Toward the sustainable bioremediation of table olive processing wastewaters coupled with the generation of value-added products

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1 Toward the sustainable bioremediation of table olive processing wastewaters coupled with the generation of value-added products Eugenia Papadaki, Fani Th. Mantzouridou Laboratory of Food Chemistry & Technology, School of Chemistry, AUTH

2 2.6 million tons Argentina 3% Egypt 13% Morocco Peru Iran 5% 2% 2% Syria 6% Turkey 15% USA 3% Algeria 9% Italy 3% Greece 8% Spain 23% International Olive Oil Council (IOC), Economic: World table olive figures (2017)

3 Papadaki & Mantzouridou, Biochem. Eng. J. 2016, 112, Spanish-style green olives Californianstyle black-ripe olives Naturally black olives Wastewaters Storage in acidified brine or water (optional) Lye treatment Lye treatment and air oxidation Lye effluent Washings Washings and color fixing Washing water effluent Brining - Fermentation Brining Brining - Fermentation Brine Packaging

4 mᵌ wastewaters / ton olive Papadaki & Mantzouridou, Biochem. Eng. J. 2016, 112, Lye Washing waters Brine Total Naturally black olives Spanish-style green olives Californian-style black-ripe olives

5 Spanish-style green olives Californian-style black-ripe olives Naturally black olives Parameter Lye Washing waters Brine Lye Washing waters Brine Brine ph Electrical conductivity (ms/cm) Chemical oxygen demand - COD (g O 2 /L) Total phenolic compounds (g/l) Organic matter Phytotoxic activity Antimicrobial activity Requirements for the discharge of agro food industrial wastewaters set by the European Union COD < 125 mg O 2 /L Papadaki & Mantzouridou, Biochem. Eng. J. 2016, 112, European Parliament. Council Directive 91/271/EEC, Off. J. Eur. Communities 1991, L135, 40 52

6 Papadaki & Mantzouridou, Biochem. Eng. J. 2016, 112, Eutrophication of aquatic ecosystems Shifts in soil microflora Production of malodorous gases Wastewater detoxification (Ozonation, photocatalysis, wet air oxidation, electrochemical oxidation) Recovery of value-added products (Ultrafiltration, evaporation) Polluted side streams High energy requirements High cost technology Wastewater detoxification Production of high valueadded products Green technology Low cost technology

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8 Aggelis et al., Appl. Microbiol. Biotechnol. 2002, 59, Pleurotus ostreatus in lye effluents LACCASE ACTIVITY 52-76% Total phenolic compounds MANGANESE PEROXIDASE ACTIVITY NO REDUCTION OF PHYTOTOXICITY 50% Color

9 Kyriacou et al., Process Biochem. 2005, 40, Aspergillus niger in lye and washing water effluents 1 Biological treatment 9000 L Air-lift Bioreactor 71% COD 65% Total phenolic compounds 1.6% H₂O₂ Electrochemical oxidation Chemical Treatment Sedimentation 0.4% Ca(OH)₂ 93% COD 98% COD

10 Papadaki et al., J. Agric. Food. Chem. 2018, 66, Aspergillus niger in lye and washing water effluents 2 RP HPLC 280 nm OH RP HPLC 240 nm HO HO O O COOCH 3 COOCH 3 HO OH O O OH CH 3 CH 3 O g/l Initial concentration of total phenolic compounds 84% Total phenolic compounds (initial concentration 1.5 g/l) Untreated Treated Phytotoxicity NaOH Phenolics Lepidium sativum Lactuca sativa Phytotoxicity Phytotoxicity

11 Activated sludge 1 Single-step biological process in brine 88% COD 98% Total phenolic compounds Sequencing stirred batch reactor Air flow 550 L/h, dissolved oxygen mg/l No need for biomass acclimatization to salinity Multi-step biological process in lye effluents Anaerobic process 10 L Draw-and-fill 1 L Stirred Draw-and-fill 50% COD 13% Total phenolic compounds Ferrer-Polonio et al., Chem. Eng. J. 2015, 273, Aggelis et al., J. Agr. Eng. Res. 2001, 80, Aerobic process 84% COD 28% Total phenolic compounds In single step aerobic process, no phenol degradation was determined

12 Activated sludge 2 Mixture of table olive processing wastewaters from various processings 92% TOC (Total organic carbon) 83% Total phenolic compounds 20 L submerged membrane bioreactor Pore size 0.04 μm, flux operation 10 L/m 2 h, air flow rate 8 N dm 3 /min Stable performance at moderate biomass concentration (<10 g/l) Patsios et al., J. Chem. Technol. Biot. 2015, 91,

