F O L I A V E T E R I N A R I A5, 2 , 3 - 4 : 1 3 5 - 1 3 92, 0 0 8 www.uvm.sk BIOLOGICALEFFECTSOF ROSEMARY(rRosmarinas officinalis1,./ BSSENTIALOIL (A Review) Faixov6,Z.r, Faix, S.2 rUniversity of Veterinary Medicine, Komensk6ho73, 041 g1 Ko5ice 2lnstitute of Animal Physiology Slovak Academyof Sciences,Solt6sovej4, 040 0l KoSice The Slovak Republic [email protected] ABSTRACT Many herbs and plant extractsare addedto the diet not only for their aromaticpropertiesbut they havebeenidentified as a sourceof variousphytochemicals, manyof which possess an importantbiologicalactivity.Resultsof many experiments showed that rosemary essential oil hadantimicrobial, antioxidant, anti-carcinogenic, cognition-improving and certainglucoselevel loweringpropertieswhich makesit usefulas a naturalanimal feedadditive.This reviewdescribes themostimportantbiological activitiesof rosemary(Rosmarinus olficinatisL.) essentialoil in animalsand humans.In vitro and,in vivoeffectsof rosemarv essentialoil are discussed. Key words:essentialoil; metabolism;rosemary INTRODUCTION The useof antibioticsas growthpromotersin animalfeed is facingreducedsocial acceptance due to the appearance of residuesand resistantstrainsof bacteria;antibioticuse hasbeenbannedin the EuropeanUnion sinceJanuary2006 (Regulationl83l /2003/EC). Many herbsand plant extractshaveantimicrobialactivities and antioxidantpropertieswhich makethem usefulas natural animalfeedadditives.Si et al. (28) reportedthan cinnamon has antimicrobialactivity againstSalmonellaserotypeTyphymuriumDT 104of swinein vitro study.Thesefindingsare in agreemew n ti t h t h e r e p o r t so f P r a b u s e e n i v a sa n e t a l . (25). They reportedthat cinnamonclove,geranium,lemon, orangeand rosemaryoils exhibitedsignificantantibacterial propertiesagainstboth gram-negative and gram-positive bacteria in vitro. Antioxidanteffectof aromaticplantsis dueto the presence of hydroxylgroupsin their phenoliccompounds (30). Most essential oils are classified as GenerallyRecognized as Safe(GRAS)and havebeenapprovedfor food and beverage consumption by US Food and Drug administration (31). Plantextracts(aniseoil, cinnamonoil, andgarlicoil) may allowone to manipulatewith rumenmicrobialfermentationby decreasing total volatilefatty acid concentrationand reducing ammoniaN concentration (6). Essentialoils derivedfrom sage,rosemary, thymeand other herbswere shownto inhibit osteoclastactivity and increase bone densityin vitro (26). Recentlyit was reportedthat some essentialoils, such as cinnamon,havethe potentialto influencefavourablythe insulinsystemand aflfectbeneficially bloodglucoseand lipid metabolism in peoplewith type 2 diabetes(17). The greatestattention from among all herbs and spices actingas biologically activecompounds has beenfocusedon rosemary. The objectivesof the current review were, therefore.to describebiologicalactivitiesof rosemary(Rosmarinus officinalis l.) essential oil in animalsand humans. Essentialoil composition The essential oil of rosemarycontainsseveralcompounds at a rather differentconcentrations. It is characterized by two or three major componentsat fairly high concentrations (20 to 70%) compared presentin traces. to othercompounds 135 Generally,these major componentsdetermine the biological propertiesof the essentialoil and can act in synergicmanner oils might be more effective against food-borne pathogens%nd spoilagebacteriawhen applied to readyto use food containing a high protein level at acidic pH, as well as lower levels of or regulate one another. D e b e r s a c e t a l . ( 8 ) r e p o r t e dt h a t t h e m a j o rc o m p o n e n ot f fats or carbohydrates. dried leavesof Rosmarinusofficinalis (L.), supplied by Robertet ( G r a s s eF , rance)w , a sm o n o t e r p e n e o x i d e 1 , 8c i n e o l e( 3 6 . 1 % ) . Antioxidant properties Many monoterpeneand sesquiterpene hydrocarbonswere present in largeamounts(32.2%). The authorsidentified c_pinene, camphene,limonene, myrcene and B-pinene,B-caryophylene, p-cymene.They alsodetectedthe monoterpeneketonecamphor ( 1 2 . 