Research ArticleToxicological Evaluation of Essential Oils from SomePlants of Rutaceae Family

Iram Liaqat 1 Naila Riaz2 Qurat-ul-Ain Saleem2 Hafiz Muhammad Tahir1

Muhammad Arshad3 and Najma Arshad 2

1Department of Zoology Government College University Lahore Pakistan2Department of Zoology University of the Punjab Quaid-e-Azam Campus Lahore Pakistan3Department of Zoology University of Education Lahore Pakistan

Correspondence should be addressed to Najma Arshad najmaarshadgmailcom

Received 25 January 2018 Accepted 12 April 2018 Published 6 May 2018

Academic Editor Nativ Dudai

Copyright copy 2018 Iram Liaqat et al This is an open access article distributed under the Creative Commons Attribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

Essential oils are produced as secondary metabolites by aromatic plants predominantly belonging to families Apiaceae LamiaceaeMyrtaceae and Rutaceae The family Rutaceae has great economic importance for its numerous edible fruits and essential oilsIn the present study essential oils of seven plants of family Rutaceae Aegle marmelos Murraya koenigii Citrus reticulata BlancoZanthoxylum armatum Skimmia laureola Murraya paniculata and Boenninghausenia albiflora were used for their toxicologicalassessment Seven groups of selected essential oils-treated Wistar rats were established against control group (119899 = 5) that receivedwater for 14 days animals were offered feed and water ad libitum and treated with essential oils at 400mgkg body weightHematological studies revealed significant elevation in TEC in animals treated with essential oils ofM koenigii S laureola and Balbiflora while an elevation in PCV and depletion in MCV were observed in animals treated with M paniculata and B albiflorarespectively Serological investigations demonstrated significant depletion in triglycerides and elevation in blood sodium level inanimals treated with essential oils of A marmelos and C reticulata Blanco Boenninghausenia albiflora affected many markersincluding RBC MCV triglycerides HDL LDL urea and sodium In conclusion all oils except B albiflora can be considered safefor internal use

1 Introduction

Medicinal plants have been used in traditional treatmentsfor numerous human diseases for thousands of years par-ticularly in rural areas of developing countries [1] About80 of people in these countries use traditional medicinesfor healthcare purposes [2] The natural products obtainedfrom medicinal plants are abundant source of biologicallyactive compounds many of these products are being usedin the development of new products for pharmaceuticals andagriculture

Essential oils are formed by aromatic plants as secondarymetabolites and are volatile natural complex compoundscharacterized by a strong odor These oils were mostly usedfor their medicinal properties like antibactericidal antivi-ral antifungal antioxidants anti-inflammatory anticancerantihistamine and antidiabetic activities and were also used

in preservation of foods All these characteristics have notmuch changed up to the present day except that today theirmechanism of antimicrobial action is well understood [3]

Rutaceae is a family of flowering plants and is commonlyknown as the citrus family and is placed in the orderSapindales The flowers of the species are divided into fouror five parts and have strong scents They range in formand size from herbs to shrubs and small trees The familyis cosmopolitan and contains 154 genera Some members ofthe family are plants with highly fragrant flowers and areused in commercial oil production Some constituents ofessential oils such as citronella and bergamot are obtainedby distillation from plants of this family [4]

Inappropriate use of essential oils can leave adverse effectson human such as skin irritation headache and nauseaCaution is generally required if essential oils are to be takeninternally or used on food commodities because of the

HindawiEvidence-Based Complementary and Alternative MedicineVolume 2018 Article ID 4394687 7 pageshttpsdoiorg10115520184394687

2 Evidence-Based Complementary and Alternative Medicine

possible cancer-causing effects of some of them [5] Appliedat nonrecommendeddoses essential oils can cause functionaldamage to organs such as stomach and liver in animal andprobably in human [6]

In the following study we focused on in vitro antimi-crobial toxicity and in vivo toxicological aspects of essentialoils For this purpose an experiment was conducted to checkthe toxicity of essential oils on Wistar rats Seven differentessential oils of the family Rutaceae were selected Aeglemarmelos (bael)Murraya koenigii (curry plant) Citrus retic-ulata Blanco (honey plant) Zanthoxylum armatum (timber)Skimmia laureola (Skimmia) Murraya paniculata (Marwa)and Boenninghausenia albiflora (Pissu Mar)

2 Materials and Methods

The plants were collected from different hilly areas of Pak-istan Essential oils from leaves and stems of all plantswere extracted by hydrodistillation method In this methodthe plant material is almost entirely covered with water assuspension and is placed on a burner Water is made to boiland essential oil is carried over to the condenser along withthe steam Oil forms a layer on water from where water iswithdrawn and oil is collected The distillation period cantake from 15 to 30 minutes or longer

21 In Vitro Toxicity of Essential Oils

211 Antimicrobial Assay The antimicrobial activity of theselected essential oils was checked against two Gram-positive(Staphylococcus aureus and Staphylococcus epidermidis) andtwo Gram-negative (Escherichia coli and Klebsiella pneu-moniae) bacterial isolates of clinical importance using discdiffusion and microdilution assays and minimum inhibitoryconcentration (MIC) and minimum bactericidal concentra-tion (MBC) were determined

212 Disc Diffusion Method Sterilized cotton swabs dippedin respective cultureswere swabbed on solidified agar surfacePresterilized filter paper discs with a diameter of 6mmwere placed on the swabbed agar plate With the help of amicropipette 5 120583l of the respective oil was placed on the discand plates were incubated for 24 hours at 37∘C At the endof the incubation period the diameter of the inhibition zoneformed around the disc was measured in mm

213 Determination of Minimum Inhibitory Concentration(MIC) and Minimum Bactericidal Concentration (MBC) byMicrodilution Method Serial dilutions of essential oils innutrient brothwere prepared and 100120583l of bacterial inoculumwas added Plates were incubated for 24 hours at 37∘CThe concentration of oil which leads to absence of visiblegrowth of target strain was declared as MIC while for MBCmaterial from the wells showing no growth was subculturedand concentration that caused complete elimination of livebacteria was declared as MBC

22 In Vivo Toxicological Assessment of Essential Oils

221 Experimental Design The experiment was conductedon 24 healthy Wistar rats (Rattus norvegicus) purchased

from University of Veterinary and Animal Sciences (UVAS)Lahore weighing 150 grams and they were kept separatelyin 1210158401015840 times 1810158401015840 iron cages in the animal house of ZoologyDepartment of University of the Punjab Lahore at roomtemperature and 12-hour day and light period Chick feed3 (Hi-Tech Pakistan) was purchased from local marketand animals were provided with feed and water ad libitumAnimals were acclimatized for a period of 7 days beforestarting the experiment

222 Animal Grouping After acclimatization animals weredivided randomly into eight groups (ie I to VIII) Eachgroup was composed of 5 animals Group I was maintainedas control and was given water Essential oils from Aeglemarmelos Murraya koenigii Citrus reticulata Blanco Zan-thoxylum armatum Skimmia laureola Murraya paniculataand Boenninghausenia albiflora plants were given to animalsfrom group II to group VIII respectively

223 Dose Preparation Tween 20 was used for dissolvingoils Each oil was separately mixed in tween 20 in a ratio of1 1 vv in a vial and labeled Animals in groups II to VIIIwere given oral dose of the respective oil by using feedingneedle and the treatment was continued for 14 days A pilotexperiment was run to investigate maximal tolerated dose(MTD) of essential oils by using 200 400 and 800mgkgto a group of 3 rats each (data not shown)On the basis ofbehavioral study including no weight loss gt 20 as comparedto the first weighing procedure (to be performed the daybefore the administration) and no death the volume of thedose given was 400mgkg body weight in this study At theend of the experiment following 24 hours of last dose bloodsamples were collected from all animals one by one For thispurpose animals were anesthetized by chloroform and bloodwas drawn directly from the heart with the help of disposablesterilized plastic syringes (3 cc) A total of 05ml of thisblood was placed in EDTA coated vials (B2B Pakistan) forhematological study Rest of the bloodwas kept in refrigeratorfor 1 hour and centrifuged at 3000 rpm (revolution perminute) for 15 minutes and serum was separated in the cleanvial and stored at minus20∘C

224Hematological Study Thehemoglobin (Hb) contents ofblood were estimated by using Randox kit while total ery-throcyte count (TEC) and total leukocyte count (TLC) weremeasured using automated hematological analyzer followingusermanualThe packed cell volume (PCV)wasmeasured bymicrohematocrit method These hematological values wereused for calculation of hematological indices which includedMCV MCH and MCHC

225 Biochemical Parameters Biochemical parameters weredivided into four categories liver function test (LFT) renalfunction test (RFT) lipid profile and electrolytes LFTincluded bilirubin alanine aminotransferase (ALT) aspartateaminotransferase (AST) and alkaline phosphatase (ALP)and RFT included blood urea and creatinine LFT and RFTwere measured photometrically by using diagnostic kits forDiaSys diagnostic system Lipid profile included total choles-terol triglycerides high-density lipoproteins (HDL) and

Evidence-Based Complementary and Alternative Medicine 3

Table 1 Zone of inhibition of different essential oils against four bacterial isolates

Oils Gram-positive bacteria Gram-negative bacteriaS aureus (mm) S epidermidis (mm) E coli (mm) K pneumonia (mm)

I 24 18 18 20II 10 14 10 14III 16 12 11 10IV 14 15 11 10V 18 18 13 14VI 14 15 9 12VII 18 20 8 15Zone of inhibition of selected essential oils against S aureus S epidermidis E coli and K pneumoniae (I Aegle marmelos II Murraya koenigii III Citrusreticulata Blanco IV Zanthoxylum armatum V Skimmia laureola VIMurraya paniculata and VII Boenninghausenia albiflora)

Table 2 Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of different oils against four bacterialisolates ( vv)

Essential oilsGram-positive bacteria Gram-negative bacteria

S aureus S epidermidis E coli K pneumoniaeMIC MBC MIC MBC MIC MBC MIC MBC

Aegle marmelos 5 10 5 5 5 10 25 25Murraya koenigii 25 25 25 5 5 5 25 25Citrus reticulataBlanco 5 5 25 5 10 20 5 10

Zanthoxylumarmatum 0312 063 125 25 5 5 25 25

Skimmia laureola 0625 063 125 125 5 5 5 5Murrayapaniculata 125 25 125 25 25 25 5 5

Boenninghauseniaalbiflora 25 25 0156 016 25 5 5 5

low-density lipoproteins (LDL) All these parameters weremeasured using kits provided by Beckman diagnostics onautomatic analyzer (Beckman Coulter) Electrolytes includedblood sodium and potassium level which were measured byflame photometric method

23 Statistical Analysis The values of oil-treated groups werecompared with control by using SPSS software The data wasexpressed asmeanplusmn SEM(standard error ofmean) Statisticalcomparison was made for each oil with negative control byusing independent sample t-test The results were consideredas statistically significant when 119901 le 005

3 Results

31 In Vitro Antimicrobial Activity

311 Disc Diffusion Assay The antibacterial activity of sevenselected essential oils against four bacterial species is sum-marized in Table 1 The results of the disc diffusion methodrevealed that the selected essential oils exhibit varying mag-nitudes of antibacterial activity Almost all of the essential oilswere active against the selected isolates According to resultsA marmelos had the highest antimicrobial activity against S

aureus E coli and K pneumoniae while Boenninghauseniaalbiflora expressed highest activity against S epidermidis

312 Microdilution Assay Results ofMIC andMBC revealedthat all the selected essential oils possess antimicrobialactivity against two Gram-positive (S aureus and S epider-midis) and two Gram-negative (E coli and K pneumoniae)bacterial isolates All the oils were found to be more activeagainst Gram-positive isolates as compared toGram-negativeisolatesZanthoxylumarmatum andMurraya paniculatawerethe most active essential oils and had smallest MIC Citrusreticulata had least antimicrobial activity Among all thetested strains S epidermidis was the most sensitive strainfollowed by S aureus and K pneumoniae while E coli wasthe most resistant strain (Table 2)

32 Toxicological Evaluation

321 Hematological Analysis Hematological studies re-vealed a significant elevation in TEC in groups VI and VIIIThe elevation was 48 and 128 respectively PCV waselevated by 2255 in group VII MCV decreased by 4642in group VIII There was no alteration in TLC MCH andMCHC level in any treated group (Figure 1)

4 Evidence-Based Complementary and Alternative Medicine

0

10

20

30

40

50

60

A-1 A-2 A-3 A-4 A-5 A-6 A-7

UN

ITS

VALU

ES IN

RES

PECT

IVE

HEMATOLOGICAL PARAMETERS

ControlA marmelosM koenigiiC reticulata Blanco

Z armatumS laureolaM paniculataB albiflora

lowast

lowast

lowast

lowast

Figure 1 Comparison of hematological parameters in the essentialoil- (EO-) fed animals and non-EO-treated (control) animals (I)A-1 hemoglobin (mgdl) A-2 total erythrocyte count (times106120583l) A-3 total leucocyte count (times103120583l) A-4 packed cell volume () A-5 mean corpuscular volume (fLlowast10cell) A-6 mean corpuscularhemoglobin (pgcell) and A-7 mean corpuscular hemoglobin con-centration (gdl) Data is presented as mean plusmn SEM Asterisks showsignificant difference from control

