NANOPARTICULAS MATÁLICAS QF-435-AULA-8 Prof. Nelson Durán-IQ-UNICAMP.

NANOPARTICULAS

MATÁLICAS

QF-435-AULA-8

Prof. Nelson Durán-IQ-UNICAMP

Chem. Eur. J. 2004, 10, 5570 – 5579

Drechsler, Erdogan, and Rotello

Elechiguerra et al. J. Nanobiotechnol. 396) 2005

METALLIC NANOPARTICLESMETALLIC NANOPARTICLES

AS CARRIERSAS CARRIERS

Gimenez et al. J. Biomed. Nanotechnol. 1, 1-7 (2005)

Lewin et al. Nat. Biotechnol. 18, 410 (2000)

HAuCl4 + NaHB4

Ho et al. Anal. Chem. 2004, 76, 7162-7168. Li et al. nnotechnology 2005, 16, 1912-1917

METALLIC NANOPARTICLES AS CARRIERS

Alves et al. Patente Braz. PIBr 0502657-1 (2005).

Lewin et al. Nat Biotechnol. 18, 410 (2000)

http://www.massmedic.com/docs/rotello.pdf

Mixed Monolayer Protected Gold Clusters (MMPCs)

http://www.massmedic.com/docs/rotello.pdf

Exemplo: Np's de Au funcionalizadas com anti-câncer para

Leucemia.• Preparação:

– Redução de Ácido tetracloroáurico(III) com Borohidreto de Sódio.

– Funcionalização após 30s da adição do agente redutor

Podsiadlo et al. Langmuir. 24, 568, 2008

6-mercaptopurine

PRATA EM PRODUTOS

Pasta de Dente

Que fazem as nanopartículas? Escova de dentes

Refrigerador

Purificador de Ar

Que fazem as nanopartículas? Sem comentários

MATERIAS PARA FERIMENTOS

Área superficial grande de partículas de prata tamanho nano melhora a

efetividade antibacteriana

-melhora os tempos de recuperação-mata bactéria mais rápido com nanoparticulas que outras formas de prata

Como preparar as nanopartícula? Nanopartículas de prata

Sal do metal + agente redutor

E.C. Constable, Depart. Chemistry, University of Basel, Switzerland

Como preparar as nanopartícula? Nanopartículas de Au e Pd

Arredondadas, lisas de 18 nm departículas de Au por reduçãode citrato

Hexagonal de 30 nm de partículasde Pd preparada por redução deuma sal de Pd com baixa concentração de ácido ascórbico

Como preparar as nanopartícula? Nanopartículas de metais

Conhecidas da antiguidade

Ouro coloidal usada para vidros tingido com vermelho e púrpura dos tempos medievais até agora

Sintese de nanoparticulas metalicas

85oCFeCl3 + 6H2O + FeCl2 . 4H2O + NH4OH --------------

HAuCl4 + NaBH4 ---------------

[Ag(NH3)2]+ Ac. ascorbico/1 h r.t. --------------

PREPARATIONChemical Synthesis of silver nanoparticles

Como preparar nanopartículas? Nanocubos de prata

Como preparar as nanopartícula? Nanopartículas de óxidos metálicos

Caracterização de nanopartículas de TiO2 por TEM

Como caracterizar as nanopartículas?

BIOSYNTHESIS OF SILVER NANOPARTICLES BY

Fusarium oxysporumAhmad et al. Colloids Surf. B. Biointerfaces 2003, 28,313-318. Mukherjee et al.ChemBioChem. 2002, 3,461-463.Pune-India

Aspergillus fumigatusBhaisa and D´Souza,Colloids Surf. B. Bio-interfaces 2006, 47,166-164Munbai-India

BIOSYNTHESIS OF SILVER NANOPARTICLES BY FUNGI

Phanerochaete chrysoporium Vigneshwaran et al. Colloids Surf. B. Biointerfaces, 2006, 53, 55-59Munbai-IndiaAspergillus flavus Vigneshwaran et al.Mat. Lett. 2007, 61, 1413-1418Munbai-IndiaPleurotus sajor-cajuVigneshwaran et al.Indian Pat. Appl. 2007:709864

