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Staphylococcus Aureus DIFFERENTIATIONDDIFFERENTIATIONIFFERENTIATION OFOOOFFF CHILEANCHILEAN NMECNEMCNEMC EID EIDEIDENDEMICEEENDEMICYSYSYS HONE HONEHONE NEMCHONEYSHHHONEYS NMC NMCBSDAEBSDAEBSDAE EIDEID YSHONEBASEDBBBASED YSONE YSONEONONON BSDAEONOOON SDAESDAE ONNN BOTANICALBBBOTANICALOTANICALOTANICAL ORIGINORIGIN ANDAND BIOLOGICALBIOLOGICAL PROPERTIESPROPERTIES.PPPROPERTIESROPERTIESROPERTIES.... MEDITERRANEANMMEDITERRANEANEDITERRANEAN ZONEZONE OFOF CHILECHILE ,, ONEONE OFOF THETHE WORLDWORLD ‘‘‘S‘‘SS HOTSPOT,HHOTSPOT,OTSPOT, EARTHEARTH BIOLOGICALLYBIOLOGICALLY RICHESTRICHEST ,, GREATGREAT ENDEMISMENDEMISM ANDAANDND MOSTMOST ENDANGEREDENDANGERED TERRESTRIALTERRESTRIAL ECOREGIONECOREGION Diversity of the 3,429 vascular plant species, of which 1,605 (46.8%) are endemic . Apicultural season in Central Chile starts at end of winter (middle August) and ends at the beginning of autumn (April). Honey harvesting takes place from December to April. Geographical and Botanical Origin. Certification lnCahieChilean Native/EndemiclnCahie osHyen Honeys osHyen onHeyHoney onHey Potential Biological Activity nigtHesrvaHarvesting nigtHesrva e vitNaNativeP atl ePlant vitNa P atl Honey Extracts Communities Laboratory Analyses ,esviBeehBeehives, ,esviBeeh Chmecial ChemicalBiiologcal Chmecial Biological BiiologcalPharmacological GSPGPS GSP gnTsetiTesting gnTseti rPoflieProfilesProprteie rPoflie Properties sProprteieIn situ validation •Obtaining New Natural WHAT WE DO Produtcs Products Produtcswith added value and CHILEAN IDENTITY Laboratorylsaysen aaanalysesanalyses: nItrvoi lsaysen: In vitrod Ivna andoniV nItrvoi dInssAsya na Vivo IvAssays oniV sAssya Certification of Botanical Origin and free of residues ilBooigcal Biological iBlooigcal Pharmacolycal Chemical Profile PrrtpeioseProperties Prrtpeiose TsetignTesting Tsetign ------CChromatographichromatographic ---Antibacterial-Antibacterial nassyilaanalysis nassyila ---Antifungical-Antifungical ---Searching-Searching of Chemical Antitumoral Activity n atdoxinitA---Antioxidant-Antioxidant n atodxnitAMrkaersMarkers Mrkaers In Situ Validation ---Antirradical-Antirradical oycuItni---Inocuity-Inocuity ocyuInti Obtaining the Honey Extracts with Biological Activities Determination of Stability and Cytotoxicity of Honey Extracts SAMPLING 665 Honey samples have been collected in 80 Beehives, from 2001 to 2008 Seasons (7 seasons) in Mediterranean arid, semiarid and humid areas. DDesertesert Scrub Shrub Vegetation Temperate Forest Deciduos Forest . 13 UNIFLORAL NATIVE HONEYS ARE PRODUCED IN CHILE Common Geographical Distribution SL Vegetation Type Native Species namenamename Tessaria absinthioides Brea 20º 30' ---37º 29 Andean and Desert Vegetation Schinus latifolius Molle 30º 30' ---35º 40' Coast and Andesn Matorral, Central Valley Cissus striata Voqui 30º 30' -----43º 30' Coast & Andean Matorral and Forest Aristotelia chilensis Maqui 30º 40' ----46º 30' Coast & Andean Matorral and Forest Quillaja saponaria Quillay 31º 10' -----8º 30 Coastal and Andean Matorral Escallonia pulverulenta Corontillo 31º 40' ----39º 30' Coastal and Andean Matorral 31º 50' ---35º 50' Coastal and Andean Matorral, Central Trevoa quinquenervia Tralhuén Valley Escallonia rubra Siete camisas 31º 50' ----35º 32 Coastal and Andean Matorral Retanilla trinervia Tevo 32º 40' ----35º 