UN

ITS

VALU

ES IN

RES

PECT

IVE

ControlA marmelosM koenigiiC reticulata Blanco

Z armatumS laureolaM paniculataB albiflora

0102030405060708090

B-1 B-2 B-3 B-4 B-5 B-6LIVER AND RENAL FUNCTION TESTS

lowast

lowast

lowast

lowast

lowast

Figure 2 Comparison of liver and renal function tests of essen-tial oil- (EO-) fed animals and control animals B-1 bilirubin(times100mgdl) B-2 alanine transaminase (IUL) and aspartatetransaminase (IUL) B-3 alkaline phosphatase (Udl) B-4 urea(mgdl) and B-5 creatinine (times100mgdl) Data are presented asmean plusmn SEM Asterisks on respective groups show significantdifference from control group at 119901 le 005

322 Liver Function Test and Renal Function Test There wasa nonsignificant decrease in bilirubin ALT AST and ALPlevels which are markers for liver injury An elevated level ofblood urea was observed in groups V VI VII and VIII with2580 3870 4731 and 7311 increase respectivelyCreatinine level remained unchanged in all treatment groups(Figure 2)

323 Lipid Profile Statistical analysis revealed a significantdepletion in triglycerides in groups II IV VI VII and

ControlA marmelosM koenigiiC reticulata

Z armatumS laureolaM paniculataB albiflora

0

50

100

150

200

Cholesterol Triglyceride HDL LDL

Valu

es (m

gdl

)

LIPID PROFILE

lowastlowast

lowast

lowast

lowast

lowast

lowast

Figure 3 Comparison of lipid profile (mgdL) in essential oil-(EO-) fed animals and non-EO-treated control animals (I) Data arepresented as mean plusmn SEM while asterisks on respective EO groupsshow significant difference from control at 119901 le 005

Valu

es in

resp

ectiv

e uni

ts

ControlA marmelosM koenigiiC reticulata

Z armatumS laureolaM paniculataB albiflora

0369

121518

Sodium (mmoledl) Potassium (mmoleL)Serum Electrolytes

lowastlowastlowast

lowastlowastlowast

Figure 4 Comparison of blood electrolytes in essential oil- (EO-)fed animals and control animals (I) Data are presented as mean plusmnSEM Asterisks on respective EO groups show significant differencefrom control at 119901 le 005

VIII The level increased by 4933 4849 3193 4147and 4615 respectively HDL and LDL levels significantlydecreased in group VIII by 3070 and 3045 respectively(Figure 3)

324 Electrolytes The results revealed that all the selectedoils have potential to elevate blood sodium level Statisticalanalysis indicated a significant difference in groups II IV VVI VII and VIII The increase in level was 1189 116516 1262 1747 and 1383 respectively There was noalteration observed in blood potassium level in any essentialoil fed group (Figure 4)

4 Discussion

The increased use of antimicrobial agents has resulted in thedevelopment of resistant strains of bacteria So there is a need

Evidence-Based Complementary and Alternative Medicine 5

to develop effective antimicrobial agents with new modes ofaction against these pathogenic microbes Essential oils havebeen recognized as a rich source of new bioactive secondarymetabolites which possess the potential of treatment ofmanyinfectious diseases In the current study the potential ofselected essential oils as antimicrobial agents was screenedagainst four resistant clinical isolates including two Gram-positive (S aureus and S epidermidis) and twoGram-negative(E coli andK pneumoniae) bacteriaThe antibacterial activitywas initially screened by using disc diffusion method andthen theminimum inhibitory concentration (MIC) andmin-imum bactericidal concentration (MBC) were determined byusing microdilution method

The results of the disc diffusion method revealed that theselected essential oils exhibit varying magnitudes of antibac-terial activity Almost all of the essential oils were activeagainst the four clinical isolates According to results Aeglemarmelos was found to possess the highest antimicrobialactivity against S aureus E coli and K pneumoniae whileBoenninghausenia albiflora shows highest activity against Sepidermidis All the oils were found to be more active againstGram-positive bacterial isolates (S aureus and S epidermidis)and were less active against Gram-negative bacteria (E coliand K pneumoniae) [7]

Results of MIC and MBC also revealed that all the oilswere more active against Gram-positive as compared toGram-negative bacterial isolates Zanthoxylum armatum andMurraya paniculata were the most active essential oils andwere active against all the selected isolates Citrus reticulataBlanco was the least active oil with low antibacterial activityGram-negative bacteria are more resistant to essential oilsthen Gram-positive bacteria due to permeability barrier pro-vided by extra lipopolysaccharidemembraneThe antimicro-bial activity of essential oils could be due to their hydrophobiccharacteristic due to which these oils are capable of breakingthe lipids of bacterial cell membrane and making them morepermeable [8]

The current study involved the investigation of hemato-logical and serological changes in rats fed with essential oilsThe hematological parameters can be influenced by toxicityof essential oils as these oils have ability to initiate the acutephase response [9] Significant difference was observed inRBC PCV and MCV levels between the treated and controlgroups However in most of the oil feeding groups the HbTEC TLC PCV MCV MCH and MCHC were in normalrange

Hemoglobin is a metalloprotein that contains iron andis present in red blood cells of all vertebrates It transportsoxygen from respiratory organs to the rest of the body Inthe current study the results indicate that the hemoglobinlevel nonsignificantly increased in oil-fed groups The reasonfor this slight increase in hemoglobin was an increase inthe number of red blood cells Erythrocytes are the mostcommon type of blood cells and are the main source oftransferring oxygen to the body tissues RBCs are formed inbone marrow and stimulated by decrease in oxygen which isdetected by kidneys [10] In the current study a significantelevation in TEC was noticed in the groups of animalswhich were treated with M koenigii S laureola and B

albiflora essential oils Studies had shown that morphologicalchanges occur in RBCs in response to various treatmentsby toxic agents [11] According to Suwalsky et al [12]Balbisia peduncularis extract caused changes in the normalerythrocyte morphology An elevation in total erythrocyteswas noticed in some EO-treated groups Packed cell volume(PCV) or hematocrit is the volume percentage of red bloodcells in blood PCV was slightly increased in all essential oil-fed groups but a significant elevation was observed only ingroup treatedwith essential oil ofM paniculataThis increasein PCV may be due to an increase in RBCs number in theoil-fed groups [13] This finding is supported by an elevatederythrocytes number in the same group Mean corpuscularvolume (MCV) is ameasure of the average red blood cell sizeThe results indicated significant decrease in the level of MCVin the group treated with essential oil of B albiflora Thisreduction in MCV was maybe due to decrease in size of redblood cell [13] There was no significant difference observedinMCHandMCHC in any oil-fed groupThat was because ofthe hemoglobin level whichwas in normal range in all groups

Leukocytes are involved in the defense mechanism ofthe body against foreign materials In the present studyno significant change was observed in any oil-fed groupwhen compared to control group All the groups indicatethe number of white blood cells in normal range This is theindication that the selected essential oils perhaps do not affectleucopoiesis or half-life of leukocytes

The serological parameters selected for the study includedthe liver function test renal function test lipid profileand serum electrolytes The selected essential oils cause anonsignificant depletion in the level of all themarkers for liverinjury which included bilirubin alanine aminotransferase(ALT) aspartate aminotransferase (AST) and alkaline Phos-phatase (ALP) This is an indication that all these oils do nothave toxic effects particularly in reference to liver functiontest

Renal function test is an indication of the state of kidneyUrea and creatinine are usually considered as markers withrespect to kidney function Urea is the nitrogenous wasteproduct of the body Raised level of blood urea is anindication of renal dysfunction An elevated level of bloodurea was observed in some of the essential oil-fed groupsSignificant elevation was observed in groups treated with Zarmatum S laureolaM paniculata and B albiflora essentialoils This increase in urea level may be due to epithelialnecrosis to the renal tubules [14] On the other hand bloodcreatinine level remained unaltered in all essential oi-fedgroups Indicating EO at this dose may cause partial toxicityto kidneys According to Abarikwu et al [15] whenever theurea level increased and creatinine level is reduced there is nokidney damage Nisha et al [16] reported that renal toxicityshould be considered only when creatinine and urea levelincreased parallel to each other

Lipid profile is the term given to the evaluation of totalcholesterol triglycerides high-density lipoproteins (HDL)and low-density lipoproteins (LDL) This test is mostly usedto identify hyperlipidemia which is risk factor for heartdiseases Cholesterol is an ester and is an important part ofmammalian cell membrane [17] It is formed in the liver

6 Evidence-Based Complementary and Alternative Medicine

In this study no variation was observed in cholesterol levelin any essential oil-fed group According to Rajadurai andPrince [18] the cholesterol level slightly decreases in rats intreatment withAeglemarmelos leaf extractsMurraya koenigiileaf extracts also decrease cholesterol level in rats [19]

Triglyceride is also an ester and is derived from glyceroland three fatty acids High triglyceride is directly related tocoronary heart disease Our results indicate a decrease intriglycerides level in all essential oil-fed groups as comparedto control group Statistical analysis indicated a significantdepletion in groups treated with A marmelos C reticulataBlanco S laureola M paniculata and B albiflora Ourfindings are in accordance with Rajadurai et al [18] andRajadurai et al [19] they reported that the leaf extracts of Amarmelos cause a decrease in triglycerides level in rat models

Recent studies suggest that triglyceride itself is inde-pendently related to coronary heart disease [20 21] andmost of the antihypercholesterolemic drugs do not decreasetriglycerides levels but plant extracts do These findingsindicate beneficial effects of essential oils instead of theirtoxicological potential

High-density lipoprotein (HDL) is also known as goodcholesterol It transports other lipids like cholesterol andtriglycerides in the blood stream Various studies have shownthat an increase in HDL-cholesterol is associated with adecrease in coronary risk [22] Statistical analysis indicateda significant decrease in HDL in animals treated with Balbiflora essential oil So this essential oil may be consideredas toxic because it is decreasing good cholesterol

Low-density lipoproteins (LDL) are also known as badcholesterol LDL collects in the walls of blood vessels andcauses the blockage of the arteries Higher LDL levels maycause sudden blood clot in an artery and increase risk of heartattack In present study significant difference was observedin animals treated with B albiflora Our findings are inaccordance with Kesari et al [19] who reported that theadministration of leaf extracts ofM koenigii causes a decreasein the level of LDL of normal and diabetic rats So theseessential oils play a beneficiary role

Sodium and potassium are electrolytes present in bloodand other body fluids These help keep the water andelectrolyte balance of the body and are also important inthe proper functioning of nerves and muscles The hormonealdosterone controls the level of sodium and potassium in thebody The results revealed that all the selected essential oilshave potential to elevate blood sodium level Statistical analy-sis indicated a significant elevation in animals treated withAmarmelos C reticulataBlanco S laureolaM paniculata andB albiflora Similar findings have been reported by Odeyemiet al [23] who concluded that all essential oils may not besafe as these can lead to leakage of electrolytes from the cellsHowever in current study blood potassium level remainedunaltered

In conclusion although EOs were found to have somefavorable insinuations (decrease in triglycerides and LDLalong with no adverse effects on LFT and RFT) they mayleave specific undesired effects (eg increase in urea (Zarmatum) depletion in HDL (B albiflora) induction inerythropoiesis above normal level (M koenigii S laureola

and B albiflora) and increase in sodium (A marmelos CreticulataBlanco S laureolaM paniculata andB albiflora))which point to their toxic potential

5 Conclusion

As reduction in triglycerides is considered as beneficial andchange in urea without alteration in creatinine is not consid-ered as toxic output of any tested material it could be con-cluded that all oils except B albiflora can be considered safefor internal use with caution and sodium level may be contin-uously monitored B albiflora affected many markers includ-ing RBC MCV triglycerides HDL LDL urea and sodium

Data Availability

The data used to support the findings of this study areincluded within the article

Conflicts of Interest

The authors declare that they have no conflicts of interest

References

[1] H R Chitme R Chandra and S Kaushik ldquoStudies on anti-diarrheal activity of Calotropis gigantea R Br in experimentalanimalsrdquo Journal of Pharmacy and Pharmaceutical Sciences vol7 no 1 pp 70ndash75 2004

[2] H-S Kim ldquoDo not put too much value on conventionalmedicinesrdquo Journal of Ethnopharmacology vol 100 no 1-2 pp37ndash39 2005

[3] F Bakkali S Averbeck D Averbeck and M Idaomar ldquoBio-logical effects of essential oilsmdasha reviewrdquo Food and ChemicalToxicology vol 46 no 2 pp 446ndash475 2008

[4] S A Aziz M A Sukari M Rahmani M Kitajima NAimi and N J Ahpandi ldquoCoumarins from murraya pan-iculata (rutaceae) (koumarin daripada murraya paniculata(rutaceae))rdquoMalaysian Journal of Analytical Sciences vol 14 no1 pp 1ndash5 2010