Phoma spChen et al. Lett. Appl. Microbiol.2003, 37, 105-108Beijing, China

Fusarium oxysporumDurán et al. J. Nanobiotechnol.2005. 3:8, 1-7.Campinas-SP-Brazil

Durán et al. J. Biomed. Nanotechnol. 2007, 3, 203-208.Campinas, SP-Brazil

BIOSYNTHESIS OF SILVER NANOPARTICLES BY FUNGI

BIOSYNTHESIS OF SILVER BIOSYNTHESIS OF SILVER NANOPARTICLESNANOPARTICLES

BACTERIA AND YEASTBACTERIA AND YEAST

Enterobacter clocaeShahverdi et al. ProcessBiochem. 2007, 42, 919-923Teheran-Iran

Aeromonas sp.Fu et al. Chin.J. Chem. Eng. 2006,14, 114-116.Xiamen, China

Yeast strainKowshik et al.Nanotechnology 2003,14, 95-100. Berlin, Germany

BIOSYNTHESIS OF SILVER NANOPARTICLES BY BACTERIA AND YEAST

Bacterial Biosynthesis of Cadmium Sulfide Nanocrystals

Sweeney et al. Chem. Biol. 11, 1553 ()2004)

N P Fe

Sulfeto de Cadmio assistido por Lactobacillus sp.

Jha et al. NANO: Brief Reports and ReviewsVol. 2, No. 4 (2007) 239–242

Biogenic formation of photoactive arsenic-sulfidenanotubes by Shewanella sp. strain HN-41.Lee et al., PNAS, 104, 20410–20415 (2007)

BIOSYNTHESIS OF SILVER BIOSYNTHESIS OF SILVER NANOPARTICLESNANOPARTICLES

BY PLANT EXTRACTSBY PLANT EXTRACTS

Geranium (Pelargonium graveolens)Shankar et al. Biotechnol Prog.2003, 19, 1627-1631Pune, India

Neem leaf brothShankar et al.J. Colloid Interf. Sci. 2004, 275,496-502. Pune,India

BIOSYNTHESIS OF SILVER NANOPARTICLES BY PLANT EXTRACTS

Cinnamomum camphora leafHuang et al. Nanotechnology, 2007, 18, 1-11Xiamen, China

Alfalfa grassGardea-Torresdey, Langmuir 2003, 19, 1357-1361Texas, USA/Mexico, Mexico

Aloe vera plant extract Chandran et al., Biotechnol Prog. 2006, 22, 577-583.Pune, IndiaEmblica Officinalis Ankamwar et al., J. Nanosc. Nanotechnol. 5, 1665-1671. Pune, India

BIOSYNTHESIS OF SILVER NANOPARTICLES BY PLANT EXTRACTS

MECHANSTIC ASPECTS OF MECHANSTIC ASPECTS OF BIOSYNTHESIS OF SILVER BIOSYNTHESIS OF SILVER

NANOPARTICLESNANOPARTICLES

The silver-binding peptides from Pseudomonas stutzeriAG259 cells were obtained by using a combinatorial approach to identify these peptides from a phage display library of random peptides. The interaction of peptide with the metal clusters provides a chemically reducing environment around the cluster, thereby allowing further accelerated reduction of silver ions at the interface between peptide and metal.

Naik et al. Nature Mater. 2002, 1, 169-172. Ohio, USA

MECHANSTIC ASPECTS OF BIOSYNTHESIS OF SILVER NANOPARTICLESMECHANSTIC ASPECTS OF BIOSYNTHESIS OF SILVER NANOPARTICLES

Si and Mandal, Chem Eur. J. 2007, 13, 3160-3168. Kolkata, IndiaSimilar results with tryptophan and goldSelvakannan et al. J. Colloids Interf. Sci. 2004, 269, 97-102Bhattacharjee et al. J. Nanosci. Nanotechnol. 2005, 5, 1141-1147.

