Proiect 91 din 02.08.2010

BIOCATALIZATORI IN MEDII NECONVENTIONALE. APLICATIE IN VALORIFICAREA BIO-GLICEROLULUI CA PRECURSOR IN DOMENIUL BIORAFINARIE

Director proiect: Lector dr. Madalina Valentina Sandulescu-Tudorache

Abstract

Proiectul propune realizarea si caracterizarea unor noi tipuri de catalizatori enzimatici pe baza de lipaze aplicati in domeniul valorificarii bio-glicerolului ca produs secundar cu pondere majora rezultat in urma fabricarii biodieselului. Proiectul are un caracter dual. In primul rand, se propune un studiu teoretic asupra comportamentului catalitic al biocatalizatorilor construiti prin imobilizarea enzimei lipaza pe suprafata micro/nano-particulelor magnetice in medii neconventionale de tip lichide ionice si apa saraca in deuteriu. Din punct de vedere practic, proiectul propune o noua metoda de sinteza biocatalitica a carbonatului de glicerol prin transesterificarea biocatalitica a bio-glicerolului in prezenta catalizatorilor de tip lipaza studiati. Astfel, se lanseaza ideea unei noi metode de valorificare a bio-glicerolului avand rol de precursor al unui produs de biorafinarie, carbonatul de glicerol.

Biocatalizatorii rezultati vor fi caracterizati cu tehnici precum SPR si detectie Electrochimica. Se va realiza un studiu laborios al comportamentului catalitic al biocatalizatorilor (activitate catalitica, chemo-, regio- si enantioselectivitate) in medii neconventionale (ex. lichide ionice si apa saraca in deuteriu) in scopul elucidarii mecanismului catalitic in conditiile mentionate.

Aplicarea biocatalizatorilor-lipaza in sinteza biocatalitica a carbonatului de glicerol implica generarea unor sisteme experimentale biocatalitice originale atat prin design-ul bioreactorului (structuri de tip lipaza-particula magnetica actionate de un camp magnetic alternativ) cat si prin principiul de functionare al sistemului bazat pe recircularea unui flux continuu a amestecului de reactie (glicerol si dimetil carbonat) in interiorul sistemului. Experimentele propuse vor permite optimizarea sistemelor biocatalitice si in final valorificarea lor in obtinerea eficienta a carbonatului de glicerol.

 

THE PROJECT PROPOSES THE DEVELOPMENT AND CHARACTERIZATION OF NEW TIP OF ENZYME-CATALYST WITH LIPASE. THE CATALYST WILL BE APPLIED IN THE FIELD OF BIO-GLYCEROL CHEMISTRY, ESPECIALLY FOR SYNTHESIS OF IMPORTANT BIORAFINARY PRODUCTS. THE PROJECT WILL BE DEVELOPED FOLLOWING TWO IMPORTANT DIRECTIONS. ONE DIRECTION IS REPREZENTED BY A THEORETICAL STUDY OF LIPASE-CATALYST IN NON-CONVENTIONAL MEDIUM (I.E. IONIC LIQUIDS / DEUTERIUM DEPLETED WATER), WHEN THE ENZYME IS IMMOBILIZED ON THE SOLID SUPPORT (MAGNETIC NANO-/MICROPARTICLES). THE OTHER DIRECTION IS IN FACT THE IMPLEMENTATION OF THEORY BY DEVELOPING AN INTERESTING APPLICATION OF DEVELOPED BIOCATALYST FOR SYNTHESIS OF GLYCEROL CARBONATE. THIS IS A NEW ALTERNATIVE FOR INDUSTRIAL SYNTHESIS OF GLYCEROL CARBONATE BASED ON HETEROGENEOUS TRANSESTERIFICATION OF GLYCEROL IN THE PRESENCE OF LIPASE-BIOCATALYST.

THE BIOCATALYST WILL BE CHARACTERISED BAZED ON SPR AND ELECTROCHEMICAL DETECTION TECHNIQUES FOLLOWING SEVERAL ASPECTS SUCH AS CATALYTIC ACTIVITY, CHEMO-, REGIO- AND ENANTIOSELECTIVITY.