30' Coastal and Andean Matorral Weinmannia trichosperma Tineo 35º 50' -----51º 50' Coastal and Andean Temperate Forest Gevuina avellana Avellano 35º 45 ---44º 10' Coastal and Andean Temperate Forest Eucryphia cordifolia Ulmo 36º 50' -----3º 20' Coastal and Andean Temperate Forest Caldcluvia paniculata Tiaca 36º 50-----47º 30' Coastal and Andean Temperate Forest AREAAREA OF DISTRIBUTION (((EscalloniaEscallonia pulverulentapulverulenta))) SAXIFRAGACEAESAXIFRAGACEAE CORONTILLO AREAAREA OF DISTRIBUTION (((GevuinaGevuina avellana))) PROTEACEAEPROTEACEAE AVELLANO AREAAREA OF DISTRIBUTION (((EucryphiaEucryphia cordifolia))) EUCRYPHIEUCRYPHI ACEAE ULMO Bactericidal Properties of Honeys Inhibition Halo Zone of Bacterial Growth Experimental Procedure : Antibacterial activity of honeys and eextractsxtracts was performed by microdilution of trypticase/soy broth technique (37°x 24 h) Statistical analysis ANOVA a=0,05. Average values of in vitro growthgrowthgrowthInhibition halo zone for Human Pathogenic Bacteria (cm222))) Letters show the CChileanChileandifferencehilean between HoneysHoneys several unifloral andand honeys HumanHuman for the Growth PathogensPathogens Inhibition Halo Zone, after comparison of results for the same Pathogenic Bacteria. Statistical Analisys: Tukey Test (95% p>0,05) Unifloral Honeys Escherichia Enterobacter Pseudomonas Salmonella Staphylococcus colicolicoli (Migula) aerogenesaerogenesaerogenes aeruginosaaeruginosaaeruginosa typhityphityphi STH aureusaureusaureus subsp. ATCC 25922 Hormaeche Schroeter ATCC 2370 aureusaureusaureus Rosenbach ATCC ATCC 13048 27853 25923 Ulmo (Eucryphia 19,13 a 22,45 a 17,98 a 18,26 a 8,13 a cordifolia) Quillay (Quillaja 15,79 ab 17,66 b 13,15 bc 16,01 b 10,82 b saponaria) Corontilla (Escallonia 17,31 ab 13,54 ab 4,83 c No No pulverulenta) inhibition inhibition Yuyo ( Brassica rapa) 6,12 cd s. inhibición 3,93 c No No inhibition inhibition Alfalfa chilota (Lotus 6,47 c 6,78 b 21,73 b No No uliginosus) inhibition inhibition Maqui (Aristotelia 7,98 cd 13,62 ab 24,67 b No No chilensis) inhibition inhibition Avellano (Gevuina 13,11 bc s. inhibición s. inhibición 15,90 b 10,82 b avellana) IINHIBITIONIINHIBITION OFOF ERWINIAERWINIA CAROTOVORACAROTOVORA GROWTHGROWTH Honey Mean area of inhibition (cm2) Bacteria Culture Control 0,00a X – 042003-M182 25,81b ± 3,75 b X- 032004-M335 Ulmo 22,21b ± 3,49 b X- 122002- M085 Honey 21,61b ± 2,52 b X-42002-M084 20,41b ± 3,06 b X-2004-M296 16,20bc ± 3,25 c V- 2004- M294 13,80c ± 3,75 c V- 112001-M007 13,20c ± 2,28 c RM- 1120002M121 9,00c ± 7,20 d V- 092002-M017 5,40c ± 6,23d Statistic Analisis : 1. Andeva ( α=0.05); F 2,57> 2,510 probability = 0,04 2. Tukey Test (α= 0,05) Honeys of Different Origin but of the same Beekeeper showed different biological activity (Alberto Poch) Multiple Range Tests ------------------------------------------------------------------------------- Method: 95,0 percent Tukey HSD Count Mean Homogeneous Groups ------------------------------------------------------------------------------- Honey 338 10 3,0 X Quillay False Honey 10 3,18 X Artificial Honey 343 10 3,36 X Polifloral Honey 340 10 4,59 X Polifloral Miel 342 10 15,3 X Ulmo ------------------------------------------------------------------------------- Contrast Difference +/- Limits ------------------------------------------------------------------------------- Miel 