Selvakannan et al. Langmuir, 2004, 20, 7825-7836.Pune, India

Specific for AuStacik et al. J. Mater Chem. 2005, 15, 749-753

Fusarium oxysporumDurán et al. J. Nanobiotechnol. 2005. 3:8, 1-7.Campinas-SP-Brazil

Durán et al. J. Biomed. Nanotechnol. 2007, 3, 203-208.Campinas, SP-Brazil

Fusarium moniliforme was negative in quinone production

SILVER NANOPARTICLES AGAINST PATHOGENS

Without any antibiotics5 ug/mL of silver nanoparticles

150 ug/mL of amoxicillin

150 ug/mL of amoxicillin + 5 ug/mL of silver nanoparticles

P. Li, J. Li, C. Wu, Q. Wu,J. Li. Synergistic antibacterial effects of β-lactam antibiotic combined with silver nanoparticles. Nanotechnology 16, 1912–1917 (2005)

SILVER NANOPARTICLES AS ANTIBIOTIC CARRIERS

E. coli 5 x 106 cfu bac., a) 5 ug mL-1 silver nanoparticles b) 0.150 ug mL-1 antibiotic,c) 0.150 ug mL-1 antibiotic + a), d) b) + 10 ug mL-1 silver nanoparticles

AMOXICILLIN MECHANISM

Li et al., Nanotechnology 16, 1912 (2005)

CLINDAMYCIN

CLINCLIN

CLINChemical and fungal synthesis

Brocchi et al. J. Nanosci. Nanotechnol., submitted (2008)

MINIMUM INHIBITION CONCENTRATION (MIC)

Durán et al., Crit. Rev. Microbiol. Submitted (2008)

VANCOMICINA EM NANOPARTICULAS PARA AUMENTARSUA ATIVIDADE ANTIMICROBIANA

Gu et al. Nano Lett. 3, 1261-1263 (2003)

SINTESE DE NANOPARTICULAS DE OURO COMVANCOMICINA

SILVER NANOPARTICLESSILVER NANOPARTICLES

AND NEGLECTED DISEASESAND NEGLECTED DISEASES

Cutaneous leishmaniasis

SILVER NANOPARTICLES AND NEGLECTED DISEASES

CLINCLIN

Clindamycin associated to silver nanoparticles could be effective in these neglected diseases

SILVER NANOPARTICLES AND ITS TOXICITY

Mitochondrial function through MTS

Membrane leakage though lactate dihydrogenase ( LDH)

Ag nanoparticles 15 nm

Silver nanoparticles (15 nm) reduced mitochondrial function drastically and increased membrane leakage. Our data show that the C18-4 cells are more sensitive than the BRL 3A cells in that respect (Braydich-Stolle et al. ToxSci Advance Access published July 13, 2005).

C18-4 spermatogonial stem cells

Pal et al., 73, 712 (2007)

SKIN PERMEATION IN VITRO

Human abdominal skin

The nanoparticles did not reach the receptor compartment

20-50 nm

ANTIBACTERIAL EFFECT OF SILVER NANOPARTICLES

Silver nanoparticles from Aspergillus nigerGade et al., Appl. Microbiol. Biotechnol., submitted (2008) (Amravati-India/Campinas SP-Brazil)

PREPARATION

Biosynthesis of silver nanoparticles

Fusarium oxysporum was grown by 7 days

Absorptions were measured in UV-Vis

The biomass was filtrated and AgNO3 (10 mM) was added in the fungal liquid

The biomass was filtrated and resuspended in sterile water

Durán et al. 2005,2007,2008

Coleção de 20 linhagens de Fusarium

Culturas de Fusarium spp

Cultura em meio

líquido (YM)0,017/10-3 M de AgNO3

Caldo fungalBiomassa em

água destl.

