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Abstract 4-Carboxybenzaldehyde (4-CBA), a major component of purified terephthalic acid wastewater, is toxic to living organisms and required to be removed before the discharge of treated wastewater in a natural water body. In this work, the adsorptive removal of 4-CBA was studied using bagasse fly ash (BFA), a waste product from the sugar industry. The adsorption capacity of BFA was compared with the commercially available adsorbent, granular activated carbon (GAC). A 4-factorial, 5-stage central composite design (CCD) was used to optimize 4-CBA removal and adsorption uptake by BFA using response surface methodology (RSM). The variables considered for the study were pH, adsorbent dose (m), initial concentration (Co) and time (t). At the optimum treatment conditions of pH = 4, m = 9 g/L, Co = 100 mg/L and t = 7.5 h, the removal efficiency and adsorption uptake of 4-CBA on BFA were found to be 79% and 9.9 mg/g, respectively. BET surface area of BFA was determined to be 284 m2/g. A kinetic study was performed using a first- and second pseudo-order model. The adsorption equilibrium data were fitted for various adsorption models. A positive value $$\Delta H^{0}$$ Δ H 0 indicates that the adsorption process is endothermic. This study indicated that BFA is a cost-effective adsorbent for the removal of 4-CBA with high adsorption capacity and fast kinetics. Graphical Abstract

Constructed wetlands are commonly used to treat domestic wastewater, but the extent to which microplastics are retained in constructed wetlands and the impact this has on their efficiency are not known. In this context, we investigated retention of microplastics (microbeads and fibres) and their effects on the removal of carbonaceous (expressed as chemical oxygen demand (COD), biochemical oxygen demand (BOD[sub]5[sup]-), and dissolved organic carbon (DOC)), nitrogenous (N-NH[sub]4[sup]+, N-NO[sub]3[sup]-, and N-NO[sub]2[sup]-), and phosphorous (P-PO[sub]4[sup]{3-}) matters in a horizontal sub-surface flow laboratory constructed wetland (LCW) under continuous flow. The LCW was in operation for 257 days, with the first 78 days without microplastics and microplastics added weekly during the next 179 days. The presence of microplastics had no significant effect on COD, BOD[sub]5, DOC, N-NH[sub]4[sup]+, and N-NO[sub]3- removal, while a statistically significant decrease in N-NO[sub]2[sup]- and an increase in P-PO[sub]4[sup]{3-} concentrations in the effluent were observed after the addition of microplastics. Microplastics were efficiently retained in the LCW, and only 0.296% and 0.003% of the total microbeads and fibres, respectively, were detected in the effluent. Thus, it is evident that constructed wetlands are an effective barrier to the release of microplastics into the aquatic environment, but the high retention and thus continuous accumulation of microplastics may alter the nutrient cycling in constructed wetlands.

Zaradi vse večje naseljenosti obalnih okolij je morski ekosistem izpostavljen različnim antropogenim motnjam, med katere uvrščamo tudi onesnaževanje z odpadnimi vodami. Vpliv vnosa hranil in mikroorganizmov z očiščeno odpadno vodo iz čistilnih naprav na obalno morsko združbo bakterij je slabo poznan, zato smo se v nalogi osredotočili na odziv metabolizma morskih bakterij na vnos očiščene odpadne vode iz komunalne čistilne naprave v modelnem eksperimentalnem sistemu. Ugotovili smo, da je vnos hranil iz odpadne vode, s katerim smo znižali molarno C:N razmerje raztopljene organske snovi v eksperimentalni steklenici, povečal bakterijsko rast, celično dihanje in produkcijo bakterijske biomase. Najvišjo bakterijsko abundanco smo izmerili v steklenicah, obogatenih z nefiltrirano odpadno vodo, k čemur so pripomogle tudi bakterije iz odpadne vode. Izračuni so