[5] M Mcguffin C Hobbs R Upton and A Goldberg AmericanHerbal Products 1997 American Herbal Products

[6] S L Ngahang Kamte F Ranjbarian K Cianfaglione et alldquoIdentification of highly effective antitrypanosomal compoundsin essential oils from the Apiaceae familyrdquo Ecotoxicology andEnvironmental Safety vol 156 pp 154ndash165 2018

[7] Y Wang Y Zhang Y Shi X Pan Y Lu and P CaoldquoAntibacterial effects of cinnamon (Cinnamomum zeylanicum)bark essential oil on Porphyromonas gingivalisrdquo MicrobialPathogenesis vol 116 pp 26ndash32 2018

[8] R Tardugno F Pellati R Iseppi M Bondi G Bruzzesi and SBenvenuti ldquoPhytochemical composition and in vitro screeningof the antimicrobial activity of essential oils on oral pathogenicbacteriardquo Natural Product Research vol 32 no 5 pp 544ndash5512018

[9] A Veerappan S Miyazaki M Kadarkaraisamy and D Ran-ganathan ldquoAcute and subacute toxicity studies of Aegle marme-los Corr an Indianmedicinal plantrdquo Phytomedicine vol 14 no2-3 pp 209ndash215 2007

[10] J Palis and G B Segel ldquoDevelopmental biology of erythro-poiesisrdquo Blood Reviews vol 12 no 2 pp 106ndash114 1998

Evidence-Based Complementary and Alternative Medicine 7

[11] A Brum S A Pereira L Cardoso et al ldquoBlood biochemicalparameters and melanomacrophage centers in Nile tilapiafed essential oils of clove basil and gingerrdquo Fish amp ShellfishImmunology vol 74 pp 444ndash449 2018

[12] M Suwalsky P Vargas M Avello F Villena and C PSotomayor ldquoHuman erythrocytes are affected in vitro byflavonoids of Aristotelia chilensis (Maqui) leavesrdquo InternationalJournal of Pharmaceutics vol 363 no 1-2 pp 85ndash90 2008

[13] O T Oyesina A M Salihu O L Aribidesi and A D AdedayoldquoEffect of essential oil of the leaves of Eucalyptus globulesonheamatological parameters of wistar ratsrdquo African Journal ofBiochemistry Research vol 6 no 4 pp 46ndash49 2012

[14] W Janssen Forensic Histopathology Springer Berlin Germany1984

[15] S O Abarikwu R C Njoku and C L Onuah ldquoAged coconutoil with a high peroxide value induces oxidative stress and tissuedamage in mercury-treated ratsrdquo Journal of Basic and ClinicalPhysiology and Pharmacology 2018

[16] R Nisha S R Srinivasa Kannan K Thanga Mariappan andP Jagatha ldquoBiochemical evaluation of creatinine and urea inpatients with renal failure undergoing hemodialysisrdquo Journal ofClinical Pathology and LaboratoryMedicine vol 1 no 2 pp 1ndash52017

[17] B R Zirkin and V Papadopoulos ldquoLeydig Cells FormationFunction and Regulationrdquo Biology of Reproduction 2018

[18] M Rajadurai and P S M Prince ldquoComparative effects of Aeglemarmelos extract and alpha-tocopherol on serum lipids lipidperoxides and cardiac enzyme levels in rats with isoproterenol-induced myocardial infarctionrdquo Singapore Medical Journal vol46 no 2 pp 78ndash81 2005

[19] A N Kesari S Kesari S K Singh R K Gupta and GWatal ldquoStudies on the glycemic and lipidemic effect of Murrayakoenigii in experimental animalsrdquo Journal of Ethnopharmacol-ogy vol 112 no 2 pp 305ndash311 2007

[20] S Saljoughian S Roohinejad A E-D A Bekhit et al ldquoTheeffects of food essential oils on cardiovascular diseases Areviewrdquo Critical Reviews in Food Science and Nutrition vol 10pp 1ndash18 2017

[21] VD Gooty A R Sinaiko J R Ryder D R Dengel D R Jacobsand J Steinberger ldquoAssociation Between Carotid Intima MediaThickness Age and Cardiovascular Risk Factors in Childrenand Adolescentsrdquo Metabolic Syndrome and Related Disordersvol 16 no 3 pp 122ndash126 2018

[22] A Mancini E Imperlini E Nigro et al ldquoBiological andnutritional properties of palm oil and palmitic acid Effects onhealthrdquoMolecules vol 20 no 9 pp 17339ndash17361 2015

[23] O O Odeyemi M T Yakubu P J Masika and A J AfolayanldquoToxicological evaluation of the essential oil from menthalongifolia l subsp capensis leaves in ratsrdquo Journal of MedicinalFood vol 12 no 3 pp 669ndash674 2009

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Evidence-Based Complementary and Alternative Medicine 3

Table 1 Zone of inhibition of different essential oils against four bacterial isolates

Oils Gram-positive bacteria Gram-negative bacteriaS aureus (mm) S epidermidis (mm) E coli (mm) K pneumonia (mm)

I 24 18 18 20II 10 14 10 14III 16 12 11 10IV 14 15 11 10V 18 18 13 14VI 14 15 9 12VII 18 20 8 15Zone of inhibition of selected essential oils against S aureus S epidermidis E coli and K pneumoniae (I Aegle marmelos II Murraya koenigii III Citrusreticulata Blanco IV Zanthoxylum armatum V Skimmia laureola VIMurraya paniculata and VII Boenninghausenia albiflora)

Table 2 Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of different oils against four bacterialisolates ( vv)

Essential oilsGram-positive bacteria Gram-negative bacteria

S aureus S epidermidis E coli K pneumoniaeMIC MBC MIC MBC MIC MBC MIC MBC

Aegle marmelos 5 10 5 5 5 10 25 25Murraya koenigii 25 25 25 5 5 5 25 25Citrus reticulataBlanco 5 5 25 5 10 20 5 10

Zanthoxylumarmatum 0312 063 125 25 5 5 25 25

Skimmia laureola 0625 063 125 125 5 5 5 5Murrayapaniculata 125 25 125 25 25 25 5 5

Boenninghauseniaalbiflora 25 25 0156 016 25 5 5 5

low-density lipoproteins (LDL) All these parameters weremeasured using kits provided by Beckman diagnostics onautomatic analyzer (Beckman Coulter) Electrolytes includedblood sodium and potassium level which were measured byflame photometric method

23 Statistical Analysis The values of oil-treated groups werecompared with control by using SPSS software The data wasexpressed asmeanplusmn SEM(standard error ofmean) Statisticalcomparison was made for each oil with negative control byusing independent sample t-test The results were consideredas statistically significant when 119901 le 005

3 Results

31 In Vitro Antimicrobial Activity

311 Disc Diffusion Assay The antibacterial activity of sevenselected essential oils against four bacterial species is sum-marized in Table 1 The results of the disc diffusion methodrevealed that the selected essential oils exhibit varying mag-nitudes of antibacterial activity Almost all of the essential oilswere active against the selected isolates According to resultsA marmelos had the highest antimicrobial activity against S

aureus E coli and K pneumoniae while Boenninghauseniaalbiflora expressed highest activity against S epidermidis

312 Microdilution Assay Results ofMIC andMBC revealedthat all the selected essential oils possess antimicrobialactivity against two Gram-positive (S aureus and S epider-midis) and two Gram-negative (E coli and K pneumoniae)bacterial isolates All the oils were found to be more activeagainst Gram-positive isolates as compared toGram-negativeisolatesZanthoxylumarmatum andMurraya paniculatawerethe most active essential oils and had smallest MIC Citrusreticulata had least antimicrobial activity Among all thetested strains S epidermidis was the most sensitive strainfollowed by S aureus and K pneumoniae while E coli wasthe most resistant strain (Table 2)

32 Toxicological Evaluation

321 Hematological Analysis Hematological studies re-vealed a significant elevation in TEC in groups VI and VIIIThe elevation was 48 and 128 respectively PCV waselevated by 2255 in group VII MCV decreased by 4642in group VIII There was no alteration in TLC MCH andMCHC level in any treated group (Figure 1)

4 Evidence-Based Complementary and Alternative Medicine

0

10

20

30

40

50

60

A-1 A-2 A-3 A-4 A-5 A-6 A-7

UN

ITS

VALU

ES IN

RES

PECT

IVE

HEMATOLOGICAL PARAMETERS

ControlA marmelosM koenigiiC reticulata Blanco

Z armatumS laureolaM paniculataB albiflora

lowast

lowast

lowast

lowast

Figure 1 Comparison of hematological parameters in the essentialoil- (EO-) fed animals and non-EO-treated (control) animals (I)A-1 hemoglobin (mgdl) A-2 total erythrocyte count (times106120583l) A-3 total leucocyte count (times103120583l) A-4 packed cell volume () A-5 mean corpuscular volume (fLlowast10cell) A-6 mean corpuscularhemoglobin (pgcell) and A-7 mean corpuscular hemoglobin con-centration (gdl) Data is presented as mean plusmn SEM Asterisks showsignificant difference from control

UN

ITS

VALU

ES IN

RES

PECT

IVE

ControlA marmelosM koenigiiC reticulata Blanco

Z armatumS laureolaM paniculataB albiflora

0102030405060708090

B-1 B-2 B-3 B-4 B-5 B-6LIVER AND RENAL FUNCTION TESTS

lowast

lowast

lowast

lowast

lowast

Figure 2 Comparison of liver and renal function tests of essen-tial oil- (EO-) fed animals and control animals B-1 bilirubin(times100mgdl) B-2 alanine transaminase (IUL) and aspartatetransaminase (IUL) B-3 alkaline phosphatase (Udl) B-4 urea(mgdl) and B-5 creatinine (times100mgdl) Data are presented asmean plusmn SEM Asterisks on respective groups show significantdifference from control group at 119901 le 005

322 Liver Function Test and Renal Function Test There wasa nonsignificant decrease in bilirubin ALT AST and ALPlevels which are markers for liver injury An elevated level ofblood urea was observed in groups V VI VII and VIII with2580 3870 4731 and 7311 increase respectivelyCreatinine level remained unchanged in all treatment groups(Figure 2)

323 Lipid Profile Statistical analysis revealed a significantdepletion in triglycerides in groups II IV VI VII and

ControlA marmelosM koenigiiC reticulata

Z armatumS laureolaM paniculataB albiflora

0

50

100

150

200

Cholesterol Triglyceride HDL LDL

Valu

es (m

gdl

)

LIPID PROFILE

lowastlowast

lowast

lowast

lowast

lowast

lowast

Figure 3 Comparison of lipid profile (mgdL) in essential oil-(EO-) fed animals and non-EO-treated control animals (I) Data arepresented as mean plusmn SEM while asterisks on respective EO groupsshow significant difference from control at 119901 le 005

Valu

es in

resp

ectiv

e uni

ts

ControlA marmelosM koenigiiC reticulata

Z armatumS laureolaM paniculataB albiflora

0369

121518

Sodium (mmoledl) Potassium (mmoleL)Serum Electrolytes

lowastlowastlowast

lowastlowastlowast

Figure 4 Comparison of blood electrolytes in essential oil- (EO-)fed animals and control animals (I) Data are presented as mean plusmnSEM Asterisks on respective EO groups show significant differencefrom control at 119901 le 005

VIII The level increased by 4933 4849 3193 4147and 4615 respectively HDL and LDL levels significantlydecreased in group VIII by 3070 and 3045 respectively(Figure 3)

324 Electrolytes The results revealed that all the selectedoils have potential to elevate blood sodium level Statisticalanalysis indicated a significant difference in groups II IV VVI VII and VIII The increase in level was 1189 116516 1262 1747 and 1383 respectively There was noalteration observed in blood potassium level in any essentialoil fed group (Figure 4)

4 Discussion

The increased use of antimicrobial agents has resulted in thedevelopment of resistant strains of bacteria So there is a need

Evidence-Based Complementary and Alternative Medicine 5

to develop effective antimicrobial agents with new modes ofaction against these pathogenic microbes Essential oils havebeen recognized as a rich source of new bioactive secondarymetabolites which possess the potential of treatment ofmanyinfectious diseases In the current study the potential ofselected essential oils as antimicrobial agents was screenedagainst four resistant clinical isolates including two Gram-positive (S aureus and S epidermidis) and twoGram-negative(E coli andK pneumoniae) bacteriaThe antibacterial activitywas initially screened by using disc diffusion method andthen theminimum inhibitory concentration (MIC) andmin-imum bactericidal concentration (MBC) were determined byusing microdilution method

The results of the disc diffusion method revealed that theselected essential oils exhibit varying magnitudes of antibac-terial activity Almost all of the essential oils were activeagainst the four clinical isolates According to results Aeglemarmelos was found to possess the highest antimicrobialactivity against S aureus E coli and K pneumoniae whileBoenninghausenia albiflora shows highest activity against Sepidermidis All the oils were found to be more active againstGram-positive bacterial isolates (S aureus and S epidermidis)and were less active against Gram-negative bacteria (E coliand K pneumoniae) [7]