8 % ) a n d m o n o t e r p e n ea l c o h o l s( 9 . 6 % ) , s u c h a s b o r n e o l . Afl these compoundsamounted to 93.4% (wlw) of essential oil. All remaining substanceswere minor volatile compounds and six of them were still unidentified. In the dichlormethane extract (DCME) from rosemary leavesonly 28.1% of the compoundspresentwere quantified. The main compoundscharacterizedin DCME were flavonoids and phenolic diterpenes.Two flavones were identified as cirs! maritin and genkwantin.Six structuresof phenolic diterpenes were detectedin that extract, namely carnosic acid (14.j %) and its methylatedform (7.4%), carnosol(3.8 %), rosmanol(1.4%), epirosmanol methyl ether (0.8 %) and epirosmanol (traces). Dichlormethane extracted also other lipophilic chemicals, probably correspondingto pigments,sterolsand hydroxylated fatty acids (8). Water-solubleextract from rosemary leaves was rich in rosemarinicacid ( 1.3%) and flavonoids(3%). A wide rangeof oxygenatedmonoterpeneswere identified: oxides,such as l,g cin_ eole, and alcohols,such as borneol and c-terpineol. Chemical composition of rosemary essentialoil can vary betweenregionsand it dependsmostly on climate, soil com_ position, plant organ, age and stage of vegetativecycle. Cytotoxicity of rosemary essential oils Oxygenatedmonoterpenesof essentialoil seem to exhibit a variabledegreeofcytotoxicity. As typical lipophilic substances, they passthrough the cell wall and cytoplasmicmembraneand disrupt their structure.In bacteria,the membranedamageis relatedto the loss of ions and reduction in membranepotential, collapseof the proton pump and ATp depletion. This cytotoxic property is used to treat individuals affected by some human or animal pathogensor parasites(20). B o z i n e t a l . ( 5 ) t e s t e d a n t i m i c r o b i a la n d a n t i o x i d a n t activities of the essential oils of rosemary (Rosmarinus ffici_ nalis L.) and sage (Salvia officinatis L.). Their antimicrobial activity was tested against 13 bacterial strains and 6 fungi, including Candica albicans and 5 dermatocytes. The highest antibacterial activity of both essential oils was expressedon E. coli, Salmonella typhi, S. enreritidis and Shigetta sonei. Essential oil of rosemary exhibited a significant rate of antifungal activity. Fu etal. (10) reported that essentialoils from clove and rosemaryalone and in combination exerteda significant antimicrobial effect against Staphylococcus epidellnidis,Escherichia coli and Candica albicans. The cytotoxic effect of rosemary is of great importance in preservationof agricultural or marine products. The antimicrobial efficacy of plant essentialoils depends on food c o m p o s i t i o nG . utie rrez et al. (14) found out that essential r36 Free radicals/reactiveoxygen speciesare associatedwith many biologicalphenomena,such as infllammation,aging,and carcinogenesis.The antioxidant activity of polar extracts of rosemaryis relatedto the content of phenolic compounds(i. e. carnosol,carnosicacid). Constituentsin rosemaryhave shown a varietyof pharmacologicalactivitiesfor cancerchemopreven_ tion and therapy in in vitro and,in vivo models (29). Cigarettesmoke, containing reactiveoxygen specles,may activate the second step of benzo(a) pyrene metabolic way and thus may be partially responsiblefor the formation of t h e c r i t i c a ll u n g t u m o r i g e n i ca d d u c t .A l e x a n d r o v e t a t . ( l ) reported that cigarettefilter containing rosemary extract caused higher than 70% decreasein benzo(a) pyrene adducts levels arising due to cigarettesmoke in MCF cells. This approach may reduce the lung cancer risk in addicted smokers. Fahim et al. (9) found that rosemary ethanolic extract (0.15g/100g b.w.) given to rats for 3 weeks showed hepatoprotective and antimutagenic effects attributed to the presence of a relatively high percentageof phenolic compounds with high antioxidantactivity. C h e u n g e , T