pokazali, da so večji del asimiliranega ogljika iz odpadne vode bakterije usmerile v proces celičnega dihanja, kar se ujema z nizkimi vrednostmi bakterijske učinkovitosti rasti. Ker se z odpadno vodo v morja vnašajo tudi patogeni organizmi, med katerimi so potencialno lahko tudi bakterije rodu Vibrio, smo v drugem delu naloge nastavili eksperiment z različnimi čistimi kulturami Vibrio spp. Seve, izolirane iz različnih lokacij, smo izpostavili gojiščem z različno vsebnostjo hranil. Rast, celično dihanje in produkcija biomase posameznega izolata je bila najvišja v tistem tretmaju, ki je bil po hranilih najbolj podoben mestu izolacije bakterijskega seva, kar kaže, da so bili ti sevi dobro prilagojeni na okoljske razmere, iz katerih so bili izolirani. The marine ecosystem can be affected by various anthropogenic activities, including wastewater pollution, due to population growth in coastal areas. The impact of nutrients and microorganisms from wastewater treatment plants on the bacterial community in coastal waters is poorly understood. Therefore, we focused our study on the response of marine bacterial metabolism to the addition of wastewater in microcosm experiments. We found that nutrient uptake from wastewater reduced the C:N molar ratio of dissolved organic matter in the experimental bottle and enhanced bacterial growth, respiration, and production. The highest bacterial density was measured in bottles with unfiltered wastewater, which also contained bacteria from wastewater treatment plants. Calculations showed that bacteria consumed most of the assimilated carbon in the respiration process, which is consistent with the low values of bacterial growth efficiency. Pathogenic organisms may also enter the sea with the wastewater, which can potentially include bacteria of the genus Vibrio. In the second part of the thesis, we focused on pure Vibrio cultures. We exposed Vibrio isolates to growth media with different nutrient concentrations. Growth, respiration, and production of each isolate were highest at the nutrient conditions most similar to the isolation site, suggesting good bacterial adaptation to the environmental conditions from which strains were isolated.

Aminoglikozidni antibiotik gentamicin sulfat v okolju predstavlja vedno večji problem, saj ga je nemogoče v celoti odstraniti iz odpadnih vod. Gentamicin sulfat je najbolj uporabljen antibiotik v skupini aminoglikozidnih antibiotikov in se uporablja za zdravljenje gram-negativnih okužb, izoliran pa je iz bakterije Micromonospora purpurea. Napredni oksidacijski procesi so najučinkovitejše nove metode, ki se uporabljajo za čiščenje in dezinfekcijo odpadnih vod, predvsem farmacevtskih ter za čiščenje biološko strupenih ali nerazgradljivih snovi. Osredotočili smo se na ozonacijo, saj velja za eno najuspešnejših metod čiščenja. V diplomski nalogi smo pod izbranimi pogoji ozonirali raztopino GS in učinkovitost procesa spremljali z določanjem kemijske potrebe po kisiku in koncentracije celotnega organskega ogljika. The aminoglycoside antibiotic gentamicin sulfate is an increasing problem in the environment, as it is impossible to completely remove it in wastewater. Gentamicin sulfate is the best antibiotic in the group of aminoglycoside antibiotics and is used to treat gram-negative infection and it’s isolated from bacteria Micromonospora purpurea. Advanced oxidation processes are the most effective new methods used for treatment and disinfection of wastewater, especially pharmaceuticals and for cleaning biologically toxic or non-degradable materials. We focused on ozonation, as it is considered one of the most successful treatment methods. In a degree thesis, we ozonated the GS solution under selected conditions and monitored the effectiveness of the process by determining chemical oxygen demand and concentration of total organic carbon.