NEW EXPERIMENTAL BIOCATALYTIC SYSTEM DEDICATED TO PROPOSED SYNTHESIS WILL BE SET UP WITH AN ORIGINAL DESIGN (BIOREACTOR BASED ON LIPASE-MAGNETIC PARTICLE STRUCTURE MANIPULATED BY ALTERNATIVE MAGNETIC FIELD) AND SPECIAL WORKING PRINCIPLE (RE-CYCLING CONTINEOUS FLOW). FINALLY, THE SUCCESS OF THE PROJECT WILL GIVE A NEW ALTERNATIVE FOR GLYCEROL CARBONATE SYNTHESIS ACCORDING WITH GREEN CHEMISTRY LEGISLATION.

Stage I       2010

 

Obiective

  • Documentare stiintifica si de laborator
  • Managementul proiectului

Objectives (English)

  • Theoretical and practical up date.
  • Project management

 

Rezultate

Documentarea stiintifica realizata in cadrul acestei etape a permis constituirea unei imagini de ansamblu asupra ideii stiintifice care face obiectivul acestui proiect. Astfel, membrii echipei de cercetare s-au familiarizat si au fost informati asupra cercetarilor de laborator raportate pana in prezent in domeniul biorafinariei pentru sinteza de carbonat de glicerol din bio-glicerol. Studiul de literatura realizat a demonstrat ca subiectul propus prezinta un interes deosebit in acest moment in lumea stiintifica. Sinteza carbonatului de glicerol se realizeaz in prezent pe baza unor metode clasice care implica costuri ridicate (ex. instalatii tehnologice, etape de purificare, etc.) dar si riscul unei toxicitati avansate (ex. utilizarea unor catalizatori acido-bazici in cataliza omogena, solventi organici, etc.). Documentarea de laborator a implicat realizarea unui necesar de reactivi si substante, cat si instrumental de laborator utile experimentelor propuse pentru etapa urmatoare. Toate acestea au fost achizitionate in cadrul etapei UNICE/2010.

Results (English)

The research team performed a lite-search on the topic of the project in order to update the information and for a better understanding of what will be developed in the future research activity. The researchers became familiar with notions such as biorefinery, bio-glycerol and value-added products. Also, we demonstrated that the project topic is important for nowadays chemistry because the current methods of glycerol carbonate synthesis is characterized by several drawbacks (e.g. many steps of the synthesis, high toxicity of the reagents, etc.). This is why we propose a green synthesis of glycerol carbonate based on glycerol and dimethyl carbonate interaction assisted by lipase enzyme. The synthesis system was developed under solvent-free conditions.

 

Diseminare (Dissemination)

Conferinte (Conferences)

  • BioTrends 2010, Dortmund, Germania - poster.
  • RENEXPO 2010, Bucuresti, Romania - poster.
  • Conferinta Diaspora 2010, Bucuresti, Romania - poster.

Publicatii (Publications)

  • M. Tudorache et al., Biocatalytic microreactor incorporating HRP anchored on micro-/nano- lithographic patterns for flow oxidation of phenols, Journal of Molecular Catalysis B: Enzymatic, accepted with revisions, december 2010.
  • L. Protesescu; M. Tudorache; S. Neatu; N. Grecu; E. Kemnitz; P. Filip; V. Parvulescu; S. Coman, Unusual Behaviour of a Novel Heterogeneous Chiral Dimer CrIII-Salen Complex in the Epoxidation/Epoxide Ring-Opening Reaction of trans-Methylcinnamate Ester, The Journal of Physical Chemistry, accepted with revisions, december 2010.

Stage II      2011

 

Obiective

  • Investigarea activitatii catalitice a lipazelor
  • Prepararea si caracterizarea biocatalizatorilor pe baza de lipaza
  • Managementul proiectului

Objectives (English)

  • Investigations of the lipase catalytic activity
  • Preparation and characterization of the biocatalyst based on the lipase enzyme
  • Project management

 

Rezultate

Studiul stiintific, atat la nivel teoretic cat si la nivel experimental realizat in cadrul acestei faze au permis construirea unui biocatalizator enzimatic de tip enzima-particula magnetica care sa sustina desfasurarea unui proces de sinteza a carbonatului de glicerol cu performante bune. In cadrul acestei faze, cercetatorii in formare si-au insusit cunostinte temeinice referitoare la domeniul Biochimie Tehnologica cum ar fi notiuni legate de activitatea enzimatica, metode de imobilizare a enzimei pe un suport solid, factori experimentali care influenteaza activitatea enzimatica in urma imobilizarii, etc. Produsul final al acestei faze il constitue biocatalizatorul lipaza-particula magnetica, enzima provenita din sursa Aspergillus niger.