338 - Miel 340 -1,59 1,83602 Miel 338 - Miel 342 *-12,3 1,83602 Miel 338 - Miel 343 -0,36 1,83602 Miel 338 - Miel Falsa -0,18 1,83602 Miel 340 - Miel 342 *-10,71 1,83602 Miel 340 - Miel 343 1,23 1,83602 Miel 340 - Miel Falsa 1,41 1,83602 Miel 342 - Miel 343 *11,94 1,83602 Miel 342 - Miel Falsa *12,12 1,83602 Miel 343 - Miel Falsa 0,18 1,83602 ------------------------------------------------------------------------------- Certificación de Origen Botánico Muestra Miel X-2005-M342 Apicultor: Alberto Poch Beekeeper Eucryphia cordifolia Mr. Alberto Poch Lotus uliginosus Blepharocalyx cruckshanksii Weinmannia trichosperma Tristerix tetrandrus Luma apiculata Trifolium repens Hypochaeris radicata Amomyrtus luma Eucalyptus globulus M elilotus indicus Rubus idaeus Plantago sp. 0,0 20,0 40,0 60,0 80,0 100,0 P o rc e n ta je Muestra Miel X-032005-M342 Rango 95% confianza Especie Nombre Común N° granos Porcentaje +/- Min Max Eucryphia cordifolia Ulmo 1120 70,088 2,245 67,843 72,333 Lotus uliginosus Alfalfa chilota 225 14,080 1,705 12,375 15,785 Blepharocalyx cruckshanksii Temu 126 7,885 1,321 6,563 9,206 Weinmannia trichosperma Tineo 105 6,571 1,215 5,356 7,786 Otras 9 especies (<1%) 1,377 0,571 0,806 1,948 M uestra M iel Río San Pedro Código X-032006-M511 E ucryphia cordifolia Lotus uliginosus Castanea sativa W einm annia trichosperm a R u b u s s p . Amomyrtus luma A extoxicon punctatum C aldcluvia paniculata H ypochaeris radicata Lum a apiculata Beekeeper Trifolium repens C issus striata Mr. Ian Staples Taraxacum officinalis Gevuina avellana Raphanus sativus 0 ,0 2 0 ,0 4 0 ,0 6 0 ,0 8 0 ,0 1 0 0 ,0 P o r c e n t a je E s p e c i e Nombre Común n ° p ó le n e s P o r c e n t a je EMV M in M a x Eucryphia cordifolia U lm o 1 0 5 3 89,389 1,759 8 7 ,6 3 0 9 1 ,1 4 8 Lotus uliginosus A lfalfa chilota, lotera 4 8 4 ,0 7 5 1 , 1 2 9 2 ,9 4 6 5 ,2 0 4 Castanea sativa C a s ta ñ o 2 0 1 ,6 9 8 0 , 7 3 8 0 ,9 6 0 2 ,4 3 6 W einmannia trichosperma T in e o 1 8 1 ,5 2 8 0 , 7 0 0 0 ,8 2 8 2 ,2 2 9 R u b u s s p . 1 3 1 ,1 0 4 0 , 5 9 7 0 ,5 0 7 1 ,7 0 0 Amomyrtus luma L u m a 6 0 ,5 0 9 0 , 4 0 7 0 ,1 0 3 0 ,9 1 6 Aextoxicon punctatum O liv illo 5 0 ,4 2 4 0 , 3 7 1 0 ,0 5 3 0 ,7 9 6 Caldcluvia paniculata T ia c a 4 0 ,3 4 0 0 , 3 3 2 0 ,0 0 7 0 ,6 7 2 Hypochaeris radicata H ierba del chancho 3 0 ,2 5 5 0 , 2 8 8 - 0 , 0 3 3 0 ,5 4 2 Trifolium repens T rébol blanco 2 0 ,1 7 0 0 , 2 3 5 - 0 , 0 6 5 0 ,4 0 5 Luma apiculata A r r a y á n 2 0 ,1 7 0 0 , 2 3 5 - 0 , 0 6 5 0 ,4 0 5 Raphanus sativus R ábano silvestre 1 0 ,0 8 5 0 , 1 6 6 - 0 , 0 8 1 0 ,2 5 1 Gevuina avellana A v e lla n o 1 0 ,0 8 5 0 , 1 6 6 - 0 , 0 8 1 0 ,2 5 1 Taraxacum officinalis D iente de león 1 0 ,0 8 5 0 , 1 6 6 - 0 , 0 8 1 0 ,2 5 1 Cissus striata V o q u i 1 0 ,0 8 5 0 , 1 6 6 - 0 , 0 8 1 0 ,2 5 1 Water Solution of Ulmo Honey 342 in different concentrations tested against de Escherichia coli 45000000 40000000 y = 2E+09x 4 - 3E+09x 3 + 2E+09x 2 - 5E+08x + 4E+07 R 2 = 0,8838 35000000 30000000 25000000 20000000 MIC = Minimal Inhibitory Concentration 15000000 10000000 UFC(Colonias Bactaerianas/ml) 5000000 0 0% 10% 15%20% 30% 40% 50% 60% 70% 80% Concentracción % (v/v) Lineal reggression between the concentration of ulmo honey versus the concentration of bacteria UFC (unit former colonies per ml) Escherichia coli.
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