatric Research vol 49 no 6 pp 755ndash762 2001

[43] O Erejuwa ldquoOxidative stress in diabetes mellitus is there arole for hypoglycemic drugs andor antioxidantsrdquo inOxidativeStress and Diseases V Lushchak Ed InTech

[44] T PKrishnakantha andB R Lokesh ldquoScavenging of superoxideanions by spice principlesrdquo Indian Journal of Biochemistry andBiophysics vol 30 no 2 pp 133ndash134 1993

[45] R W Scholz K S Graham E Gumpricht and C C ReddyldquoMechanism of interaction of vitamin E and glutathione in theprotection against membrane lipid peroxidationrdquo Annals of theNew York Academy of Sciences vol 570 pp 514ndash517 1989

[46] A Meister and M E Anderson ldquoGlutathionerdquo Annual Reviewof Biochemistry vol 52 pp 711ndash760 1983

[47] M Rastogi R Ojha G V Rajamanickam A Agrawal AAggarwal andG PDubey ldquoCurcuminoidsmodulates oxidativedamage and mitochondrial dysfunction in diabetic rat brainrdquoFree Radical Research vol 42 no 11-12 pp 999ndash1005 2008

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of


Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology


Molecular Biology International

GenomicsInternational Journal of


The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014


BioinformaticsAdvances in

Marine BiologyJournal of



Signal TransductionJournal of


BioMed Research International

Evolutionary BiologyInternational Journal of



Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014


Genetics Research International


Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational



Enzyme Research



Microbiology


the pancreas causing no production of insulin Type II DMor adult onset diabetes involves a combination of reducedinsulin production with increased cell insulin resistance Thethird type of DM is known as gestational DM which occursduring pregnancy [5] Although the exact mechanism of howDM may cause oxidative stress is not known it is highlyprobable that long termhyperglycaemia causes autooxidationof glucose which will eventually lead to microvascular andmacrovascular complications [6] Oxidative stress resultsfrom the inability of endogenous antioxidants to neutralizethe abnormally high level of reactive oxygen species (ROS)which can react with and damage complex cellular moleculessuch as lipids proteins and DNA [6] This results in changesin the activity of endogenous antioxidant enzymes such assuperoxide dismutase (SOD) glutathione peroxidase (GPx)and malondialdehyde level (MDA) as well as DNA damagein DM patients [7 8] Antioxidant enzymes and dietarycompounds such as vitamins C E and flavonoids may havean effect in reducing the increased serum glucose levelin DM and secondary complications of neuropathy andnephropathy aswell asmacrovascular complications of strokeand cardiovascular problems in people with longstandinguncontrolled DM [9]

In the past decade rigorous research had been performedto find out the best way to overcome the complications asobserved in DM either by modern or alternative medicineIn Malaysia research on herbal medicine in alleviating DMusing animal and human studies has gained widemomentumin the past few years [10] A formulated polyherbal drugconsisting of extract of root of M paradisiaca seed of Tindica seed of E jambolana and leaf of C indica was foundto have a potent antidiabetogenic efficacy in STZ-induceddiabetic rats by reducing oxidative stress and carbohydratemetabolic profile and enzymes activities such glucose-6-phosphatase lactate dehydrogenase and hexokinase as wellas glucose-6-phosphate dehydrogenase in liver skeletal mus-cle and cardiac muscle towards the normal range [11] Aizzatet al [12] reported a reduction in lipid peroxidation asmeasured by MDA levels and reduction in DNA damageevaluated by Comet assay in STZ-induced diabetic rats whensupplemented with 150mgkg body weightChlorella vulgaris

For centuries honey has been used as a substitute forsugar as well as providing medicinal benefit In the pastdecade numerous biological and nutritional effects of honeyhave been reported such as antimicrobial antioxidant antivi-ral antiparasitic anti-inflammatory antimutagenic anti-cancer and immunosuppressive activities [13 14] In addi-tion Yao et al [15] showed that higher concentration of gelamhoney (5 gkg body weight) reduced oxidative damage betterthan lower concentration (25 gkg body weight) in youngand middle aged rats by modulating antioxidant enzymeactivities Ginger which is an underground rhizome of plantZingiber officinale belonging to the family Zingiberaceaewas reported to have hypoglycaemic anti-inflammatoryantitumorigenic and antioxidant effects in in vitro and in vivostudies [16ndash20] However to the best of our knowledge noprevious studies have explored the antidiabetic and antioxi-dant properties of the combination of gelamhoney and gingeron the oxidative stress and metabolic profile of STZ-induced

diabetic rats although a study by Patel et al [21] reportedthe synergistic effect of the combination of honey and gingeron the antibacterial property on isolates of extracted cariousteeth during orthodontic treatment Thus we embarked onthis project to evaluate whether the combination effect ofgelam honey and ginger would synergize in their actions inreducing oxidative stress and alleviating the complications ofdiabetes

