drug screening in soil algae - hevs.ch

DIE UNTERNEHMERISCHE HOCHSCHULE ® 6020 Innsbruck / Austria www.mci.edu

MCI MANAGEMENT CENTER INNSBRUCK Universitätsstraße 15 1

DRUG SCREENING IN SOIL ALGAE

Maren Kruus

05.09.2017

Summer School on Advanced Biotechnology 2017

Sion

http://www.google.at/url?sa=i&rct=j&q=&esrc=s&source=images&cd=&cad=rja&uact=8&ved=0CAQQjRw&url=http://www.presseportal.de/pm/66904/mci-management-center-innsbruck&ei=0xoIVLv-No7H7AaRm4HQAg&bvm=bv.74649129,d.ZGU&psig=AFQjCNE86H69qXFtZc1VN33AQlu5Y5MhzQ&ust=1409903699929425



Agenda

• Soil Algae

• Products and applications

• Workflow

• Cryopreservation

• Algae biomass cultivation

• Physico-chemical stress cultivation

• Harvesting and Extraction

• Screening



Soil Algae

• Estimated >500 Culturestrains, Slant agar cultures

• Soil-, Air- and Lichen-algae

• Origin: alpin area central europe

Features of Soil-Algae

• Primary producers

• Grout of the soil

• Symbiosis

• Survival under extreme conditions

Algae collection

ASIB, Institute of Botany, University Innsbruck

Prof. Georg Gärtner

MCI ASIB 505 collection maintains 58 green soilalgae and cyanobacteria strains wich are underactive research

Production of secondary metabolites



Products and applications

Docosahexaenoic Acid(DHA) C22H32O2

EicosapentaenoicAcid (EPA) C20H30O2

Mycosporine-like amino acids (MAAs)

http://bggworld.com/astaxanthin-astazinetm/

http://www.matreya.com/Products

AstaxanthinCanthaxanthin

Annual Review of Physiology, Volume 64, 2002

C40H52O4C40H52O2

http://www.theseus.fi/bitstream/handle/10024/121971/kruus_maren.pdf?sequence=1



Workflow

MCI Department ofBiotechnology & Food Engineering

Cultivation Harvesting TestingExtraction Analytics

Biomass

Stress

Solvent BioactivityCryopreservation



Cryopreservation

Advances of using cryopreservation as a long term:• Maintain their viability• At the low temperature cells are an inactive metabolically• Retaining microalgae ability and genetic integrity over long

periods• Lower collection maintenance costs• Safe

http://www.fertilityinstitutesandiego.com/our-services/fertility-treatment/in-vitro-fertilization-ivf/embryo-cryopreservation/

Cryoprotectant:Dimethyl sulfoxide (DMSO)Methanol (MeOH)

Thawing Culture for biomassproduction

Storage in liquid nitrogen

Algae culture added intocryovial

Pre-freezing



Biomass cultivation

Long term cultivation Agar culture Liquid culture Upscaling

Flat-platebioreactor

Polycarbonate bioreactor

Bubble columnSchott bottle



Stress cultivation

Light

Temperature

Nutrients

Salinity

pH

Microalgae High nitrates andhigh light

Biomass

First stage Second stage

Combination ofstress factores

Enhanced high valuables

combounds(pigments, lipids)

High salinityHigh lightHigh temperature Low nitrates

Lipid, astaxanthinand β-carotene

Biomass

Lipid contentandsaturation of

fatty acides

BiomassGrowth rate Lipid andluteincontent

Stress factors

Lipid andcarotenoidproductivity

Standard cultureconditions

Saturation offatty acid andastaxanthinproduction



Workflow



Biomass

Stress




Harvesting and Extraction

AlgalCulture

Centrifugationand biomass

washing

Lyophilization

Storage Celldisruption

Extractionand

separation

Concentrationand re-dissolve

FiltrationBioassay

andanalytics

ExtractionHarvesting

Main parameters driving the selection of an extraction method:• Extracted molecules• Velocity• Amount of solvent use /toxicity of solvent• Reproducibility• Extraction yield• Extraction sensitivity (pigments)• Selectivity• Protection of extraction molecules against

chemical transformation• Dimension• Cost• Easiness• Solvent safe equipments

• Liquid-liquid• Solid-liquid

Distruption solvents:• Methanol• Ethanol • Hexane• Dimethylformamid

(DMF)• Dichloromethane (DCM)• Acetone• Dimethyl sulfoxide

(DMSO)

Mechanical celldistruption:• Bead mill• Homogenization• Ultrasound• Sonication• Glass beads• Grinding (mortar

and pestle)• French pressure





Biomass

Stress




Testing and Analytics

• Disk Diffusion Method for

Antibacterial/antibiotics and

Antifungal Susceptibility Testing

• Toxicity, Inflammation, Oxidative

Stress (Peter Leitner)

Screening methods

Bioactivity Analytics (HPLC/GC – MS)

• PUFAs (DHA, EPA)• Mycosporine-like amino

acids (MAAs)• Pigments (Astaxanthin,

Cantaxanthin)

http://www.asmscience.org/content/education/imagegallery/image.2716 http://www.mdpi.com/1660-3397/10/6/1400/htm




Project High-lights

Results:

New Prasiola sp. strain (MCI-28) New structure of MAA Taxonimic classification: New tree and new

genera MCI-28 MCI ASIB 505 collection 40 strains classified 30 strains cryopreserved 105 extracts ready for analyse

Hits wich are under active research:

Algae strain MCI-31 Chromochloris zofingiensisare able to produce pigment Astaxanthin(C40H52O4) under stress conditions.

Algae strain MCI-45 Vischeria sp. are able toproduce PUFAs like EPA

Antimicrobial screening have showed somepositive results and further studies are required

Maximize the MAA production using UV-LED in 2stage



Thank you

FH-Prof. Dr. Christoph GriesbeckDr. Alexander TrockenbacherSebastian Perkams, MScPeter Leitner, MScBettina Rainer, MSc

Prof. Lukas HuberProf. Günther BonnDr. Ronald GstirDr. Thomas JakschitzDr. Oliver LutzMartin Fischnaller, MScDaniel Nothdurfter BSc

Institute of Botany, Prof. Georg Gärtner

Dr. rer. Nat. Daniel RemiasDI (FH) Klaus Krennhuber

COIN – Cooperation & Innovation




Thank you for your

attention!

drug screening in soil algae - hevs.ch

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