Results (English)

In this work-package the enzyme biocatalyst has been developed. The biocatalyst design was the enzyme immobilized on the particle surface. The biocatalyst was dedicated to glycerol carbonate synthesis. On this stage, the young researchers came in contact with new and useful knowledge from Technological Biochemistry (e.g. enzyme activity, approach of the enzyme immobilization on the slid support and experimental parameters affecting the enzyme catalytic capacity after immobilization). The end-product of this stage was the biocatalyst based on lipase.

 

Diseminare (Dissemination)

Conferinte (Conferences)

  • FEZA, Iulie 2011, Valencia, Spania – poster.
  • BIOMASS, Iulie 2011, Berlin, Germania – poster.
  • EUCOMC, Iulie 2011, Toulouse, Franta – poster.
  • BioTrans, Octombrie 2011, Giardini Naxos, Italia – poster (s-a castigat premiul pt cel mai bun poster la sesiunea New biocatalitic applications).
  • ICEST, Noiembrie 2011, Kusadasi, Turcia – oral.

Publicatii (Publications)

  • M. Tudorache, L. Protesescu, S. Coman, V.I. Parvulescu, Efficient bio-conversion of glycerol to glycerol carbonate catalyzed by lipase extracted from Aspergillus niger, acceptat spre a fi publicat in Green Chemistry (5,7 factor de impact), Noiembrie 2011.
  • M.Tudorache, L. Protesescu, A. Negoi, V.I. Parvulescu, Magnetic beads bioreactor based on Aspergillus niger lipase for green synthesis of glycerol carbonate, trimis spre publicare in Journal of Molecular Catalysis B: Enzymatic, Decembrie 2011.

Stage III     2012

 

Obiective

  • Construirea bioreactorului pe baza biocatalizatorului lipaza-pm
  • Realizarea sistemului biocatalitic
  • Managementul proiectului

Objectives (English)

  • Construction of the bioreactor with lipase immobilized on the magnetic particles as the biocatalyst design
  • Biocatalytic system
  • Project management

 

Rezultate

In urma activitatii stiintifice desfasurate in cadrul acestei etapa, membrii echipei de cercetare au reusit sa construiasca si sa optimizeze un sistem biocatalitic dedicat sintezei glicerol carbonatului. De asemenea, metoda biocatalitica de sinteza a fost optimizata pentru a reusi sa producem glicerol carbonat intr-un mod eficient. Produsul final al acestei etape il constitue sistemul biocatalitic de sinteza a glicerol carbonatului. Testarea biocatalizatorilor de tip „enzima immobilizata” cu design-uri diferite a permis alegerea catalizatorului optim pentru metoda de sinteza propusa. Rezultatele obtinute vor fi utilizate in etapa urmatoare pentru a testa biocatalizatorii studiati pana in prezent in medii neconventionale.

Results (English)

A biocatalytic system for glycerol carbonate production has been developed and optimized on this stage. The optimization process allowed to set up new and performing synthesis system for the production of glycerol carbonate with high efficiency. The system was tested for all of the biocatalysts developed in the previous stage. Also, the best biocatalyst was chosen for performing biocatalysis of glycerol to glycerol carbonate. The same biocatalyst will be also used on non-conventional environment for the next stage.

 

Diseminare (Dissemination)

Conferinte (Conferences)

  • 8th International Conference on Renewable Resources & Biorefineries, June 2012, Toulouse, France –oral.
  • 244th ACS National Meeting in Philadelphia, Pennsylvania, August 19-23, 2012 – oral.
  • BioCat2012, Hamburg, Germany, September 2-6, 2012 – oral.
  • Cat4Bio, Thesaloniky, Greece, July 2012 – poster.
  • ECB15, Istanbul, Turkey, September 23-26, 2012 – poster.
  • COST Actin CM0903 (UBIOCHEM), 3rd Workshop, Thessaloniki (Greece), 1-3 November, 2012 – poster.
  • Green solvents for Synthesis, Boppard, Germany, 8-10 October, 2012 –oral.
  • 15th International Congress on Catalysis, Munich, Germany, 1-6 July, 2012 – poster.
  • ICCEC, Lyon, France, 2-6 September, 2012 – oral.