Odpadna voda je voda, ki nastane po uporabi sladke, surove, pitne ali slane vode v različnih namenskih aplikacijah ali procesih. V grobem lahko odpadne vode delimo na tri glavne tipe: komunalne oz. gospodinjske odpadne vode, industrijske odpadne vode in meteorne oz. padavinske odpadne vode. Pri onesnaževalih v odpadnih, ki prihajajo iz različnih virov, gre za naravna onesnaževala ali sintetična onesnaževala. Kadar je voda onesnažena in je potrebno dekontaminiranje, je treba izbrati najboljši pristop čiščenja, da se dosežejo cilji dekontaminacije. Pri tem se uporabljajo različni postopki in tehnike. Membranska filtracija postaja pogosta tehnika za doseganje primerne kvalitete vode. Uporaba ogljikovih nanocevk pri tem kaže velik potencial zaradi kemije površine, morfologije, kemijske in fizikalne interakcije z organskimi in anorganskimi snovmi, dobrih adsorpcijskih lastnosti in številnih možnosti funkcionalizacije oz. modifikacije. V študijah so že razvili različne membranske filtre za praktično uporabo in z njimi dokazali njihovo visoko učinkovitost. Med najučinkovitejše spada elektrokemijski filter narejen iz skupkov ogljikovih nanocevk sklopljen z in situ generacijo H2O2. Kljub temu pa visoka cena materiala, zamašitve filtra in ekološka problematika ogljikovih nanocevk predstavljajo problem za vsakdanjo uporabo pri čiščenju odpadnih vod. V ta namen bi bilo potrebno narediti več študij, predvsem na večji skali. Wastewater is water that is produced after fresh, raw, drinkable, or saline water has been used in various applications or processes. Roughly speaking, wastewater can be divided into three main types: municipal or domestic wastewater, industrial wastewater and stormwater or precipitation wastewater. The pollutants in wastewater come from a variety of sources, either natural pollutants or synthetic pollutants. When water is contaminated and decontamination is necessary, the best treatment approach must be chosen to achieve the decontamination objectives. Different processes and techniques are used. Membrane filtration is becoming a common technique to achieve adequate water quality. The use of carbon nanotubes in this context shows great potential due to their surface chemistry, morphology, chemical and physical interaction with organic and inorganic substances, good adsorption properties and many possibilities for functionalisation or modification. Various membrane filters have already been developed for practical applications and their high efficiency has been demonstrated in studies. Among the most effective is an electrochemical filter made of carbon nanotube clusters coupled with in situ generation of H2O2. However, the high cost of the material, the fouling of the filter and the ecological problems of carbon nanotubes are a challenge for everyday use in wastewater treatment. More studies should be carried out for this purpose, especially on a larger scale.

V diplomskem delu so na kratko opisani antibiotiki, njihove vrste, delovanje, kemijska struktura in zakaj postanejo bakterije odporne na njihovo delovanje. Prav tako so v diplomskem delu opisane čistilne naprave in njihovo delovanje, katerim sledi podrobnejši opis različnih vrst naprednih oksidacijskih procesov z opisom reakcij, ki prihajajo v veljavo v sistemih čistilnih naprav. Napredni oksidacijski procesi lahko povečajo biorazgradljivost odpadnih vod in s tem pripomorejo pri odstranjevanju antibiotikov prisotnih v odpadnih vodah in zmanjševanju njihovih emisij v okolje. In the diploma thesis, there is a brief description of antibiotics, their types, action mechanisms, chemical structure and why bacteria become resistant to them. After that, a description of wastewater treatment plants and their working progress takes place, followed by a more detailed description of advanced oxidation processes with a closer look into the reactions of different advanced oxidation processes, which are being enforced in wastewater treatment plant systems. Advanced oxidation processes can enhance the biodegradability of wastewater, which helps with the removal of antibiotics present in wastewater and to reduce their emissions into the environment.