12 Horas 72 Horas

Espectro UV-vis

MEV - nanopartículas

MATERIAIS E MÉTODOS

Solução coloidal de reação

Durán et al. J. Nanobiotechnol. 3, 1-7 (2005).

Para seleção da linhagem com melhor eficiência, levamos em consideração os dois métodos de biossíntese, tanto usando a biomassa quanto usando o líquido fungal

No caldo, foram introduzindo,017/10-3

M de AgNO3

Após ser separada a biomassa da água destilada. Este peso úmido foi adicionada em uma solução de 0,017/10-3

M de AgNO3 .Nanopartículas de prata.

Métodos

Souza et al. 58º SBPC, Florianopolis (2006)

Staphylococcus aureus Klebisiela pneumoniae

Algodão 100% Poliéster

Nanopartículas

Impregnação em

tecido têxtil para

atividade biocida

Teste oligodinâmica

Utilizando as nanopartículas

da melhor linhagem

Durán et al. Patente Braz. PIBr 0605681-4 (2006)

PRÉ-ENSAIO E RECUPERAÇÃO DAS NANOPARTÍCULAS EM EFLUENTES

A água de lavagem do tecido têxtil impregnado com nanopartículas de prata, foi submetida ao tratamento C. violaceum.

100mL/100mL 100mL/60mL 100mL/20mL

48 Horas sob

agitação

Aeração (0,5 L/mim)

Ausência de ar

Análise espectro UV-vis

Durán et al. J. Biomed. Nanotechnol. 3, 203-208 (2007).

RESULTADOS

•Espectro UV-vis de linhagens de Fusarium

produtoras de nanopartículas de prata.

Souza et al. 58º SBPC, Florianopolis, S.C. (2006)

Silver nanoparticlesSize: 1,6 nm (biosynthesis)

Durán et al. J. Nanobiotechnol. 2005

Silver nanoparticlesSize: 175 nm

(Chemical synthesis)

Espectro UV-VÍSMEV do extrato líquido

Impregnação das nanopartículas em a) algodão e b) polyester

Amostras centrifugadas

1) Algodão 100% e 2) polyester 100% 3) Cultura de Staphylococus

aureus e 4) Teste com tecido algodão impregnado com

nanopartículas de prata frente à cultura bacteriana e 5) teste do

tecido polyester impregnado com nanopartículas de prata frente à

cultura bacteriana 1

ATIVIDADE BIOCIDA DO TECIDO IMPREGNADO (ALGODÃO) COM NANOPARTÍCULAS DE PRATA

frente a bactéria S. aureus

MICROSCOPIA ELETRÔNICA DE VARREDURA DO TRATAMENTO DE ALGODÃO COM NANOPARTICULAS DE PRATA

Tecido Controle (sem nanopartículas de prata)

Tecido com nanopartículas de

ATIVIDADE BIOCIDA DO TECIDO IMPREGNADO

(POLIÉSTER) COM NANOPARTÍCULAS DE PRATA frente a

bactéria S. aureus

M.O do Poliéster

impregnado com

M.O das amostras não impregnadas

•Os tecidos impregnados com nanopartículas, após sucessivas lavagens, perdem gradativamente sua atividade biocida, uma vez que as nanopartículas são liberadas durante este processo.

•Para evitar o descarte de nanopartículas no ambiente, foram realizados ensaios com diferentes concentrações de inóculo da bactéria, Chromobacterium violaceum.

POSSÍVEL RECUPERAÇÃO

Espectros A e B, correspondentes às amostras controle (A, sem diluição e B, diluição de 10 vezes). (C, D e E submetidas à aeração). (F, G e H submetidas à ausência de ar) todas tratadas com Chromobacterium violaceum.

EDS: Espectro de nanopartículas de prata em bactéria Chromobacterium violaceum.

CONCLUSÃO

•Todas as 20 linhagens de Fusarium utilizadas neste trabalho

apresentaram capacidade de produção de nanopartículas de prata,

entretanto, foram as três mais eficientes separadas (dados

apresentados no Relatório Parcial). Destas, foi selecionada a

linhagem 07SD.