13 Nannochloropsis gaditana in washing waters 69% TOC (Total organic carbon) 72% Total phenolic compounds 1 L glass bottle aeration rate 0.5 vvm Natural light of 14h of light 10h of dark Sequential adaptation of microalga to wastewater at increased substrate concentrations (10-80%) ensures the process efficiency Serrano et al., J. Environ. Sci. Health A 2017, 1 6

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15 Methane (biogas) production using activated sludge 1 Primary component of biogas (55-75%) Methane In 2010 Europe s biogas production reached 10.9 millions of tons Biogas is a clean renewable fuel that can replace fossil fuels and reduce the negative environmental impact of the latter During anaerobic digestion, organic compounds are converted into biogas through the enzymatic activities of naturally occurring microorganisms under conditions of oxygen depletion Mes et al., ISBN , 2003, Papadaki & Mantzouridou, Biochem. Eng. J. 2016, 112,

16 Methane (biogas) production using activated sludge 2 Treatment of brine Methane (295 ml/g COD degraded ) 20 L Spherical magnetically 81-94% COD stirred digester Batch operation, 35 C Co-treatment of lye and washing water effluents with animal manure Methane (300 ml/g VS added ) 50 L stainless steel complete mix anaerobic digester Feed rate 1.66 L/d, agitation 60 rpm for 6 min/h, 55 C 81% TOC (total organic carbon) 17% Total phenolic compounds Olive mill wastewaters 260 ml CH4/g COD Cassava wastewaters 183 ml CH4/g COD Coconut oil cake 320 ml CH4/g VS Beltran et al., Hazard. Mater. 2008, 154, ; Zarkadas & Pilidis, Bioresour. Technol. 2011, 102, ; Martín et al., Process Biochem. 1991, 26, ; Intanoo et al., Bioresour. Technol. 2014, 173, ; Prabhudessai et al., J. Energy 2013, 1 7

17 Lactic acid and hydroxytyrosol production using lactic acid bacteria 1 Lactic acid Acidulant, flavor enhancer, antimicrobial preservative mainly in food and pharmaceutical industries, as a raw material in plastics production Hydroxytyrosol Antimicrobial, antioxidant, anticancer, anti-inflammatory, antidiabetic and neuroprotective properties Wang et al., J. Biosci. Bioeng. 2015, 119, Fernández-Mar et al., Food Chem. 2012, 130,

18 Lactic acid and hydroxytyrosol production using lactic acid bacteria 2 Treatment of washing waters with Lactobacillus pentosus 500 L Batch fermenter Initial ph value 5, 9 months, C 12.5 g/l Lactic acid 3.4 g/l Hydroxytyrosol 20% COD to 38g/L Evaporation to 10% (After ph correction to 8.3) Lactic acid /10 g ( 98%) Hydroxytyrosol 250 /25 mg ( 98%) Concentrates (COD 399g/L) g/l lactic acid 36.4 g/l Hydroxytyrosol Distillates (COD 2.1g/L) Not detected lactic acid <0.01g/L Hydroxytyrosol Barley starch (Lactobacillus casei) 162 g/l Lactic acid Sugar molasses (Enterococcus faecalis) 96 g/l Lactic acid Olive mill wastewaters (liquid-liquid extraction) 1.2 g/l Hydroxytyrosol Brenes et al., Chem. Technol. Biotechnol. 2004, 79, ; Linko & Yavanainen, Enzyme Microb. Technol. 1996, 19, ; Wee et al., Enzyme Microb. Technol. 2004, 35, Kalogerakis et al., J. Environ. Manage. 2013, 128,

19 Toward the development of lactic acid bacteria starter cultures for table olive production Lactobacillus plantarum 1High tolerance to acidic conditions 2Extracellular polysaccharide production 3Hydrolysis of oleuropein Enterococcus faecium Lactic acid bacteria isolates from brine 1High tolerance to acidic conditions 2Extracellular polysaccharide production 3Lipolytic activity 4Hydrolysis of oleuropein Lactobacillus spp. Enterococcus spp. Leuconostoc mesenteroides 1 Tolerance to acidic conditions 2Lipolytic activity 3Extracellular polysaccharide production 4Hydrolysis of oleuropein Leuconostoc spp. Control of fermentation Elimination of debittering step Fendri et al., Environ. Technol. 2013, 34,

20 Table olive processing wastewaters are high polluted, but also have the potential to be used as fermentation feedstocks. The use of fungi, activated sludge and microalgae for the treatment of table olive processing wastewaters was really effective, causing important detoxification of these wastes. Findings are expected to provide useful information for the subsequent treatment of residual contaminants. To ensure the economic feasibility of the process, the production/recovery of important to the market chemicals through the wastewater treatment should be targeted. Biotechnological production of methane, lactic acid and hydroxytyrosol is competitive to other wastes. Oleuropeinolytic lactic acid bacteria isolates have the potential to be used as starter cultures for table olive fermentation by eliminating the debittering step.

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