Results of MIC and MBC also revealed that all the oilswere more active against Gram-positive as compared toGram-negative bacterial isolates Zanthoxylum armatum andMurraya paniculata were the most active essential oils andwere active against all the selected isolates Citrus reticulataBlanco was the least active oil with low antibacterial activityGram-negative bacteria are more resistant to essential oilsthen Gram-positive bacteria due to permeability barrier pro-vided by extra lipopolysaccharidemembraneThe antimicro-bial activity of essential oils could be due to their hydrophobiccharacteristic due to which these oils are capable of breakingthe lipids of bacterial cell membrane and making them morepermeable [8]

The current study involved the investigation of hemato-logical and serological changes in rats fed with essential oilsThe hematological parameters can be influenced by toxicityof essential oils as these oils have ability to initiate the acutephase response [9] Significant difference was observed inRBC PCV and MCV levels between the treated and controlgroups However in most of the oil feeding groups the HbTEC TLC PCV MCV MCH and MCHC were in normalrange

Hemoglobin is a metalloprotein that contains iron andis present in red blood cells of all vertebrates It transportsoxygen from respiratory organs to the rest of the body Inthe current study the results indicate that the hemoglobinlevel nonsignificantly increased in oil-fed groups The reasonfor this slight increase in hemoglobin was an increase inthe number of red blood cells Erythrocytes are the mostcommon type of blood cells and are the main source oftransferring oxygen to the body tissues RBCs are formed inbone marrow and stimulated by decrease in oxygen which isdetected by kidneys [10] In the current study a significantelevation in TEC was noticed in the groups of animalswhich were treated with M koenigii S laureola and B

albiflora essential oils Studies had shown that morphologicalchanges occur in RBCs in response to various treatmentsby toxic agents [11] According to Suwalsky et al [12]Balbisia peduncularis extract caused changes in the normalerythrocyte morphology An elevation in total erythrocyteswas noticed in some EO-treated groups Packed cell volume(PCV) or hematocrit is the volume percentage of red bloodcells in blood PCV was slightly increased in all essential oil-fed groups but a significant elevation was observed only ingroup treatedwith essential oil ofM paniculataThis increasein PCV may be due to an increase in RBCs number in theoil-fed groups [13] This finding is supported by an elevatederythrocytes number in the same group Mean corpuscularvolume (MCV) is ameasure of the average red blood cell sizeThe results indicated significant decrease in the level of MCVin the group treated with essential oil of B albiflora Thisreduction in MCV was maybe due to decrease in size of redblood cell [13] There was no significant difference observedinMCHandMCHC in any oil-fed groupThat was because ofthe hemoglobin level whichwas in normal range in all groups

Leukocytes are involved in the defense mechanism ofthe body against foreign materials In the present studyno significant change was observed in any oil-fed groupwhen compared to control group All the groups indicatethe number of white blood cells in normal range This is theindication that the selected essential oils perhaps do not affectleucopoiesis or half-life of leukocytes

The serological parameters selected for the study includedthe liver function test renal function test lipid profileand serum electrolytes The selected essential oils cause anonsignificant depletion in the level of all themarkers for liverinjury which included bilirubin alanine aminotransferase(ALT) aspartate aminotransferase (AST) and alkaline Phos-phatase (ALP) This is an indication that all these oils do nothave toxic effects particularly in reference to liver functiontest

Renal function test is an indication of the state of kidneyUrea and creatinine are usually considered as markers withrespect to kidney function Urea is the nitrogenous wasteproduct of the body Raised level of blood urea is anindication of renal dysfunction An elevated level of bloodurea was observed in some of the essential oil-fed groupsSignificant elevation was observed in groups treated with Zarmatum S laureolaM paniculata and B albiflora essentialoils This increase in urea level may be due to epithelialnecrosis to the renal tubules [14] On the other hand bloodcreatinine level remained unaltered in all essential oi-fedgroups Indicating EO at this dose may cause partial toxicityto kidneys According to Abarikwu et al [15] whenever theurea level increased and creatinine level is reduced there is nokidney damage Nisha et al [16] reported that renal toxicityshould be considered only when creatinine and urea levelincreased parallel to each other

Lipid profile is the term given to the evaluation of totalcholesterol triglycerides high-density lipoproteins (HDL)and low-density lipoproteins (LDL) This test is mostly usedto identify hyperlipidemia which is risk factor for heartdiseases Cholesterol is an ester and is an important part ofmammalian cell membrane [17] It is formed in the liver

6 Evidence-Based Complementary and Alternative Medicine

In this study no variation was observed in cholesterol levelin any essential oil-fed group According to Rajadurai andPrince [18] the cholesterol level slightly decreases in rats intreatment withAeglemarmelos leaf extractsMurraya koenigiileaf extracts also decrease cholesterol level in rats [19]

Triglyceride is also an ester and is derived from glyceroland three fatty acids High triglyceride is directly related tocoronary heart disease Our results indicate a decrease intriglycerides level in all essential oil-fed groups as comparedto control group Statistical analysis indicated a significantdepletion in groups treated with A marmelos C reticulataBlanco S laureola M paniculata and B albiflora Ourfindings are in accordance with Rajadurai et al [18] andRajadurai et al [19] they reported that the leaf extracts of Amarmelos cause a decrease in triglycerides level in rat models

Recent studies suggest that triglyceride itself is inde-pendently related to coronary heart disease [20 21] andmost of the antihypercholesterolemic drugs do not decreasetriglycerides levels but plant extracts do These findingsindicate beneficial effects of essential oils instead of theirtoxicological potential

High-density lipoprotein (HDL) is also known as goodcholesterol It transports other lipids like cholesterol andtriglycerides in the blood stream Various studies have shownthat an increase in HDL-cholesterol is associated with adecrease in coronary risk [22] Statistical analysis indicateda significant decrease in HDL in animals treated with Balbiflora essential oil So this essential oil may be consideredas toxic because it is decreasing good cholesterol

Low-density lipoproteins (LDL) are also known as badcholesterol LDL collects in the walls of blood vessels andcauses the blockage of the arteries Higher LDL levels maycause sudden blood clot in an artery and increase risk of heartattack In present study significant difference was observedin animals treated with B albiflora Our findings are inaccordance with Kesari et al [19] who reported that theadministration of leaf extracts ofM koenigii causes a decreasein the level of LDL of normal and diabetic rats So theseessential oils play a beneficiary role

Sodium and potassium are electrolytes present in bloodand other body fluids These help keep the water andelectrolyte balance of the body and are also important inthe proper functioning of nerves and muscles The hormonealdosterone controls the level of sodium and potassium in thebody The results revealed that all the selected essential oilshave potential to elevate blood sodium level Statistical analy-sis indicated a significant elevation in animals treated withAmarmelos C reticulataBlanco S laureolaM paniculata andB albiflora Similar findings have been reported by Odeyemiet al [23] who concluded that all essential oils may not besafe as these can lead to leakage of electrolytes from the cellsHowever in current study blood potassium level remainedunaltered

In conclusion although EOs were found to have somefavorable insinuations (decrease in triglycerides and LDLalong with no adverse effects on LFT and RFT) they mayleave specific undesired effects (eg increase in urea (Zarmatum) depletion in HDL (B albiflora) induction inerythropoiesis above normal level (M koenigii S laureola

and B albiflora) and increase in sodium (A marmelos CreticulataBlanco S laureolaM paniculata andB albiflora))which point to their toxic potential

5 Conclusion

As reduction in triglycerides is considered as beneficial andchange in urea without alteration in creatinine is not consid-ered as toxic output of any tested material it could be con-cluded that all oils except B albiflora can be considered safefor internal use with caution and sodium level may be contin-uously monitored B albiflora affected many markers includ-ing RBC MCV triglycerides HDL LDL urea and sodium

Data Availability

The data used to support the findings of this study areincluded within the article

Conflicts of Interest

The authors declare that they have no conflicts of interest

References

[1] H R Chitme R Chandra and S Kaushik ldquoStudies on anti-diarrheal activity of Calotropis gigantea R Br in experimentalanimalsrdquo Journal of Pharmacy and Pharmaceutical Sciences vol7 no 1 pp 70ndash75 2004

[2] H-S Kim ldquoDo not put too much value on conventionalmedicinesrdquo Journal of Ethnopharmacology vol 100 no 1-2 pp37ndash39 2005

[3] F Bakkali S Averbeck D Averbeck and M Idaomar ldquoBio-logical effects of essential oilsmdasha reviewrdquo Food and ChemicalToxicology vol 46 no 2 pp 446ndash475 2008

[4] S A Aziz M A Sukari M Rahmani M Kitajima NAimi and N J Ahpandi ldquoCoumarins from murraya pan-iculata (rutaceae) (koumarin daripada murraya paniculata(rutaceae))rdquoMalaysian Journal of Analytical Sciences vol 14 no1 pp 1ndash5 2010

[5] M Mcguffin C Hobbs R Upton and A Goldberg AmericanHerbal Products 1997 American Herbal Products

[6] S L Ngahang Kamte F Ranjbarian K Cianfaglione et alldquoIdentification of highly effective antitrypanosomal compoundsin essential oils from the Apiaceae familyrdquo Ecotoxicology andEnvironmental Safety vol 156 pp 154ndash165 2018

[7] Y Wang Y Zhang Y Shi X Pan Y Lu and P CaoldquoAntibacterial effects of cinnamon (Cinnamomum zeylanicum)bark essential oil on Porphyromonas gingivalisrdquo MicrobialPathogenesis vol 116 pp 26ndash32 2018

[8] R Tardugno F Pellati R Iseppi M Bondi G Bruzzesi and SBenvenuti ldquoPhytochemical composition and in vitro screeningof the antimicrobial activity of essential oils on oral pathogenicbacteriardquo Natural Product Research vol 32 no 5 pp 544ndash5512018

[9] A Veerappan S Miyazaki M Kadarkaraisamy and D Ran-ganathan ldquoAcute and subacute toxicity studies of Aegle marme-los Corr an Indianmedicinal plantrdquo Phytomedicine vol 14 no2-3 pp 209ndash215 2007

[10] J Palis and G B Segel ldquoDevelopmental biology of erythro-poiesisrdquo Blood Reviews vol 12 no 2 pp 106ndash114 1998

Evidence-Based Complementary and Alternative Medicine 7

[11] A Brum S A Pereira L Cardoso et al ldquoBlood biochemicalparameters and melanomacrophage centers in Nile tilapiafed essential oils of clove basil and gingerrdquo Fish amp ShellfishImmunology vol 74 pp 444ndash449 2018

[12] M Suwalsky P Vargas M Avello F Villena and C PSotomayor ldquoHuman erythrocytes are affected in vitro byflavonoids of Aristotelia chilensis (Maqui) leavesrdquo InternationalJournal of Pharmaceutics vol 363 no 1-2 pp 85ndash90 2008

[13] O T Oyesina A M Salihu O L Aribidesi and A D AdedayoldquoEffect of essential oil of the leaves of Eucalyptus globulesonheamatological parameters of wistar ratsrdquo African Journal ofBiochemistry Research vol 6 no 4 pp 46ndash49 2012

[14] W Janssen Forensic Histopathology Springer Berlin Germany1984

[15] S O Abarikwu R C Njoku and C L Onuah ldquoAged coconutoil with a high peroxide value induces oxidative stress and tissuedamage in mercury-treated ratsrdquo Journal of Basic and ClinicalPhysiology and Pharmacology 2018

[16] R Nisha S R Srinivasa Kannan K Thanga Mariappan andP Jagatha ldquoBiochemical evaluation of creatinine and urea inpatients with renal failure undergoing hemodialysisrdquo Journal ofClinical Pathology and LaboratoryMedicine vol 1 no 2 pp 1ndash52017

[17] B R Zirkin and V Papadopoulos ldquoLeydig Cells FormationFunction and Regulationrdquo Biology of Reproduction 2018

[18] M Rajadurai and P S M Prince ldquoComparative effects of Aeglemarmelos extract and alpha-tocopherol on serum lipids lipidperoxides and cardiac enzyme levels in rats with isoproterenol-induced myocardial infarctionrdquo Singapore Medical Journal vol46 no 2 pp 78ndash81 2005

[19] A N Kesari S Kesari S K Singh R K Gupta and GWatal ldquoStudies on the glycemic and lipidemic effect of Murrayakoenigii in experimental animalsrdquo Journal of Ethnopharmacol-ogy vol 112 no 2 pp 305ndash311 2007

[20] S Saljoughian S Roohinejad A E-D A Bekhit et al ldquoTheeffects of food essential oils on cardiovascular diseases Areviewrdquo Critical Reviews in Food Science and Nutrition vol 10pp 1ndash18 2017

[21] VD Gooty A R Sinaiko J R Ryder D R Dengel D R Jacobsand J Steinberger ldquoAssociation Between Carotid Intima MediaThickness Age and Cardiovascular Risk Factors in Childrenand Adolescentsrdquo Metabolic Syndrome and Related Disordersvol 16 no 3 pp 122ndash126 2018