•Avaliaram-se dois métodos de biossíntese: um empregando

diretamente a biomassa do fungo e outro apenas o líquido fungal.

Este último mostrou melhores resultados. Logo, deu-se

continuidade ao trabalho empregando este método.

•Os tecidos impregnados com as nanopartículas de prata

apresentaram atividade biocida frente a Staphylococcus aureus e

Klebsiella pneunomiae.

•A possibilidade de recuperação das nanopartículas de prata após

lavagens sucessivas dos tecidos utilizando Chromobacterium

violaceum foi comprovada, podendo ser empregada no

reaproveitamento contínuo dessas nanopartículas (ainda em teste)

Jun Tian, Kenneth K. Y. Wong, Chi-Ming Ho, Chun-Nam Lok,Wing-Yiu Yu,] Chi-Ming Che, Jen-Fu Chiu, and Paul K. H. Tam. Topical delivery of silver nanoparticles promotes wound healing.ChemMedChem.(2007)2,129-136.

• Preparation of the dressings:• Nanosilver-coated dressing: Initial experiments were carried

out by using silver nanoparticle grafted dressing (silver content: 2.75 mgg1, Anson Nanotechnology Group Co., Ltd. Hong Kong). A piece of silver nanoparticle-coated dressing (4H3 cm2) weighed 0.1737 g. This corresponds to 0.4777 mg of silver nanoparticles on each piece. Transmission electron microscopy revealed that silver nanoparticles were spherical, with diameters of 149.8 nm based on 750 particle measurements on five sites of interest. Subsequently experiments were performed with silver nanoparticles (1 mm) synthesized by borohydride reduction of AgNO3 in the presence of citrate as a stabilizing agent, as previously described.[45] Similar efficacy with silver nanoparticle dressing was confirmed (data not shown).

SSDNTND

Silver Nano Silver sulfadiazine

E:epidermidHF: hair follicles

CULTURA DE MICRORGANISMOS NO PANO DA FERIDA NOS DIAS 1, 3, 5, 7 E 10 NOS ANIMAIS TRATADOS COM ND (GRIS), SSD (BRANCO E NÃO TRATADO (GRIS ESCURO)

silver nanoparticles can modulate local and systemic inflammatory response following burn injury by cytokine modulation.

ND▄ No treatment▲ SSD

IL-6 TGF-beta IL-10

VGEF IFN-gama

IL-6 aumenta inflamaçãoTGF-beta auenta inflamaçãoIl-10regula citokinasVGEF (vascular endotelial growth factor) promove cicatrizaçãoIFN-gama aumenta colageno

PATENTES EM COSMÉTICOS • Facial mask comprising a nanometer metal coating. Ko, Chuan Tao. (Taiwan). U.S. Pat. Appl. Publ.

(2007), 5pp. CODEN: USXXCO US 2007125383 • Silver-containing an antimicrobial body care product. Hanke, Bernhard. (Procter and Gamble Company,

USA). PCT Int. Appl. (2000), 22 pp. CODEN: PIXXD2 WO 2000078281 • Antimicrobial gel formulation containing silver nanoparticles for treating female genital infections, its

preparation method and uses. Chen, Hu. (Shenzhen Agt Pharm. Co., Ltd., Peop. Rep. China). Faming Zhuanli Shenqing Gongkai Shuomingshu (2005), 16pp. CODEN: CNXXEV CN 1672689 A

• Manufacture of cosmetics containing silver nanoparticles and Carbopol. Liu, Baning; Ma, Xiaoling; Liu, Xi. (Peop. Rep. China). Faming Zhuanli Shenqing Gongkai Shuomingshu (2004), No pp. given. CODEN: CNXXEV CN 1539403 A

• Formulation of UV absorbers by incorporation in solid lipid nanoparticles. Herzog, Bernd. (Ciba Specialty Chemicals Corporation, USA). U.S. Pat. Appl. Publ. (2003), 16 pp. CODEN: USXXCO US 2003235540 A1