2 Materials and Methods

21 Animals and Diet A total of 60 male Sprague-Dawleyrats were obtained from by Animal Care Unit UniversitiKebangsaan Malaysia (Bangi Malaysia) The studywas approved by the Animal Ethics Committee of theFaculty of Medicine Universiti Kebangsaan Malaysia (FPBIOKIMIA2012YASMIN26-SEPTEMBER463-SEPTEM-BER-2012-AUGUST-2013) Prior to the experiment ratsweighing 200ndash250 gm were randomly chosen They werehoused individually in polycarbonate cages at a temperatureof 22∘C with a 12-hour light-12-hour dark-cycle and wasquarantined for one week and fed with a standard labchow and water ad libitum The rats were divided into twogroups which were control (28 rats) and diabetic groups(32 rats) The control group was further divided into fourgroups (i) control group fed with normal lab chow (ii)control group fed with gelam honey (20 gkg body weight)(iii) control group fed with ginger (60mgkg body weight)and (iv) control group fed with gelam honey and ginger(20 gkg bodyweight honey + 60mgkg bodyweight ginger)Diabetes was induced by a single intramuscular injectionof streptozotocin (STZ) (55mgkg body weight) STZ wasprepared by dissolving it in cold 09 NaCl It was injectedinto overnight fasting rats through the thigh muscle bulkThe rats were observed closely for symptoms of DM such aspolydipsia polyuria and weight loss Blood samples wereobtained from ratsrsquo tail vein 48 hours after STZ injection Ifblood glucose level was gt14mmolL rats were considereddiabeticThe diabetic rats were then divided into four groupsthat is (i) STZ-induced diabetic rats fed with normal chowfood (ii) STZ-induced diabetic rats fed with gelam honey(20 gkg body weight) (iii) STZ-induced diabetic rats fedwith ginger (60mgkg body weight) and (iv) STZ-induceddiabetic rats fed with gelam honey and ginger The bloodglucose level was taken once weekly starting from week 0 toweek 3 After 3 weeks blood samples were obtained fromperiorbital sinus vein for the evaluation of metabolic andoxidative markers such as blood glucose triglyceride (TG)superoxide dismutase (SOD) glutathione peroxidase (GPx)catalase (CAT) activities glutathione peroxidase (GPx)reduced glutathione (GSH) oxidized glutathione (GSSG)and malondialdehyde (MDA)

22 Ginger Extract by Hot Compressed Water Extraction(HCW) An amount of 75 gm of ground ginger was weighedbefore being loaded into a covered stainless steel meshcylinder and placed into the extraction cell of fabricated hot



2gas










2O2













4



3 Results




0050

100150200250300350

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion

Glu

cose

leve

l (m

mol

L)


abcd

abcd

abcd


012345678

Trig

lyce

ride l

evel

(mm

olL

)

Triglyceride

abcd

abcde a

bcdf

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion




















02468

1012141618

SOD

activ

ity (U

mg

Hb)


a a ae

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion


02468

101214161820

Cata

lase

activ

ity (U

mg

Hb)


bcd

ae ae ae

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion



002040608

11214

GPx

activ

ity (m

Um

g H

b)


d

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion


4 Discussion



2







0200400600800

100012001400160018002000

GSH

(nm

olm


a a a a

abcdef

abcdef

gN

orm

alco

ntro

l

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(a)

05

101520253035

GSHGSSG ratio

abc

abcde

aef

efg

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(b)


0

10

20

30

40

50

60

Mal

onde

alde

hyde

elev

el (n

mol

L)


bcabcde

abcd

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion



5 Conclusion








Acknowledgments


References
























































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology



2gas










2O2













4



3 Results




0050

100150200250300350

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion

Glu

cose

leve

l (m

mol

L)


abcd

abcd

abcd


012345678

Trig

lyce

ride l

evel

(mm

olL

)