[22] A Mancini E Imperlini E Nigro et al ldquoBiological andnutritional properties of palm oil and palmitic acid Effects onhealthrdquoMolecules vol 20 no 9 pp 17339ndash17361 2015

[23] O O Odeyemi M T Yakubu P J Masika and A J AfolayanldquoToxicological evaluation of the essential oil from menthalongifolia l subsp capensis leaves in ratsrdquo Journal of MedicinalFood vol 12 no 3 pp 669ndash674 2009

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Evidence-Based Complementary andAlternative Medicine

Volume 2018Hindawiwwwhindawicom

Submit your manuscripts atwwwhindawicom

4 Evidence-Based Complementary and Alternative Medicine

0

10

20

30

40

50

60

A-1 A-2 A-3 A-4 A-5 A-6 A-7

UN

ITS

VALU

ES IN

RES

PECT

IVE

HEMATOLOGICAL PARAMETERS

ControlA marmelosM koenigiiC reticulata Blanco

Z armatumS laureolaM paniculataB albiflora

lowast

lowast

lowast

lowast

Figure 1 Comparison of hematological parameters in the essentialoil- (EO-) fed animals and non-EO-treated (control) animals (I)A-1 hemoglobin (mgdl) A-2 total erythrocyte count (times106120583l) A-3 total leucocyte count (times103120583l) A-4 packed cell volume () A-5 mean corpuscular volume (fLlowast10cell) A-6 mean corpuscularhemoglobin (pgcell) and A-7 mean corpuscular hemoglobin con-centration (gdl) Data is presented as mean plusmn SEM Asterisks showsignificant difference from control

UN

ITS

VALU

ES IN

RES

PECT

IVE

ControlA marmelosM koenigiiC reticulata Blanco

Z armatumS laureolaM paniculataB albiflora

0102030405060708090

B-1 B-2 B-3 B-4 B-5 B-6LIVER AND RENAL FUNCTION TESTS

lowast

lowast

lowast

lowast

lowast

Figure 2 Comparison of liver and renal function tests of essen-tial oil- (EO-) fed animals and control animals B-1 bilirubin(times100mgdl) B-2 alanine transaminase (IUL) and aspartatetransaminase (IUL) B-3 alkaline phosphatase (Udl) B-4 urea(mgdl) and B-5 creatinine (times100mgdl) Data are presented asmean plusmn SEM Asterisks on respective groups show significantdifference from control group at 119901 le 005

322 Liver Function Test and Renal Function Test There wasa nonsignificant decrease in bilirubin ALT AST and ALPlevels which are markers for liver injury An elevated level ofblood urea was observed in groups V VI VII and VIII with2580 3870 4731 and 7311 increase respectivelyCreatinine level remained unchanged in all treatment groups(Figure 2)

323 Lipid Profile Statistical analysis revealed a significantdepletion in triglycerides in groups II IV VI VII and

ControlA marmelosM koenigiiC reticulata

Z armatumS laureolaM paniculataB albiflora

0

50

100

150

200

Cholesterol Triglyceride HDL LDL

Valu

es (m

gdl

)

LIPID PROFILE

lowastlowast

lowast

lowast

lowast

lowast

lowast

Figure 3 Comparison of lipid profile (mgdL) in essential oil-(EO-) fed animals and non-EO-treated control animals (I) Data arepresented as mean plusmn SEM while asterisks on respective EO groupsshow significant difference from control at 119901 le 005

Valu

es in

resp

ectiv

e uni

ts

ControlA marmelosM koenigiiC reticulata

Z armatumS laureolaM paniculataB albiflora

0369

121518

Sodium (mmoledl) Potassium (mmoleL)Serum Electrolytes

lowastlowastlowast

lowastlowastlowast

Figure 4 Comparison of blood electrolytes in essential oil- (EO-)fed animals and control animals (I) Data are presented as mean plusmnSEM Asterisks on respective EO groups show significant differencefrom control at 119901 le 005

VIII The level increased by 4933 4849 3193 4147and 4615 respectively HDL and LDL levels significantlydecreased in group VIII by 3070 and 3045 respectively(Figure 3)

324 Electrolytes The results revealed that all the selectedoils have potential to elevate blood sodium level Statisticalanalysis indicated a significant difference in groups II IV VVI VII and VIII The increase in level was 1189 116516 1262 1747 and 1383 respectively There was noalteration observed in blood potassium level in any essentialoil fed group (Figure 4)

4 Discussion

The increased use of antimicrobial agents has resulted in thedevelopment of resistant strains of bacteria So there is a need

Evidence-Based Complementary and Alternative Medicine 5

to develop effective antimicrobial agents with new modes ofaction against these pathogenic microbes Essential oils havebeen recognized as a rich source of new bioactive secondarymetabolites which possess the potential of treatment ofmanyinfectious diseases In the current study the potential ofselected essential oils as antimicrobial agents was screenedagainst four resistant clinical isolates including two Gram-positive (S aureus and S epidermidis) and twoGram-negative(E coli andK pneumoniae) bacteriaThe antibacterial activitywas initially screened by using disc diffusion method andthen theminimum inhibitory concentration (MIC) andmin-imum bactericidal concentration (MBC) were determined byusing microdilution method

The results of the disc diffusion method revealed that theselected essential oils exhibit varying magnitudes of antibac-terial activity Almost all of the essential oils were activeagainst the four clinical isolates According to results Aeglemarmelos was found to possess the highest antimicrobialactivity against S aureus E coli and K pneumoniae whileBoenninghausenia albiflora shows highest activity against Sepidermidis All the oils were found to be more active againstGram-positive bacterial isolates (S aureus and S epidermidis)and were less active against Gram-negative bacteria (E coliand K pneumoniae) [7]

Results of MIC and MBC also revealed that all the oilswere more active against Gram-positive as compared toGram-negative bacterial isolates Zanthoxylum armatum andMurraya paniculata were the most active essential oils andwere active against all the selected isolates Citrus reticulataBlanco was the least active oil with low antibacterial activityGram-negative bacteria are more resistant to essential oilsthen Gram-positive bacteria due to permeability barrier pro-vided by extra lipopolysaccharidemembraneThe antimicro-bial activity of essential oils could be due to their hydrophobiccharacteristic due to which these oils are capable of breakingthe lipids of bacterial cell membrane and making them morepermeable [8]

The current study involved the investigation of hemato-logical and serological changes in rats fed with essential oilsThe hematological parameters can be influenced by toxicityof essential oils as these oils have ability to initiate the acutephase response [9] Significant difference was observed inRBC PCV and MCV levels between the treated and controlgroups However in most of the oil feeding groups the HbTEC TLC PCV MCV MCH and MCHC were in normalrange

Hemoglobin is a metalloprotein that contains iron andis present in red blood cells of all vertebrates It transportsoxygen from respiratory organs to the rest of the body Inthe current study the results indicate that the hemoglobinlevel nonsignificantly increased in oil-fed groups The reasonfor this slight increase in hemoglobin was an increase inthe number of red blood cells Erythrocytes are the mostcommon type of blood cells and are the main source oftransferring oxygen to the body tissues RBCs are formed inbone marrow and stimulated by decrease in oxygen which isdetected by kidneys [10] In the current study a significantelevation in TEC was noticed in the groups of animalswhich were treated with M koenigii S laureola and B

albiflora essential oils Studies had shown that morphologicalchanges occur in RBCs in response to various treatmentsby toxic agents [11] According to Suwalsky et al [12]Balbisia peduncularis extract caused changes in the normalerythrocyte morphology An elevation in total erythrocyteswas noticed in some EO-treated groups Packed cell volume(PCV) or hematocrit is the volume percentage of red bloodcells in blood PCV was slightly increased in all essential oil-fed groups but a significant elevation was observed only ingroup treatedwith essential oil ofM paniculataThis increasein PCV may be due to an increase in RBCs number in theoil-fed groups [13] This finding is supported by an elevatederythrocytes number in the same group Mean corpuscularvolume (MCV) is ameasure of the average red blood cell sizeThe results indicated significant decrease in the level of MCVin the group treated with essential oil of B albiflora Thisreduction in MCV was maybe due to decrease in size of redblood cell [13] There was no significant difference observedinMCHandMCHC in any oil-fed groupThat was because ofthe hemoglobin level whichwas in normal range in all groups

Leukocytes are involved in the defense mechanism ofthe body against foreign materials In the present studyno significant change was observed in any oil-fed groupwhen compared to control group All the groups indicatethe number of white blood cells in normal range This is theindication that the selected essential oils perhaps do not affectleucopoiesis or half-life of leukocytes

The serological parameters selected for the study includedthe liver function test renal function test lipid profileand serum electrolytes The selected essential oils cause anonsignificant depletion in the level of all themarkers for liverinjury which included bilirubin alanine aminotransferase(ALT) aspartate aminotransferase (AST) and alkaline Phos-phatase (ALP) This is an indication that all these oils do nothave toxic effects particularly in reference to liver functiontest

Renal function test is an indication of the state of kidneyUrea and creatinine are usually considered as markers withrespect to kidney function Urea is the nitrogenous wasteproduct of the body Raised level of blood urea is anindication of renal dysfunction An elevated level of bloodurea was observed in some of the essential oil-fed groupsSignificant elevation was observed in groups treated with Zarmatum S laureolaM paniculata and B albiflora essentialoils This increase in urea level may be due to epithelialnecrosis to the renal tubules [14] On the other hand bloodcreatinine level remained unaltered in all essential oi-fedgroups Indicating EO at this dose may cause partial toxicityto kidneys According to Abarikwu et al [15] whenever theurea level increased and creatinine level is reduced there is nokidney damage Nisha et al [16] reported that renal toxicityshould be considered only when creatinine and urea levelincreased parallel to each other

Lipid profile is the term given to the evaluation of totalcholesterol triglycerides high-density lipoproteins (HDL)and low-density lipoproteins (LDL) This test is mostly usedto identify hyperlipidemia which is risk factor for heartdiseases Cholesterol is an ester and is an important part ofmammalian cell membrane [17] It is formed in the liver

6 Evidence-Based Complementary and Alternative Medicine

In this study no variation was observed in cholesterol levelin any essential oil-fed group According to Rajadurai andPrince [18] the cholesterol level slightly decreases in rats intreatment withAeglemarmelos leaf extractsMurraya koenigiileaf extracts also decrease cholesterol level in rats [19]

Triglyceride is also an ester and is derived from glyceroland three fatty acids High triglyceride is directly related tocoronary heart disease Our results indicate a decrease intriglycerides level in all essential oil-fed groups as comparedto control group Statistical analysis indicated a significantdepletion in groups treated with A marmelos C reticulataBlanco S laureola M paniculata and B albiflora Ourfindings are in accordance with Rajadurai et al [18] andRajadurai et al [19] they reported that the leaf extracts of Amarmelos cause a decrease in triglycerides level in rat models

Recent studies suggest that triglyceride itself is inde-pendently related to coronary heart disease [20 21] andmost of the antihypercholesterolemic drugs do not decreasetriglycerides levels but plant extracts do These findingsindicate beneficial effects of essential oils instead of theirtoxicological potential

High-density lipoprotein (HDL) is also known as goodcholesterol It transports other lipids like cholesterol andtriglycerides in the blood stream Various studies have shownthat an increase in HDL-cholesterol is associated with adecrease in coronary risk [22] Statistical analysis indicateda significant decrease in HDL in animals treated with Balbiflora essential oil So this essential oil may be consideredas toxic because it is decreasing good cholesterol

Low-density lipoproteins (LDL) are also known as badcholesterol LDL collects in the walls of blood vessels andcauses the blockage of the arteries Higher LDL levels maycause sudden blood clot in an artery and increase risk of heartattack In present study significant difference was observedin animals treated with B albiflora Our findings are inaccordance with Kesari et al [19] who reported that theadministration of leaf extracts ofM koenigii causes a decreasein the level of LDL of normal and diabetic rats So theseessential oils play a beneficiary role

Sodium and potassium are electrolytes present in bloodand other body fluids These help keep the water andelectrolyte balance of the body and are also important inthe proper functioning of nerves and muscles The hormonealdosterone controls the level of sodium and potassium in thebody The results revealed that all the selected essential oilshave potential to elevate blood sodium level Statistical analy-sis indicated a significant elevation in animals treated withAmarmelos C reticulataBlanco S laureolaM paniculata andB albiflora Similar findings have been reported by Odeyemiet al [23] who concluded that all essential oils may not besafe as these can lead to leakage of electrolytes from the cellsHowever in current study blood potassium level remainedunaltered

In conclusion although EOs were found to have somefavorable insinuations (decrease in triglycerides and LDLalong with no adverse effects on LFT and RFT) they mayleave specific undesired effects (eg increase in urea (Zarmatum) depletion in HDL (B albiflora) induction inerythropoiesis above normal level (M koenigii S laureola

and B albiflora) and increase in sodium (A marmelos CreticulataBlanco S laureolaM paniculata andB albiflora))which point to their toxic potential