Ostanki zdravilnih učinkovin in metabolitov v okolju so opredeljeni kot novodobna mikroonesnažila. V okolje vstopajo predvsem kot posledica uporabe v humani in veterinarski medicini. Ugotovili so, da so čistilne naprave glavni točkovni vir izpustov, saj njihova tehnologija ni prilagojena za čiščenje tovrstnih mikroonesnažil. Odpadna voda se iz čistilnih naprav zliva v površinske in/ali podtalne vode, ki so vir pitne vode. Koncentracije ostankov zdravilnih učinkovin v okolju so večinoma nizke: do nekaj μg/L v odpadnih vodah, kjer so običajno izmerjene najvišje vrednosti za posamezen analit, in v padajočem trendu v površinskih in pitnih vodah. V slednjih so koncentracije najpogosteje v območju pg/L. Kljub nizkim koncentracijam pa je z vidika toksičnosti nujno treba obravnavati kombinacijo vseh zdravilnih učinkovin, ki imajo lahko negativne učinke na izpostavljene vodne organizme in posredno tudi na človeško zdravje. Nekatere raziskave poročajo o biokopičenju v vodnih organizmih in negativnih učinkih, kot so morfološke, citološke in vedenjske spremembe. Prve raziskave pojavljanja ostankov zdravilnih učinkovin v okolju so bile objavljene že leta 1970, porast pa beležimo v zadnjih letih, kar je posledica razvoja naprednih instrumentalnih metod, ki omogočajo zaznavo in merjenje zdravilnih učinkovin že pri zelo nizkih koncentracijah. Okoljska analitika je običajno sestavljena iz dveh delov: priprave vzorca ter instrumentalne metode. Glede na objavljeno literaturo je metoda prvega izbora za pripravo vzorca ekstrakcija na trdnem nosilcu in instrumentalna analiza s tekočinsko kromatografijo sklopljena z masno spektrometrijo. V doktorski nalogi smo se najprej osredotočili na razvoj ustrezne metodologije za pripravo vzorcev. Ločeno smo razvijali postopke za ekstrakcijo iz odpadnih, površinskih in pitnih voda ter za ekstrakcijo iz ribjih tkiv. Za ekstrakcijo iz odpadnih voda smo razvili dva inovativna pristopa ekstrakcije: prvi je pol-avtomatiziran sistem SPE-DEX, ki je osnovan na ekstrakciji na trdnem nosilcu (ang. solid phase extraction, SPE), drugi pristop pa je ekstrakcija Twister®, ki je osnovana na sorptivnem mehanizmu. Oba pristopa smo ovrednotili, vendar izbrali prvega, ki se je izkazal za bolj primernega za večje število zdravilnih učinkovin (> 100). Razvito metodo smo primerjali z najpogosteje uporabljenim ročnim pristopom SPE ter potrdili primerljive validacijske parametre. Metoda SPE-DEX ima poleg tega številne prednosti: gre za avtomatiziran sistem, ki ne zahteva stalne prisotnosti operaterja, s čimer se izboljša ponovljivost med vzorci ima manjšo možnost mašenja ekstrakcijskih diskov omogoča hkratno filtracijo z ekstrakcijo neposredno na aparaturi, kar znatno skrajša čas priprave vzorca in še pomembnejše prepreči izgube analitov zaradi morebitne adsorpcije na odfiltriran material ali filter. Z metodo SPE-DEX smo preverjali prisotnost ostankov zdravilnih učinkovin in metabolitov v odpadnih vodah. Vzorčili smo na 19 čistilnih napravah, pri čemer smo vzorec 33-krat vzeli na iztoku in 13-krat na vtoku. Ekstrakte smo analizirali z visoko občutljivo metodo LC-MS/MS. Po zahtevnem razvoju in optimizaciji metode je analiza, ki je zajemala ločitev in detekcijo vseh 104 analitov, trajala 8 minut. V odpadnih vodah na iztoku smo potrdili 94 % vseh tarčnih analitov (n = 104) v vsaj enem vzorcu v povprečnih koncentracijah od 1 ng/L do 11 μg/L. Na vtoku smo potrdili prisotnost 87 % vseh tarčnih analitov (n = 104) v vsaj enem vzorcu, povprečne koncentracije pa so bile v območju od 1 ng/L do 116 μg/L. Na vtoku smo najvišjo povprečno koncentracijo ugotovili za metabolit salicilno