a i ( 1 5 ) s t u d i e da n t i - p r o l i f e r a t i vper o p e r t i e s of crude extracts of rosemary (Rosmarinus fficinatis L.) in several human cancer cell lines and their anti-oxidative properties lr? vitro in a mouse RAW 264.7 macrophage/monocyte cell line. The study showedthat the crude ethanolic rosemary extract had anti-proliferative effect on human leukemia and breast carcinomacells. The body possessesvarious antioxidative systems (free radical scavengingactivity, FRSA) that prevent oxidative stress, for example saliva exhibits such an activity. A t s u m i e t T o n o s a k i ( 2 ) m e a s u r e dt h e t o t a l s a l i v a r y FRSA induced after sniffing lavender and rosemary essential oils which are widely used in aromatherapy. Salivary FRSA valueswere increasedby stimulation with low concentrations (l 000 times dilution) of lavender or by high concentrations ( l0 times dilution) of rosemary.In contrast,both lavenderand rosemary stimulationsdecreasedcortisol levels.A significant inverse correlation was observed between the FRSA value and the cortisol levelswith each concentration of rosemary stimulation. Thesefindings show that Iavenderand rosemaryenhance FRSA and decreasethe stress hormone, cortisol, protecting body from oxidative stress. Plant extractsrich in polyphenols (pERp) could provide an important alternative antioxidant but little is known about their use in ruminants since the antioxidantcapacityof pERp could be altered by digestiveprocesses. G l a d i n e e t a l . ( 1 2 ) r e p o r t e dt h a t a s i n g l e a c u t e d o s e of PERP, given directly into the rumen highly susceptibleto lipoperoxidation,enhancedthe plasmatotal antioxidantstatus. The digestiveprocessesof PERP in vivo arc beneficial by improving the biological effect of polymeric proanthocyanidins. It was found that addition of rosemaryextract dnlayedthe oxidation of lipid fraction of minced meat balls during storage in the freezer. The antioxidative effect was related to the concentrationofthe ethanol rosemaryextractin the product (16). S i m i l a r r e s u l t sw e r e r e p o r t e db y L o p e z - B o t e e t a t . ( 1 9 ) . In contrast to the above, other experiments have reported There is a lack of data regarding the effects of dietary rosemaryextract on immunity. The results of B ab u er al. (3) indicate that rosemary dietary extract might not result in general enhancing of the immune system in young rats and will probably be effective little or no antioxidant activity of rosemary. G a l o b a r t e t a l . ( 1 1 ) f o u n d t h a t d i e t a r ys u p p l e m e n t a t i o n with 500 or 1 000 mg/kg of a commercial rosemary extracr under some stress condition, such as protein or antioxidant deficiency. had no effect on the lipid oxidative stability of eggs enriched with omega-3fatty acids. Cognition-enhancingproperties Effect on rumen microbial fermentation In dairy cattle, the use of antibiotics as feed additives, such as ionophore antibiotics,has proved to be a useful tool to reduce energy (in the form of methane) and nitrogen (in the form ol ammonia) lossesfrom the diet. However,the use of antibiotics as feed additives in dairy cows is banned in the EuropeanUnion. The use of plant extractsappearsas one of the most natural alternativesto the antibiotic use in animal nutrition. C a s t i l l e j o s e l a / . ( 7 ) s t u d i e dt h e e f f e c t so f 1 0 e s s e n t i a l oils (rosemary,hyssop, sage, tea tree, clove leave. lavandin. lavender, thyme, oregano and savor), administered in vitro at three doses (5, 50 and 500mg/l), on rumen fermentationof high concentrate,feedlot-typediet. The majority ofthe essential oils modified rumen microbial fermentation and may allowed one to manipulate with rumen fermentation to improve animal performance. Effect on bone Miihlb auer et al. (22) tested the effects of some common herbs (rosemary,thyme and sage) and their constituent essentialoils and monoterpeneson bone resorptionin ovariectomized rats. Bone resorption was inhibited by the addition of I g of powdered leaves of each herb, and the essential oils extractedfrom sageand rosemary had similar inhibitory effects. Recently our results showed that diet supplementation with graded proportions of rosemaryessentialoil (0; 0.1; 0.05 and 0.025%) causeda significantly lower plasma calcium level in broiler chicksand the effectof rosemaryessentialoil on plasma calcium levels tended to be dose-dependent(FaixovAetal. 