5 Conclusion

As reduction in triglycerides is considered as beneficial andchange in urea without alteration in creatinine is not consid-ered as toxic output of any tested material it could be con-cluded that all oils except B albiflora can be considered safefor internal use with caution and sodium level may be contin-uously monitored B albiflora affected many markers includ-ing RBC MCV triglycerides HDL LDL urea and sodium

Data Availability

The data used to support the findings of this study areincluded within the article

Conflicts of Interest

The authors declare that they have no conflicts of interest

References

[1] H R Chitme R Chandra and S Kaushik ldquoStudies on anti-diarrheal activity of Calotropis gigantea R Br in experimentalanimalsrdquo Journal of Pharmacy and Pharmaceutical Sciences vol7 no 1 pp 70ndash75 2004

[2] H-S Kim ldquoDo not put too much value on conventionalmedicinesrdquo Journal of Ethnopharmacology vol 100 no 1-2 pp37ndash39 2005

[3] F Bakkali S Averbeck D Averbeck and M Idaomar ldquoBio-logical effects of essential oilsmdasha reviewrdquo Food and ChemicalToxicology vol 46 no 2 pp 446ndash475 2008

[4] S A Aziz M A Sukari M Rahmani M Kitajima NAimi and N J Ahpandi ldquoCoumarins from murraya pan-iculata (rutaceae) (koumarin daripada murraya paniculata(rutaceae))rdquoMalaysian Journal of Analytical Sciences vol 14 no1 pp 1ndash5 2010

[5] M Mcguffin C Hobbs R Upton and A Goldberg AmericanHerbal Products 1997 American Herbal Products

[6] S L Ngahang Kamte F Ranjbarian K Cianfaglione et alldquoIdentification of highly effective antitrypanosomal compoundsin essential oils from the Apiaceae familyrdquo Ecotoxicology andEnvironmental Safety vol 156 pp 154ndash165 2018

[7] Y Wang Y Zhang Y Shi X Pan Y Lu and P CaoldquoAntibacterial effects of cinnamon (Cinnamomum zeylanicum)bark essential oil on Porphyromonas gingivalisrdquo MicrobialPathogenesis vol 116 pp 26ndash32 2018

[8] R Tardugno F Pellati R Iseppi M Bondi G Bruzzesi and SBenvenuti ldquoPhytochemical composition and in vitro screeningof the antimicrobial activity of essential oils on oral pathogenicbacteriardquo Natural Product Research vol 32 no 5 pp 544ndash5512018

[9] A Veerappan S Miyazaki M Kadarkaraisamy and D Ran-ganathan ldquoAcute and subacute toxicity studies of Aegle marme-los Corr an Indianmedicinal plantrdquo Phytomedicine vol 14 no2-3 pp 209ndash215 2007

[10] J Palis and G B Segel ldquoDevelopmental biology of erythro-poiesisrdquo Blood Reviews vol 12 no 2 pp 106ndash114 1998

Evidence-Based Complementary and Alternative Medicine 7

[11] A Brum S A Pereira L Cardoso et al ldquoBlood biochemicalparameters and melanomacrophage centers in Nile tilapiafed essential oils of clove basil and gingerrdquo Fish amp ShellfishImmunology vol 74 pp 444ndash449 2018

[12] M Suwalsky P Vargas M Avello F Villena and C PSotomayor ldquoHuman erythrocytes are affected in vitro byflavonoids of Aristotelia chilensis (Maqui) leavesrdquo InternationalJournal of Pharmaceutics vol 363 no 1-2 pp 85ndash90 2008

[13] O T Oyesina A M Salihu O L Aribidesi and A D AdedayoldquoEffect of essential oil of the leaves of Eucalyptus globulesonheamatological parameters of wistar ratsrdquo African Journal ofBiochemistry Research vol 6 no 4 pp 46ndash49 2012

[14] W Janssen Forensic Histopathology Springer Berlin Germany1984

[15] S O Abarikwu R C Njoku and C L Onuah ldquoAged coconutoil with a high peroxide value induces oxidative stress and tissuedamage in mercury-treated ratsrdquo Journal of Basic and ClinicalPhysiology and Pharmacology 2018

[16] R Nisha S R Srinivasa Kannan K Thanga Mariappan andP Jagatha ldquoBiochemical evaluation of creatinine and urea inpatients with renal failure undergoing hemodialysisrdquo Journal ofClinical Pathology and LaboratoryMedicine vol 1 no 2 pp 1ndash52017

[17] B R Zirkin and V Papadopoulos ldquoLeydig Cells FormationFunction and Regulationrdquo Biology of Reproduction 2018

[18] M Rajadurai and P S M Prince ldquoComparative effects of Aeglemarmelos extract and alpha-tocopherol on serum lipids lipidperoxides and cardiac enzyme levels in rats with isoproterenol-induced myocardial infarctionrdquo Singapore Medical Journal vol46 no 2 pp 78ndash81 2005

[19] A N Kesari S Kesari S K Singh R K Gupta and GWatal ldquoStudies on the glycemic and lipidemic effect of Murrayakoenigii in experimental animalsrdquo Journal of Ethnopharmacol-ogy vol 112 no 2 pp 305ndash311 2007

[20] S Saljoughian S Roohinejad A E-D A Bekhit et al ldquoTheeffects of food essential oils on cardiovascular diseases Areviewrdquo Critical Reviews in Food Science and Nutrition vol 10pp 1ndash18 2017

[21] VD Gooty A R Sinaiko J R Ryder D R Dengel D R Jacobsand J Steinberger ldquoAssociation Between Carotid Intima MediaThickness Age and Cardiovascular Risk Factors in Childrenand Adolescentsrdquo Metabolic Syndrome and Related Disordersvol 16 no 3 pp 122ndash126 2018

[22] A Mancini E Imperlini E Nigro et al ldquoBiological andnutritional properties of palm oil and palmitic acid Effects onhealthrdquoMolecules vol 20 no 9 pp 17339ndash17361 2015

[23] O O Odeyemi M T Yakubu P J Masika and A J AfolayanldquoToxicological evaluation of the essential oil from menthalongifolia l subsp capensis leaves in ratsrdquo Journal of MedicinalFood vol 12 no 3 pp 669ndash674 2009

Stem Cells International

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

MEDIATORSINFLAMMATION

of

EndocrinologyInternational Journal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Disease Markers

Hindawiwwwhindawicom Volume 2018

BioMed Research International

OncologyJournal of

Hindawiwwwhindawicom Volume 2013

Hindawiwwwhindawicom Volume 2018

Oxidative Medicine and Cellular Longevity

Hindawiwwwhindawicom Volume 2018

PPAR Research

Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom

The Scientific World Journal

Volume 2018

Immunology ResearchHindawiwwwhindawicom Volume 2018

Journal of

ObesityJournal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Computational and Mathematical Methods in Medicine

Hindawiwwwhindawicom Volume 2018

Behavioural Neurology

OphthalmologyJournal of

Hindawiwwwhindawicom Volume 2018

Diabetes ResearchJournal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Research and TreatmentAIDS

Hindawiwwwhindawicom Volume 2018

Gastroenterology Research and Practice

Hindawiwwwhindawicom Volume 2018

Parkinsonrsquos Disease

Evidence-Based Complementary andAlternative Medicine

Volume 2018Hindawiwwwhindawicom

Submit your manuscripts atwwwhindawicom

Evidence-Based Complementary and Alternative Medicine 5

to develop effective antimicrobial agents with new modes ofaction against these pathogenic microbes Essential oils havebeen recognized as a rich source of new bioactive secondarymetabolites which possess the potential of treatment ofmanyinfectious diseases In the current study the potential ofselected essential oils as antimicrobial agents was screenedagainst four resistant clinical isolates including two Gram-positive (S aureus and S epidermidis) and twoGram-negative(E coli andK pneumoniae) bacteriaThe antibacterial activitywas initially screened by using disc diffusion method andthen theminimum inhibitory concentration (MIC) andmin-imum bactericidal concentration (MBC) were determined byusing microdilution method

The results of the disc diffusion method revealed that theselected essential oils exhibit varying magnitudes of antibac-terial activity Almost all of the essential oils were activeagainst the four clinical isolates According to results Aeglemarmelos was found to possess the highest antimicrobialactivity against S aureus E coli and K pneumoniae whileBoenninghausenia albiflora shows highest activity against Sepidermidis All the oils were found to be more active againstGram-positive bacterial isolates (S aureus and S epidermidis)and were less active against Gram-negative bacteria (E coliand K pneumoniae) [7]

Results of MIC and MBC also revealed that all the oilswere more active against Gram-positive as compared toGram-negative bacterial isolates Zanthoxylum armatum andMurraya paniculata were the most active essential oils andwere active against all the selected isolates Citrus reticulataBlanco was the least active oil with low antibacterial activityGram-negative bacteria are more resistant to essential oilsthen Gram-positive bacteria due to permeability barrier pro-vided by extra lipopolysaccharidemembraneThe antimicro-bial activity of essential oils could be due to their hydrophobiccharacteristic due to which these oils are capable of breakingthe lipids of bacterial cell membrane and making them morepermeable [8]

The current study involved the investigation of hemato-logical and serological changes in rats fed with essential oilsThe hematological parameters can be influenced by toxicityof essential oils as these oils have ability to initiate the acutephase response [9] Significant difference was observed inRBC PCV and MCV levels between the treated and controlgroups However in most of the oil feeding groups the HbTEC TLC PCV MCV MCH and MCHC were in normalrange

Hemoglobin is a metalloprotein that contains iron andis present in red blood cells of all vertebrates It transportsoxygen from respiratory organs to the rest of the body Inthe current study the results indicate that the hemoglobinlevel nonsignificantly increased in oil-fed groups The reasonfor this slight increase in hemoglobin was an increase inthe number of red blood cells Erythrocytes are the mostcommon type of blood cells and are the main source oftransferring oxygen to the body tissues RBCs are formed inbone marrow and stimulated by decrease in oxygen which isdetected by kidneys [10] In the current study a significantelevation in TEC was noticed in the groups of animalswhich were treated with M koenigii S laureola and B

albiflora essential oils Studies had shown that morphologicalchanges occur in RBCs in response to various treatmentsby toxic agents [11] According to Suwalsky et al [12]Balbisia peduncularis extract caused changes in the normalerythrocyte morphology An elevation in total erythrocyteswas noticed in some EO-treated groups Packed cell volume(PCV) or hematocrit is the volume percentage of red bloodcells in blood PCV was slightly increased in all essential oil-fed groups but a significant elevation was observed only ingroup treatedwith essential oil ofM paniculataThis increasein PCV may be due to an increase in RBCs number in theoil-fed groups [13] This finding is supported by an elevatederythrocytes number in the same group Mean corpuscularvolume (MCV) is ameasure of the average red blood cell sizeThe results indicated significant decrease in the level of MCVin the group treated with essential oil of B albiflora Thisreduction in MCV was maybe due to decrease in size of redblood cell [13] There was no significant difference observedinMCHandMCHC in any oil-fed groupThat was because ofthe hemoglobin level whichwas in normal range in all groups

Leukocytes are involved in the defense mechanism ofthe body against foreign materials In the present studyno significant change was observed in any oil-fed groupwhen compared to control group All the groups indicatethe number of white blood cells in normal range This is theindication that the selected essential oils perhaps do not affectleucopoiesis or half-life of leukocytes

The serological parameters selected for the study includedthe liver function test renal function test lipid profileand serum electrolytes The selected essential oils cause anonsignificant depletion in the level of all themarkers for liverinjury which included bilirubin alanine aminotransferase(ALT) aspartate aminotransferase (AST) and alkaline Phos-phatase (ALP) This is an indication that all these oils do nothave toxic effects particularly in reference to liver functiontest

Renal function test is an indication of the state of kidneyUrea and creatinine are usually considered as markers withrespect to kidney function Urea is the nitrogenous wasteproduct of the body Raised level of blood urea is anindication of renal dysfunction An elevated level of bloodurea was observed in some of the essential oil-fed groupsSignificant elevation was observed in groups treated with Zarmatum S laureolaM paniculata and B albiflora essentialoils This increase in urea level may be due to epithelialnecrosis to the renal tubules [14] On the other hand bloodcreatinine level remained unaltered in all essential oi-fedgroups Indicating EO at this dose may cause partial toxicityto kidneys According to Abarikwu et al [15] whenever theurea level increased and creatinine level is reduced there is nokidney damage Nisha et al [16] reported that renal toxicityshould be considered only when creatinine and urea levelincreased parallel to each other

Lipid profile is the term given to the evaluation of totalcholesterol triglycerides high-density lipoproteins (HDL)and low-density lipoproteins (LDL) This test is mostly usedto identify hyperlipidemia which is risk factor for heartdiseases Cholesterol is an ester and is an important part ofmammalian cell membrane [17] It is formed in the liver