Publicatii (Publications)

  • M. Tudorache, L. Protesescu, A. Negoi, V.I. Parvulescu – ”Recyclable biocatalytic composites of lipase-linked magnetic macro-/nano- particles for glycerol carbonate synthesis”, Applied Catalysis A: General, 2012, 437-438, 90-95.
  • M.Tudorache, S. Nica, E. Bartha, I. Lupan, V.I. Parvulescu – „Sequential deracemization of sulfoxides via whole-cell resolution and heterogeneous oxidation”, Applied Catalysis A: General, 2012, 441-442, 42-46.

Stage IV     2013

Obiective

  • Testarea sistemului biocatalitic in medii neconventionale.
  • Managementul proiectului

Objectives (English)

  • Testing the biocatalytic system in nonconventional environment.
  • Project management.

Rezultate

Activitatea stiintifica desfasurata in cadrul acestei etape a avut drept scop principal caracterizarea eficientei sistemului biocatalitic construit si optimizat in etapele anterioare, cat si determinarea capacitatii acestui sistem de a functiona in medii neconventionale (lichide ionice si apa saraca in deuteriu). Sistemul biocatalitic dedicat sintezei glicerol carbonatului a fost testat pentru biocatalizatori cu design diferit (ELMP si CLEMPA) in care suportul necesar imobilizarii enzimei a avut structura variata. Pentru biocatalizatorii CLEMPA s-a incercat realizarea unui sistem „one-pot” de sinteza bazat pe tripla utilizare a dimetilcarbonatului drept agent de precipitare, reactant si mediu de reactie. Totodata, sinteza glicerol carbonatului s-a realizat in mediu de ultrasunete. Rezultatele obtinute au fost comparate cu cazul excitarii termice a sistemului. Mai mult, s-a incercat conturarea unei perspective de continuare a subiectului acestui proiect prin sinteza glicidolului din glicerol carbonat sau chiar glicerol via glicerol carbonat.

Results (English)

The research activity in this stage was focused on the characterization of the biocatalytic system in nonconventional environment (ionic liquid and depleted water). For this propose, both biocatalyst designs (ELMP and CLEMPA) were considered. They are different based on the solid support used for the enzyme immobilization. Moreover, “one-pot” system for glycerol carbonate synthesis was tested for CLEMPA biocatalyst. This idea came when we discovered that dimethylcarbonate used as reagent and solvent of the synthesis could be used also for biocatalyst preparation. The synthesis of glycerol carbonate was performed under termic and ultrasound conditions. Additionally, we made preliminary experiments for demonstrating the future perspectives os this project which are the synthesis of glycidol from glycerol carbonate and also direct from glycerol.

Diseminare (Dissemination)

Conferinte (Conferences)

  • CAFC, Turku, Finland, 16-19 June, 2013 –oral.
  • RomCat, Cluj, Romania, 29-31 May, 2013 – oral.
  • EU BC&E, Copenhagen, Danemark, 3-7 June, 2013 – oral.
  • ISGC2, La Rochelle, France, 21-24 May, 2013 – poster.
  • 23rd NACS, June, 2013 – poster.

Publicatii (Publications)

  • M. Tudorache, A. Negoi, L. Protesescu, V.I. Parvulescu – ”Biocatalytic alternative for bio-glycerol conversion with alkyl carbonates via a lipase-linked magnetic nano-particles assisted process”, Applied Catalysis B: Environmental, 2013, http://dx.doi.org/10.1016/j.apcatb.2012.12.033.
  • M. Tudorache, A. Nae, S. Coman, V.I. Parvulescu – ”Strategy of cross-linked enzyme aggregates onto magnetic particles adapted to the green design of biocatalytic synthesis of glycerol carbonate”, RSC Advances, 2013, 3, 4052-4058.
  • M. Tudorache, A. Negoi, B. Tudora, V.I. Parvulescu – ” Environmental-friendly strategy for biocatalytic conversion of waste glycerol to glycerol carbonate”, Applied Catalysis B: Environmental, 2013, 10.1016/j.apcatb.2013.02.049.

 

Pagină actualizată la 02 Decembrie 2014.