2 0 0 8 , u n p u b l i s h e dd a r a ) . Antidiabetic properties Rosemary (Rosmarinus officinatis) is used in traditional Turkish folk medicine for the treatment of hyperglycaemia. B a k i r e I et al. (4) investigatedpotential effect of ethanolic extract of Rosntarinus o.fficinalisleaveson glucose homeostasis in rabbits. Results of this study showed that ethanolic extracts of leavesof Rosmarinusofficinalis (L.) reduced blood glucose level in normoglycaemic and glucose-hyperglycaemicrabbits. Repeatedadministration ofl rosemary extract to alloxan-diabetic rabbits led to a decreasein blood glucose level and significant increasein serum insulin level. However,results of our experimentwith dietary graded levelsof rosemaryessentialoil in broiler chicks did not confirm the abovementionedfindings (Faixovri et al., 2008, unpublisheddata). Effect on immunity Severalnon-toxic European herbal specieshave pan-cultural traditions as treatment for cognitive deficit, including that associatedwith ageing.Particularlypromising candidatespecies include sage,lemon and rosemary. The essentialaction of rosemaryessentialoil is in stimulation of the nervous system under sympathetic control resUlting in improved memorizing and concentrating abilities (27). Moss el al. (21) assessedthe olfactory impact of essential oils of lavender and rosemary on cognitive performance and mood of healthy volunteers. They reported that rosemary produced a significant enhancementof performancein terms of overall quality of memory and secondarymemory factors, but also impaired the speedcomponent of memory compared to the control. Essentialoils of peppermintand rosemaryhave been fiound to increasethe activity level of mice (18) and diffusion of rosemary into the environment encouragedthe dogs to spend more of their time alert (i. e. standing, moving) than any other olfactory conditions (lavender,chamomile) ( l3). Rosemaryessentialoil was found to cause moderate inhibition of acetylcholinesterase (24). The anti-acetylcholinesterasd activity of rosemary essential oil is explained by synergic interaction between 1,8 cineole and 2-pinene in rosemary essentialoil. Inhibition of acetylcholinesterase is considered one of the treatment strategies against several neurologic disorders,including Alzeimer's disease,senile dementia and myasthenia gravis. Adverseeffects of rosemary Nusier et al. (23) studied the effect of dietary rosemary (250 and 500mg/kg b.w.) on fertility of adult rats over the period of 63 days. They reported that sperm motility and density were significantly decreasedin the cauda epididymidis and in the testes of rosemary-treated male rats in group 2. In addition, the treatment markedly increasedthe number of casesof floetalresorption in female rats impregnated by group 2 males, thereby reducing their fertility. CONCLUSION Many in vivo and in yitro experiments investigated effects of rosemary essentialoil in order to show that its constituents have important biological properties. Results of the relevant studies showed that there are biologicallyactive compoundsin rosemary essential oil exhibiting cytotoxic, antioxidant, anti-carcinogenic and cognition-enhancingproperties.They also have the potential to influence glucoselevel in diabetic patients, t37 rllodify rumen microbial fermentationand enhancebone resorption. Rosemary essentialoil, however, does not enhanceimmune responseand there are reports about its adverse effects on fertility. 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