6 Evidence-Based Complementary and Alternative Medicine

In this study no variation was observed in cholesterol levelin any essential oil-fed group According to Rajadurai andPrince [18] the cholesterol level slightly decreases in rats intreatment withAeglemarmelos leaf extractsMurraya koenigiileaf extracts also decrease cholesterol level in rats [19]

Triglyceride is also an ester and is derived from glyceroland three fatty acids High triglyceride is directly related tocoronary heart disease Our results indicate a decrease intriglycerides level in all essential oil-fed groups as comparedto control group Statistical analysis indicated a significantdepletion in groups treated with A marmelos C reticulataBlanco S laureola M paniculata and B albiflora Ourfindings are in accordance with Rajadurai et al [18] andRajadurai et al [19] they reported that the leaf extracts of Amarmelos cause a decrease in triglycerides level in rat models

Recent studies suggest that triglyceride itself is inde-pendently related to coronary heart disease [20 21] andmost of the antihypercholesterolemic drugs do not decreasetriglycerides levels but plant extracts do These findingsindicate beneficial effects of essential oils instead of theirtoxicological potential

High-density lipoprotein (HDL) is also known as goodcholesterol It transports other lipids like cholesterol andtriglycerides in the blood stream Various studies have shownthat an increase in HDL-cholesterol is associated with adecrease in coronary risk [22] Statistical analysis indicateda significant decrease in HDL in animals treated with Balbiflora essential oil So this essential oil may be consideredas toxic because it is decreasing good cholesterol

Low-density lipoproteins (LDL) are also known as badcholesterol LDL collects in the walls of blood vessels andcauses the blockage of the arteries Higher LDL levels maycause sudden blood clot in an artery and increase risk of heartattack In present study significant difference was observedin animals treated with B albiflora Our findings are inaccordance with Kesari et al [19] who reported that theadministration of leaf extracts ofM koenigii causes a decreasein the level of LDL of normal and diabetic rats So theseessential oils play a beneficiary role

Sodium and potassium are electrolytes present in bloodand other body fluids These help keep the water andelectrolyte balance of the body and are also important inthe proper functioning of nerves and muscles The hormonealdosterone controls the level of sodium and potassium in thebody The results revealed that all the selected essential oilshave potential to elevate blood sodium level Statistical analy-sis indicated a significant elevation in animals treated withAmarmelos C reticulataBlanco S laureolaM paniculata andB albiflora Similar findings have been reported by Odeyemiet al [23] who concluded that all essential oils may not besafe as these can lead to leakage of electrolytes from the cellsHowever in current study blood potassium level remainedunaltered

In conclusion although EOs were found to have somefavorable insinuations (decrease in triglycerides and LDLalong with no adverse effects on LFT and RFT) they mayleave specific undesired effects (eg increase in urea (Zarmatum) depletion in HDL (B albiflora) induction inerythropoiesis above normal level (M koenigii S laureola

and B albiflora) and increase in sodium (A marmelos CreticulataBlanco S laureolaM paniculata andB albiflora))which point to their toxic potential

5 Conclusion

As reduction in triglycerides is considered as beneficial andchange in urea without alteration in creatinine is not consid-ered as toxic output of any tested material it could be con-cluded that all oils except B albiflora can be considered safefor internal use with caution and sodium level may be contin-uously monitored B albiflora affected many markers includ-ing RBC MCV triglycerides HDL LDL urea and sodium

Data Availability

The data used to support the findings of this study areincluded within the article

Conflicts of Interest

The authors declare that they have no conflicts of interest

References

[1] H R Chitme R Chandra and S Kaushik ldquoStudies on anti-diarrheal activity of Calotropis gigantea R Br in experimentalanimalsrdquo Journal of Pharmacy and Pharmaceutical Sciences vol7 no 1 pp 70ndash75 2004

[2] H-S Kim ldquoDo not put too much value on conventionalmedicinesrdquo Journal of Ethnopharmacology vol 100 no 1-2 pp37ndash39 2005

[3] F Bakkali S Averbeck D Averbeck and M Idaomar ldquoBio-logical effects of essential oilsmdasha reviewrdquo Food and ChemicalToxicology vol 46 no 2 pp 446ndash475 2008

[4] S A Aziz M A Sukari M Rahmani M Kitajima NAimi and N J Ahpandi ldquoCoumarins from murraya pan-iculata (rutaceae) (koumarin daripada murraya paniculata(rutaceae))rdquoMalaysian Journal of Analytical Sciences vol 14 no1 pp 1ndash5 2010

[5] M Mcguffin C Hobbs R Upton and A Goldberg AmericanHerbal Products 1997 American Herbal Products

[6] S L Ngahang Kamte F Ranjbarian K Cianfaglione et alldquoIdentification of highly effective antitrypanosomal compoundsin essential oils from the Apiaceae familyrdquo Ecotoxicology andEnvironmental Safety vol 156 pp 154ndash165 2018

[7] Y Wang Y Zhang Y Shi X Pan Y Lu and P CaoldquoAntibacterial effects of cinnamon (Cinnamomum zeylanicum)bark essential oil on Porphyromonas gingivalisrdquo MicrobialPathogenesis vol 116 pp 26ndash32 2018

[8] R Tardugno F Pellati R Iseppi M Bondi G Bruzzesi and SBenvenuti ldquoPhytochemical composition and in vitro screeningof the antimicrobial activity of essential oils on oral pathogenicbacteriardquo Natural Product Research vol 32 no 5 pp 544ndash5512018

[9] A Veerappan S Miyazaki M Kadarkaraisamy and D Ran-ganathan ldquoAcute and subacute toxicity studies of Aegle marme-los Corr an Indianmedicinal plantrdquo Phytomedicine vol 14 no2-3 pp 209ndash215 2007

[10] J Palis and G B Segel ldquoDevelopmental biology of erythro-poiesisrdquo Blood Reviews vol 12 no 2 pp 106ndash114 1998

Evidence-Based Complementary and Alternative Medicine 7

[11] A Brum S A Pereira L Cardoso et al ldquoBlood biochemicalparameters and melanomacrophage centers in Nile tilapiafed essential oils of clove basil and gingerrdquo Fish amp ShellfishImmunology vol 74 pp 444ndash449 2018

[12] M Suwalsky P Vargas M Avello F Villena and C PSotomayor ldquoHuman erythrocytes are affected in vitro byflavonoids of Aristotelia chilensis (Maqui) leavesrdquo InternationalJournal of Pharmaceutics vol 363 no 1-2 pp 85ndash90 2008

[13] O T Oyesina A M Salihu O L Aribidesi and A D AdedayoldquoEffect of essential oil of the leaves of Eucalyptus globulesonheamatological parameters of wistar ratsrdquo African Journal ofBiochemistry Research vol 6 no 4 pp 46ndash49 2012

[14] W Janssen Forensic Histopathology Springer Berlin Germany1984

[15] S O Abarikwu R C Njoku and C L Onuah ldquoAged coconutoil with a high peroxide value induces oxidative stress and tissuedamage in mercury-treated ratsrdquo Journal of Basic and ClinicalPhysiology and Pharmacology 2018

[16] R Nisha S R Srinivasa Kannan K Thanga Mariappan andP Jagatha ldquoBiochemical evaluation of creatinine and urea inpatients with renal failure undergoing hemodialysisrdquo Journal ofClinical Pathology and LaboratoryMedicine vol 1 no 2 pp 1ndash52017

[17] B R Zirkin and V Papadopoulos ldquoLeydig Cells FormationFunction and Regulationrdquo Biology of Reproduction 2018

[18] M Rajadurai and P S M Prince ldquoComparative effects of Aeglemarmelos extract and alpha-tocopherol on serum lipids lipidperoxides and cardiac enzyme levels in rats with isoproterenol-induced myocardial infarctionrdquo Singapore Medical Journal vol46 no 2 pp 78ndash81 2005

[19] A N Kesari S Kesari S K Singh R K Gupta and GWatal ldquoStudies on the glycemic and lipidemic effect of Murrayakoenigii in experimental animalsrdquo Journal of Ethnopharmacol-ogy vol 112 no 2 pp 305ndash311 2007

[20] S Saljoughian S Roohinejad A E-D A Bekhit et al ldquoTheeffects of food essential oils on cardiovascular diseases Areviewrdquo Critical Reviews in Food Science and Nutrition vol 10pp 1ndash18 2017

[21] VD Gooty A R Sinaiko J R Ryder D R Dengel D R Jacobsand J Steinberger ldquoAssociation Between Carotid Intima MediaThickness Age and Cardiovascular Risk Factors in Childrenand Adolescentsrdquo Metabolic Syndrome and Related Disordersvol 16 no 3 pp 122ndash126 2018

[22] A Mancini E Imperlini E Nigro et al ldquoBiological andnutritional properties of palm oil and palmitic acid Effects onhealthrdquoMolecules vol 20 no 9 pp 17339ndash17361 2015

[23] O O Odeyemi M T Yakubu P J Masika and A J AfolayanldquoToxicological evaluation of the essential oil from menthalongifolia l subsp capensis leaves in ratsrdquo Journal of MedicinalFood vol 12 no 3 pp 669ndash674 2009

Stem Cells International

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

MEDIATORSINFLAMMATION

of

EndocrinologyInternational Journal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Disease Markers

Hindawiwwwhindawicom Volume 2018

BioMed Research International

OncologyJournal of

Hindawiwwwhindawicom Volume 2013

Hindawiwwwhindawicom Volume 2018

Oxidative Medicine and Cellular Longevity

Hindawiwwwhindawicom Volume 2018

PPAR Research

Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom

The Scientific World Journal

Volume 2018

Immunology ResearchHindawiwwwhindawicom Volume 2018

Journal of

ObesityJournal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Computational and Mathematical Methods in Medicine

Hindawiwwwhindawicom Volume 2018

Behavioural Neurology

OphthalmologyJournal of

Hindawiwwwhindawicom Volume 2018

Diabetes ResearchJournal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Research and TreatmentAIDS

Hindawiwwwhindawicom Volume 2018

Gastroenterology Research and Practice

Hindawiwwwhindawicom Volume 2018

Parkinsonrsquos Disease

Evidence-Based Complementary andAlternative Medicine

Volume 2018Hindawiwwwhindawicom

Submit your manuscripts atwwwhindawicom

6 Evidence-Based Complementary and Alternative Medicine

In this study no variation was observed in cholesterol levelin any essential oil-fed group According to Rajadurai andPrince [18] the cholesterol level slightly decreases in rats intreatment withAeglemarmelos leaf extractsMurraya koenigiileaf extracts also decrease cholesterol level in rats [19]

Triglyceride is also an ester and is derived from glyceroland three fatty acids High triglyceride is directly related tocoronary heart disease Our results indicate a decrease intriglycerides level in all essential oil-fed groups as comparedto control group Statistical analysis indicated a significantdepletion in groups treated with A marmelos C reticulataBlanco S laureola M paniculata and B albiflora Ourfindings are in accordance with Rajadurai et al [18] andRajadurai et al [19] they reported that the leaf extracts of Amarmelos cause a decrease in triglycerides level in rat models

Recent studies suggest that triglyceride itself is inde-pendently related to coronary heart disease [20 21] andmost of the antihypercholesterolemic drugs do not decreasetriglycerides levels but plant extracts do These findingsindicate beneficial effects of essential oils instead of theirtoxicological potential

High-density lipoprotein (HDL) is also known as goodcholesterol It transports other lipids like cholesterol andtriglycerides in the blood stream Various studies have shownthat an increase in HDL-cholesterol is associated with adecrease in coronary risk [22] Statistical analysis indicateda significant decrease in HDL in animals treated with Balbiflora essential oil So this essential oil may be consideredas toxic because it is decreasing good cholesterol

Low-density lipoproteins (LDL) are also known as badcholesterol LDL collects in the walls of blood vessels andcauses the blockage of the arteries Higher LDL levels maycause sudden blood clot in an artery and increase risk of heartattack In present study significant difference was observedin animals treated with B albiflora Our findings are inaccordance with Kesari et al [19] who reported that theadministration of leaf extracts ofM koenigii causes a decreasein the level of LDL of normal and diabetic rats So theseessential oils play a beneficiary role

Sodium and potassium are electrolytes present in bloodand other body fluids These help keep the water andelectrolyte balance of the body and are also important inthe proper functioning of nerves and muscles The hormonealdosterone controls the level of sodium and potassium in thebody The results revealed that all the selected essential oilshave potential to elevate blood sodium level Statistical analy-sis indicated a significant elevation in animals treated withAmarmelos C reticulataBlanco S laureolaM paniculata andB albiflora Similar findings have been reported by Odeyemiet al [23] who concluded that all essential oils may not besafe as these can lead to leakage of electrolytes from the cellsHowever in current study blood potassium level remainedunaltered

In conclusion although EOs were found to have somefavorable insinuations (decrease in triglycerides and LDLalong with no adverse effects on LFT and RFT) they mayleave specific undesired effects (eg increase in urea (Zarmatum) depletion in HDL (B albiflora) induction inerythropoiesis above normal level (M koenigii S laureola

and B albiflora) and increase in sodium (A marmelos CreticulataBlanco S laureolaM paniculata andB albiflora))which point to their toxic potential