Triglyceride

abcd

abcde a

bcdf

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion




















02468

1012141618

SOD

activ

ity (U

mg

Hb)


a a ae

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion


02468

101214161820

Cata

lase

activ

ity (U

mg

Hb)


bcd

ae ae ae

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion



002040608

11214

GPx

activ

ity (m

Um

g H

b)


d

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion


4 Discussion



2







0200400600800

100012001400160018002000

GSH

(nm

olm


a a a a

abcdef

abcdef

gN

orm

alco

ntro

l

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(a)

05

101520253035

GSHGSSG ratio

abc

abcde

aef

efg

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(b)


0

10

20

30

40

50

60

Mal

onde

alde

hyde

elev

el (n

mol

L)


bcabcde

abcd

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion



5 Conclusion








Acknowledgments


References
























































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology





4



3 Results




0050

100150200250300350

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion

Glu

cose

leve

l (m

mol

L)


abcd

abcd

abcd


012345678

Trig

lyce

ride l

evel

(mm

olL

)

Triglyceride

abcd

abcde a

bcdf

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion




















02468

1012141618

SOD

activ

ity (U

mg

Hb)


a a ae

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion


02468

101214161820

Cata

lase

activ

ity (U

mg

Hb)


bcd

ae ae ae

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion



002040608

11214

GPx

activ

ity (m

Um

g H

b)


d

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion


4 Discussion



2







0200400600800

100012001400160018002000

GSH

(nm

olm


a a a a

abcdef

abcdef

gN

orm

alco

ntro

l

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(a)

05

101520253035

GSHGSSG ratio

abc

abcde

aef

efg

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(b)


0

10

20

30

40

50

60

Mal

onde

alde

hyde

elev

el (n

mol

L)


bcabcde

abcd

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion



5 Conclusion








Acknowledgments


References
























































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology

















02468

1012141618

SOD

activ

ity (U

mg

Hb)


a a ae

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion


02468

101214161820

Cata

lase

activ

ity (U

mg

Hb)


bcd

ae ae ae

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion



002040608

11214

GPx

activ

ity (m

Um

g H

b)


d

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion


4 Discussion



2







0200400600800

100012001400160018002000

GSH

(nm

olm


a a a a

abcdef

abcdef

gN

orm

alco

ntro

l

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(a)

05

101520253035

GSHGSSG ratio

abc

abcde

aef

efg

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(b)


0

10

20

30

40

50

60

Mal

onde

alde

hyde

elev

el (n

mol

L)


bcabcde

abcd

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion



5 Conclusion








Acknowledgments


References
























































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology


002040608

11214

GPx

activ

ity (m

Um

g H

b)


d

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

+gi

nger

(50 m

gkg

)

Dia

betic

com

bina

tion


4 Discussion



2







0200400600800

100012001400160018002000

GSH

(nm

olm


a a a a

abcdef

abcdef

gN

orm

alco

ntro

l

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(a)

05

101520253035

GSHGSSG ratio

abc

abcde

aef

efg

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(b)


0

10

20

30

40

50

60

Mal

onde

alde

hyde

elev

el (n

mol

L)


bcabcde

abcd

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion



5 Conclusion








Acknowledgments


References
























































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology


0200400600800

100012001400160018002000

GSH

(nm

olm


a a a a

abcdef

abcdef

gN

orm

alco

ntro

l

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(a)

05

101520253035

GSHGSSG ratio

abc

abcde

aef

efg

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion

(b)


0

10

20

30

40

50

60

Mal

onde

alde

hyde

elev

el (n

mol

L)


bcabcde

abcd

Nor

mal

cont

rol

Nor

mal

+ho

ney

(2 g

kg)

Nor

mal

com

bina

tion

Dia

betic

cont

rol

Dia

betic

+ho

ney

(2 g

kg)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

+gi

nger

(40 m

gkg

)

Dia

betic

com

bina

tion



5 Conclusion








Acknowledgments


References
























































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology


Acknowledgments


References
























































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology



























Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology








Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology

Documents

Research Article Effect of the Combination of Gelam Honey ...downloads.hindawi.com/journals/bmri/2014/160695.pdf · Academic Editor: Paul W. Huber ... BIOKIMIA//YASMIN/-SEPTEMBER