kislino (116 μg/L), prisotna je bila v 92 % vzorcev (n = 13). V visokih koncentracijah so bili prisotni še paracetamol (18 μg/L) in metformin (17 μg/L), oba v vseh vzorcih, in kofein (16 μg/L) v 92 % vzorcev. Skupno je bilo v koncentracijah nad 1 μg/L prisotnih 12 analitov s pogostostjo pojavljanja nad 85 %, z izjemo lakozamida (7 μg/L), ki je bil prisoten le v 1 vzorcu. Stalno prisotnost v vseh vzorcih smo potrdili za 31 analitov od skupno 104. Na iztoku smo v najvišji povprečni koncentraciji ugotovili tramadol (11 μg/L), v vseh vzorcih (n = 33). V visokih koncentracijah so bili prisotni še: hidroklorotiazid (5 μg/L) v 97 % vzorcev, metformin (4 μg/L) v 76 % vzorcev in salicilna kislina (4 μg/L) v 37 % vzorcev. Skupno je bilo v koncentracijah nad 1 μg/L prisotnih 10 analitov s pogostostjo pojavljanja nad 60 %, z izjemo fluoksetina (1 μg/L), ki je bil prisoten le v treh vzorcih, in salicilne kisline. Stalno prisotnost v vseh vzorcih smo potrdili za 14 analitov od skupno 104. Na osnovi pridobljenih podatkov smo oblikovali ožji izbor analitov za spremljanje njihove prisotnosti v površinskih vodah. Vključili smo le analite, ki smo jih zaznali v več kot polovici vzorcev na iztoku iz čistilne naprave in v višjih koncentracijah skupno smo spremljali 44 analitov. Razvili smo metodo ekstrakcije na ročnem sistemu SPE, ki nam je omogočal zadostno koncentriranje z namenom doseganja čim nižjih mej določitve. Metodo smo ovrednotili in pokazali ustreznost validacijskih parametrov. Uporabnost metode smo preverili z analizo 6 vzorcev površinskih voda, od tega 5 vzorcev rečnih voda in 1 vzorca jezerske vode. Potrdili smo prisotnost 43 tarčnih analitov (od skupno 44), večinoma v koncentracijah pod mejo določitve, merljive koncentracije pa so bile v območju od 0,08 ng/L do 47 ng/L. V vseh vzorcih smo potrdili prisotnost 29 analitov. Najvišja absolutna izmerjena koncentracija je bila za valsartan, ki je znašala 47 ng/L (povprečna koncentracija 11 ng/L, mediana 4 ng/L), in za gabapentin, ki je dosegla 45 ng/L (samo enkrat nad LOQ (LOQ = 40 ng/L)). Pogosto sta bila zaznana še tramadol (v vseh vzorcih, n = 6) v povprečni koncentraciji 5 ng/L (mediana 1,2 ng/L) in sulfametoksazol (v 67 % vzorcev) v povprečni koncentraciji 6 ng/L (mediana 6,3 ng/L). Raziskave smo nadaljevali z razvojem metodologije za ekstrakcijo iz pitnih voda. Uporabili smo razvito metodo SPE za površinske vode in povečali volumen nanešenega nanos vzorca, da bi povišali faktor koncentriranja. Metodo smo ovrednotili in pokazali ustreznost validacijskih parametrov. Preverili smo prisotnost ostankov zdravilnih učinkovin in metabolitov v 18 vzorcih pitnih voda. V vsaj enem vzorcu smo potrdili 24 % izbranih analitov (n = 54). Koncentracije so bile izredno nizke od 0,04 do 26,0 ng/L. V vseh vzorcih je bil prisoten le azitromicin, in sicer v povprečni koncentraciji 2,6 ng/L (mediana 0,7 ng/L). V najvišjih koncentracijah sta bila ugotovljena azitromicin (26 ng/L) in kofein (14 ng/L). Sledil je izračun vseživljenjske izpostavljenosti človeka izbranim zdravilnim učinkovinam zaradi vnosa pitne vode. Pri izračunu smo upoštevali življenjsko dobo 81 let in povprečni vnos pitne vode 2 L/dan. Na osnovi pridobljenih podatkov bi vseživljenjska izpostavljenost znašala 3 mg, iz česar sklepamo, da je tveganje za človeka minimalno. Primerjali smo rezultate glede na posamezne tipe vode in ugotovili, da smo vse zdravilne učinkovine, katerih prisotnost smo potrdili v pitni vodi, zaznali tudi v površinskih in odpadnih vodah. Iz izračunanih koncentracij lahko sklepamo, da je verjetnost pojavljanja v površinskih in pitnih vodah večja, kadar