5 Conclusion

As reduction in triglycerides is considered as beneficial andchange in urea without alteration in creatinine is not consid-ered as toxic output of any tested material it could be con-cluded that all oils except B albiflora can be considered safefor internal use with caution and sodium level may be contin-uously monitored B albiflora affected many markers includ-ing RBC MCV triglycerides HDL LDL urea and sodium

Data Availability

The data used to support the findings of this study areincluded within the article

Conflicts of Interest

The authors declare that they have no conflicts of interest

References

[1] H R Chitme R Chandra and S Kaushik ldquoStudies on anti-diarrheal activity of Calotropis gigantea R Br in experimentalanimalsrdquo Journal of Pharmacy and Pharmaceutical Sciences vol7 no 1 pp 70ndash75 2004

[2] H-S Kim ldquoDo not put too much value on conventionalmedicinesrdquo Journal of Ethnopharmacology vol 100 no 1-2 pp37ndash39 2005

[3] F Bakkali S Averbeck D Averbeck and M Idaomar ldquoBio-logical effects of essential oilsmdasha reviewrdquo Food and ChemicalToxicology vol 46 no 2 pp 446ndash475 2008

[4] S A Aziz M A Sukari M Rahmani M Kitajima NAimi and N J Ahpandi ldquoCoumarins from murraya pan-iculata (rutaceae) (koumarin daripada murraya paniculata(rutaceae))rdquoMalaysian Journal of Analytical Sciences vol 14 no1 pp 1ndash5 2010

[5] M Mcguffin C Hobbs R Upton and A Goldberg AmericanHerbal Products 1997 American Herbal Products

[6] S L Ngahang Kamte F Ranjbarian K Cianfaglione et alldquoIdentification of highly effective antitrypanosomal compoundsin essential oils from the Apiaceae familyrdquo Ecotoxicology andEnvironmental Safety vol 156 pp 154ndash165 2018

[7] Y Wang Y Zhang Y Shi X Pan Y Lu and P CaoldquoAntibacterial effects of cinnamon (Cinnamomum zeylanicum)bark essential oil on Porphyromonas gingivalisrdquo MicrobialPathogenesis vol 116 pp 26ndash32 2018

[8] R Tardugno F Pellati R Iseppi M Bondi G Bruzzesi and SBenvenuti ldquoPhytochemical composition and in vitro screeningof the antimicrobial activity of essential oils on oral pathogenicbacteriardquo Natural Product Research vol 32 no 5 pp 544ndash5512018

[9] A Veerappan S Miyazaki M Kadarkaraisamy and D Ran-ganathan ldquoAcute and subacute toxicity studies of Aegle marme-los Corr an Indianmedicinal plantrdquo Phytomedicine vol 14 no2-3 pp 209ndash215 2007

[10] J Palis and G B Segel ldquoDevelopmental biology of erythro-poiesisrdquo Blood Reviews vol 12 no 2 pp 106ndash114 1998

Evidence-Based Complementary and Alternative Medicine 7

[11] A Brum S A Pereira L Cardoso et al ldquoBlood biochemicalparameters and melanomacrophage centers in Nile tilapiafed essential oils of clove basil and gingerrdquo Fish amp ShellfishImmunology vol 74 pp 444ndash449 2018

[12] M Suwalsky P Vargas M Avello F Villena and C PSotomayor ldquoHuman erythrocytes are affected in vitro byflavonoids of Aristotelia chilensis (Maqui) leavesrdquo InternationalJournal of Pharmaceutics vol 363 no 1-2 pp 85ndash90 2008

[13] O T Oyesina A M Salihu O L Aribidesi and A D AdedayoldquoEffect of essential oil of the leaves of Eucalyptus globulesonheamatological parameters of wistar ratsrdquo African Journal ofBiochemistry Research vol 6 no 4 pp 46ndash49 2012

[14] W Janssen Forensic Histopathology Springer Berlin Germany1984

[15] S O Abarikwu R C Njoku and C L Onuah ldquoAged coconutoil with a high peroxide value induces oxidative stress and tissuedamage in mercury-treated ratsrdquo Journal of Basic and ClinicalPhysiology and Pharmacology 2018

[16] R Nisha S R Srinivasa Kannan K Thanga Mariappan andP Jagatha ldquoBiochemical evaluation of creatinine and urea inpatients with renal failure undergoing hemodialysisrdquo Journal ofClinical Pathology and LaboratoryMedicine vol 1 no 2 pp 1ndash52017

[17] B R Zirkin and V Papadopoulos ldquoLeydig Cells FormationFunction and Regulationrdquo Biology of Reproduction 2018

[18] M Rajadurai and P S M Prince ldquoComparative effects of Aeglemarmelos extract and alpha-tocopherol on serum lipids lipidperoxides and cardiac enzyme levels in rats with isoproterenol-induced myocardial infarctionrdquo Singapore Medical Journal vol46 no 2 pp 78ndash81 2005

[19] A N Kesari S Kesari S K Singh R K Gupta and GWatal ldquoStudies on the glycemic and lipidemic effect of Murrayakoenigii in experimental animalsrdquo Journal of Ethnopharmacol-ogy vol 112 no 2 pp 305ndash311 2007

[20] S Saljoughian S Roohinejad A E-D A Bekhit et al ldquoTheeffects of food essential oils on cardiovascular diseases Areviewrdquo Critical Reviews in Food Science and Nutrition vol 10pp 1ndash18 2017

[21] VD Gooty A R Sinaiko J R Ryder D R Dengel D R Jacobsand J Steinberger ldquoAssociation Between Carotid Intima MediaThickness Age and Cardiovascular Risk Factors in Childrenand Adolescentsrdquo Metabolic Syndrome and Related Disordersvol 16 no 3 pp 122ndash126 2018

[22] A Mancini E Imperlini E Nigro et al ldquoBiological andnutritional properties of palm oil and palmitic acid Effects onhealthrdquoMolecules vol 20 no 9 pp 17339ndash17361 2015

[23] O O Odeyemi M T Yakubu P J Masika and A J AfolayanldquoToxicological evaluation of the essential oil from menthalongifolia l subsp capensis leaves in ratsrdquo Journal of MedicinalFood vol 12 no 3 pp 669ndash674 2009

Stem Cells International

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

MEDIATORSINFLAMMATION

of

EndocrinologyInternational Journal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Disease Markers

Hindawiwwwhindawicom Volume 2018

BioMed Research International

OncologyJournal of

Hindawiwwwhindawicom Volume 2013

Hindawiwwwhindawicom Volume 2018

Oxidative Medicine and Cellular Longevity

Hindawiwwwhindawicom Volume 2018

PPAR Research

Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom

The Scientific World Journal

Volume 2018

Immunology ResearchHindawiwwwhindawicom Volume 2018

Journal of

ObesityJournal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Computational and Mathematical Methods in Medicine

Hindawiwwwhindawicom Volume 2018

Behavioural Neurology

OphthalmologyJournal of

Hindawiwwwhindawicom Volume 2018

Diabetes ResearchJournal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Research and TreatmentAIDS

Hindawiwwwhindawicom Volume 2018

Gastroenterology Research and Practice

Hindawiwwwhindawicom Volume 2018

Parkinsonrsquos Disease

Evidence-Based Complementary andAlternative Medicine

Volume 2018Hindawiwwwhindawicom

Submit your manuscripts atwwwhindawicom

Evidence-Based Complementary and Alternative Medicine 7

[11] A Brum S A Pereira L Cardoso et al ldquoBlood biochemicalparameters and melanomacrophage centers in Nile tilapiafed essential oils of clove basil and gingerrdquo Fish amp ShellfishImmunology vol 74 pp 444ndash449 2018

[12] M Suwalsky P Vargas M Avello F Villena and C PSotomayor ldquoHuman erythrocytes are affected in vitro byflavonoids of Aristotelia chilensis (Maqui) leavesrdquo InternationalJournal of Pharmaceutics vol 363 no 1-2 pp 85ndash90 2008

[13] O T Oyesina A M Salihu O L Aribidesi and A D AdedayoldquoEffect of essential oil of the leaves of Eucalyptus globulesonheamatological parameters of wistar ratsrdquo African Journal ofBiochemistry Research vol 6 no 4 pp 46ndash49 2012

[14] W Janssen Forensic Histopathology Springer Berlin Germany1984

[15] S O Abarikwu R C Njoku and C L Onuah ldquoAged coconutoil with a high peroxide value induces oxidative stress and tissuedamage in mercury-treated ratsrdquo Journal of Basic and ClinicalPhysiology and Pharmacology 2018

[16] R Nisha S R Srinivasa Kannan K Thanga Mariappan andP Jagatha ldquoBiochemical evaluation of creatinine and urea inpatients with renal failure undergoing hemodialysisrdquo Journal ofClinical Pathology and LaboratoryMedicine vol 1 no 2 pp 1ndash52017

[17] B R Zirkin and V Papadopoulos ldquoLeydig Cells FormationFunction and Regulationrdquo Biology of Reproduction 2018

[18] M Rajadurai and P S M Prince ldquoComparative effects of Aeglemarmelos extract and alpha-tocopherol on serum lipids lipidperoxides and cardiac enzyme levels in rats with isoproterenol-induced myocardial infarctionrdquo Singapore Medical Journal vol46 no 2 pp 78ndash81 2005

[19] A N Kesari S Kesari S K Singh R K Gupta and GWatal ldquoStudies on the glycemic and lipidemic effect of Murrayakoenigii in experimental animalsrdquo Journal of Ethnopharmacol-ogy vol 112 no 2 pp 305ndash311 2007

[20] S Saljoughian S Roohinejad A E-D A Bekhit et al ldquoTheeffects of food essential oils on cardiovascular diseases Areviewrdquo Critical Reviews in Food Science and Nutrition vol 10pp 1ndash18 2017

[21] VD Gooty A R Sinaiko J R Ryder D R Dengel D R Jacobsand J Steinberger ldquoAssociation Between Carotid Intima MediaThickness Age and Cardiovascular Risk Factors in Childrenand Adolescentsrdquo Metabolic Syndrome and Related Disordersvol 16 no 3 pp 122ndash126 2018

[22] A Mancini E Imperlini E Nigro et al ldquoBiological andnutritional properties of palm oil and palmitic acid Effects onhealthrdquoMolecules vol 20 no 9 pp 17339ndash17361 2015

[23] O O Odeyemi M T Yakubu P J Masika and A J AfolayanldquoToxicological evaluation of the essential oil from menthalongifolia l subsp capensis leaves in ratsrdquo Journal of MedicinalFood vol 12 no 3 pp 669ndash674 2009

Stem Cells International

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

MEDIATORSINFLAMMATION

of

EndocrinologyInternational Journal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Disease Markers

Hindawiwwwhindawicom Volume 2018

BioMed Research International

OncologyJournal of

Hindawiwwwhindawicom Volume 2013

Hindawiwwwhindawicom Volume 2018

Oxidative Medicine and Cellular Longevity

Hindawiwwwhindawicom Volume 2018

PPAR Research

Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom

The Scientific World Journal

Volume 2018

Immunology ResearchHindawiwwwhindawicom Volume 2018

Journal of

ObesityJournal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Computational and Mathematical Methods in Medicine

Hindawiwwwhindawicom Volume 2018

Behavioural Neurology

OphthalmologyJournal of

Hindawiwwwhindawicom Volume 2018

Diabetes ResearchJournal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Research and TreatmentAIDS

Hindawiwwwhindawicom Volume 2018

Gastroenterology Research and Practice

Hindawiwwwhindawicom Volume 2018

Parkinsonrsquos Disease

Evidence-Based Complementary andAlternative Medicine

Volume 2018Hindawiwwwhindawicom

Submit your manuscripts atwwwhindawicom

Stem Cells International

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

MEDIATORSINFLAMMATION

of

EndocrinologyInternational Journal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Disease Markers

Hindawiwwwhindawicom Volume 2018

BioMed Research International

OncologyJournal of

Hindawiwwwhindawicom Volume 2013

Hindawiwwwhindawicom Volume 2018

Oxidative Medicine and Cellular Longevity

Hindawiwwwhindawicom Volume 2018

PPAR Research

Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom

The Scientific World Journal

Volume 2018

Immunology ResearchHindawiwwwhindawicom Volume 2018

Journal of

ObesityJournal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Computational and Mathematical Methods in Medicine

Hindawiwwwhindawicom Volume 2018

Behavioural Neurology

OphthalmologyJournal of

Hindawiwwwhindawicom Volume 2018

Diabetes ResearchJournal of

Hindawiwwwhindawicom Volume 2018

Hindawiwwwhindawicom Volume 2018

Research and TreatmentAIDS

Hindawiwwwhindawicom Volume 2018

Gastroenterology Research and Practice

Hindawiwwwhindawicom Volume 2018

Parkinsonrsquos Disease

Evidence-Based Complementary andAlternative Medicine

Volume 2018Hindawiwwwhindawicom

Submit your manuscripts atwwwhindawicom