How to Resolve Unresolved Complex Mixture

Vladimir P. BeškoskiDepartment of Biochemistry, Faculty of Chemistry

University of Belgrade, Belgrade, Serbia vbeskoski@chem.bg.ac.rs

Institut Vinča20.11.2018.

Seminar and Workshop: Environmental Protection

Prevention, Monitoring and Remediation -Worldwide and Our Experiences

ENVIRONMENT IN SERBIA

Europe ~ 1,800,000 potentially contaminated sites/240,000 sites require remediation!

Official data from Serbia - 357 heavy polluted sites!

2

Locations of known illegal dumps in the territory of the Republic of Serbia, REPORT ON THE STATE OF THE ENVIRONMENT IN SERBIA 2009,

Ministry of Environment and Spatial Planning

3

3

THE ESSENCE OF BIOREMEDIATION

“BIOLOGICAL AGENT”

MICROORGANISMS

HARMFUL SUBSTANCES

SUBSTANCES WITHOUT

HARMFUL EFFECT ON THE

ENVIRONMENT AND HUMANS

+ BIOMASS

GREEN BIO/TECHNOLOGY!!!

4

FROM LABORATORY…

…VIA PILOT…150m

3of soil in “Oil Refinery

Pančevo”

TO INDUSTRIAL SCALE

BIOREMEDIATION

600 m3

of soil, JKP BE

5

BIOREMEDIATION FROM LABORATORY TO INDUSTRIAL SCALE

Methods

BiochemicalMicrobiologicalChemical

6

No. SampleTPH (g/kg) Reduction

of TPH (%)0 days 120 days

1. 3NST8

103,2

48,1 53

2. 3NST8-SG* 38,5 63

3. 3NST8-BS* 14,3 86

4. 1NS Sludge

412,5

221,7 46

5. 1NS Sludge-SG 88,8 79

6. 1NS Sludge-BS 47,0 89

Reduction of Total Petroleum Hydrocarbons (TPH) in experiments on the laboratory shaker (Erlenmeyer flasks) - gravimetric method

The degradation rate was from 1,8 – 6,4 g/kg/day.*Surface active substances

LABORATORY LEVEL

77

0

1

2

3

4

5

6

7

8

9

0 20 40 60 80 100 120 140

log

No

. of

mic

roo

rgan

ism

s (C

FU/g

)

TIME (days)

TC

ANA

HD

YM

Ps.

Bac.

LABORATORY LEVEL

BASIC MICROBIOLOGICAL DATA

TC-Total Chemoorganoheterotrophs; ANA – Anaerobic microorganisms; HD –Hydrocarbon degraders; YM – Yeast and Molds; Ps. – Pseudomonas spp.;

Bac. – Bacillus spp.

Milic et al., J Serb Chem Soc 2009;74:455-60

8

Jovančićević et al., Water Air and Soil Pollution 2008; 190:299-307

Graphical representation of the dynamics of pilot

bioremediation in the function of time

~30 g/kg

2,2 g/kg

(~93%)

173 mg/kg/day

292 mg/kg/day

64 mg/kg/day

0-60 days

60-90 days

90-180 days

21,5

7,8

Total:151 mg/kg/day

9

Comparative gas chromatograms of TPH at the beginning (Start) and after the completed bioremediation (End) at the pilot biopile

Start End(180 days)

Miletić et al., CLEAN – Soil, Air, Water 2014;42:1280–83

10

Reduction of TPH

5,2

2,1

1,3

0,3

Average rate of biodegradation0-50 days: 62 mg/kg/day50-100 days: 16 mg/kg/day100-150 days: 20 mg/kg/day

GAS CHROMATOGRAPHY IN BIOREMEDIATION

Beškoski et al., Chemosphere 2011;83:34-40

GC-FID

UCMUnresolved

ComplexMixture

11

Beškoski et al., J Serb Chem Soc 2010;75:1605–16

Fragmentograms of the isoprenoids (SIM, m/z 183) of the alkane fractions in the samples S-0 and S-150

GC-MS analysis - SIM mode

12

Fragmentograms of terpanes (SIM, m/z 191) of alkane fractions in samples S-0 and S-150.

Fragmentograms of steranes (SIM, m/z 217) of alkane fractions in samples S-0 and S-

150.

GC-MS analysis - SIM mode

Novaković et al., Environ Chem Lett 2012;10:287–94

13

Comprehensive GCxGC-MS analysisFaculty of Chemistry, University of Belgrade

Comprehensive GCxGC-MS analysis

UCM finally

resolved!

BIOCHEMICAL PROFILING OF MICROORGANISMS USING COMPREHENSIVE GCxGC-MS

Djurić et al., Int J Biol Macromol 2017;104:311-21.Izrael-Živković et al., Journal of Molecular Catalysis. B, Enzymatic 2017; in press

The fatty acid methyl ester profile of

Brachybacterium sp. CH-KOV3 a) GC–MS;

b) 2D GCxGC–MS.

16

Lama et al., ACS Sustainable Chemistry & Engineering 2017;5:2415-20Beškoski et al., CLEAN – Soil, Air, Water 2017;45:1600023Kannan et al., Environ Sci Pollut R 2015, in press

•TPH (>1000)•PAHs (>16) •PCBs (209)•Pesticides (>100)•Lignin•Metabolite profiling (metabolomics)•Biodegradation products – non targeted analysis•Natural products (plant extracts, propolis, honey…)•…

Comprehensive GCxGC-MS posibilities

17

Thank you for your attention