• Pharmaceutical compositions for treating allergic dermatitis comprising nano-gold solution. Han, Sang Pil; Yoo, Eui Jae; Choi, Paeck Hi; Choi, Young Deuk. (Hanssem Health Co., Ltd., S. Korea). Repub. Korean Kongkae Taeho Kongbo (2006), No pp. given. CODEN: KRXXA7 KR 2006134261

• Hair preparations containing silver nanoparticle colloids for the treatment of bacteria-caused hair depilation. Han, Sang Pil; Yoo, Eui Jae; Choi, Paeck Hi; Choi, Young Deuk. (Hanssem Health Co., Ltd., S. Korea). Repub. Korean Kongkae Taeho Kongbo (2006), No pp. given. CODEN: KRXXA7 KR 2006110600

• Vaginal adhesive preparation containing silver nanoparticles and its preparation method. Gu, Maojian; Ren, Fuzheng; Liang, Wei; Jiang, Zongrun. (Shanghai Jnj Modern Pharmaceutical Technologies Inc., Peop. Rep. China). Faming Zhuanli Shenqing Gongkai Shuomingshu (2006), 14pp. CODEN: CNXXEV CN 1857310

• Anti-microbial condom containing silver nanoparticles, capable of preventing various adult diseases and pregnancy. Yang, Won Dong. (S. Korea). Repub. Korean Kongkae Taeho Kongbo (2005), No pp. given. CODEN: KRXXA7 KR 2005114182

• Antiaging cosmetics containing gold nanoparticles and silk fibroin for maintaining antioxidant activity. Hur, Won; Lim, Kun Bin; Ahn, Hyo Soon; Shin, Hwa Sook. (Fineco. Ltd., S. Korea). Repub. Korean Kongkae Taeho Kongbo (2006), No pp. given. CODEN: KRXXA7 KR 2006108789

SINTESE DE NANOPARTICULASDE OURO ASSOCIADA A IgG

IgG de soro humano

HAuCl4 + NaBH4

BACTERIA

SEPARAÇÃO

ANALISE

MLDI Analysis: (H. Steen and M. Mann. “The abc’s (and xyz’s) of peptide sequencing”. Nat. Rev. Mol. Cell Biol. 5, 699-711 (2003)).

Staphylococcus saprophyticus were collected from patients at the Hospital, Streptococcus pyogenes JRS 75 and JRS 4 were from collections S. pyogenes JRS 75 was obtained by mutating M protein from the strain of S. pyogenes JRS 4.29The lowest cell concentration that was detected for both Staphylococcus saprophyticus and Staphylococcus aureus in aqueous sample solutions (0.5 mL) was 3 x 10(5) cfu/mL, while the detectable cell concentration for S. saprophyticus in a urine sample was 3 x 10(7) cfu/mL

Absorption and desorption of chemotherapeutic drugsfrom a magnectically targeted carrier (MTC). Rudger et al. J. Control. Releae 74, 335-340 (2001).

Doxorubicin follows the Langmuir theory formilling, particles are resuspended and dispersed, adsorption to MTCs. The Langmuir theoryholds that the solute adsorbs to sites on the carbonsurface in a discrete one to one correspondence.

ACTIVATED CARBON

MAGNETICALLY TARGETED CARRIER (MTC)

Rudger et al. J. Control. Releae 74, 335-340 (2001)

ALVOS NAO ESPECIFICOS------------------------------------------------------------------------------------------------------

Treatment of a hepatocellular carcinoma byMTCs in clinical human trials

As shown in the left panel following the treatment of a HCC tumor with MCT-DOX the hepatic arteriesRemain patent, demonstrating particles toletaribility as evidenced by the lack of trombosis and/orembolization of the arteries. Localization and retention of the particles in the target tumor are showed in the the righ papel by magnetic resonant imagins (MIR) (the particles were in the local after 28 days)