je izmerjena zelo visoka koncentracija v odpadnih vodah (iztok) ter kadar je posamezni analit v odpadnih vodah zaznan zelo pogosto. Raziskave smo dopolnili z ugotavljanjem prisotnosti ostankov zdravilnih učinkovin v ribah. V ta namen smo razvili metodo za ekstrakcijo iz ribjega tkiva, da bi lahko opredelili tveganje za človeka zaradi uporabe rib v prehrani. Za ekstrakcijo smo uporabili metodo QuEChERS, ki smo jo kot prvi prilagodili za ekstrakcijo ostankov zdravilnih učinkovin iz ribjih tkiv. Validirano metodo smo prenesli na 13 vzorcev ribjih mišic, kože in škrg. V vseh treh tipih tkiva smo potrdili prisotnost ostankov zdravilnih učinkovini in metabolitov. V vsaj enem vzorcu smo potrdili 42 vseh tarčnih analitov (n = 56). V vseh je bil prisoten le metoprolol (< LOQ–0,7 ng/g). V več kot polovici vzorcev smo zaznali še kofein (< LOQ–3,2 ng/g), valsartan (0,1–0,4 ng/g) in metabolit kofeina 1,7-dimetilksantin (v vseh vzorcih < LOQ). Najvišje povprečne koncentracije smo izmerili za ciprofloksacin (19,2 ng/g), enalapril (16,2 ng/g) in haloperidol (14 ng/g). Koncentracije zdravilnih učinkovin v mišicah rib, katere so del prehranjevalne verige, so bile zelo nizke, in sicer med 0,07 ng/g in 1,1 ng/g (povprečno 0,3 ng/g). Posledično lahko sklepamo, da je tveganje za človeka zanemarljivo. Najvišje koncentracije smo izmerili v škrgah. Z raziskavami smo potrdili prisotnost zdravilnih učinkovin v odpadnih, površinskih in pitnih vodah ter v ribjih tkivih. Potrdili smo tudi vpliv odpadnih voda na vstopanje zdravilnih učinkovin v okolje, zato smo razvili postopek čiščenja, ki bi učinkovito razgradil ostanke mikroonesnažil. Uporabili smo elektrokemijsko oksidacijsko metodo na osnovi z borom dopirane diamantne elektrode, ki z nastajanjem reaktivnih hidroksilnih radikalov in situ omogoča mineralizacijo organskih snovi. Z razvito elektrolitsko celico smo na laboratorijskem nivoju v sintetični zmesi razgradili več kot 85 % začetne vrednosti za 7 od 10 preiskovanih spojin, za 3 spojine pa smo začetno vrednost razpolovili. Nekoliko slabšo učinkovitost, do 70 % razgradnje, smo dosegli v realnih vzorcih odpadnih voda, ki smo jih izpostavili razvitemu postopku čiščenja na laboratorijskem nivoju. Slabšo učinkovitost pripisujemo krajšemu času postopka, s katerim smo želeli simulirati realne pogoje, in tudi večji količini organskega materiala, za mineralizacijo katerega so se dodatno porabljali nastali hidroksilni radikali. V okviru evropskega projekta LIFE PharmDegrade smo razvito metodologijo prenesli v realno okolje na biološko čistilno napravo kot dodatni postopek čiščenja. Po nadaljnji optimizaciji (večje število vzporedno vezanih celic, večkratni prehod, višja gostota toka in dodatno čiščenje s filtri) smo povečali razgradnjo izbranih zdravilnih učinkovin med procesom čiščenja za povprečno 53 % oziroma smo s celotnim postopkom čiščenja v biološki čistilni napravi in z naprednim oksidacijskem postopkom uspešno razgradili več kot 86 % začetne vrednosti posameznega analita. Rezultati doktorske disertacije so osvetlili problematiko ostankov zdravilnih učinkovin in metabolitov v okolju. Kot novost na področju okoljske analitike smo razvili instrumentalno metodo za sočasno analizo 104 analitov, ki bo znatno olajšala ugotavljanje prisotnosti številnih ostankov zdravilnih učinkovin in metabolitov v okoljskih vzorcih. Z razvito metodologijo smo celovito ovrednotili njihovo prisotnost v odpadnih, površinskih in pitnih vodah, s čimer smo kot prvi predstavili podatke za področje Slovenije in zapolnili vrzel s tega dela Evrope. Na osnovi koncentracij v okoljskih vodah, s poudarkom na pitnih vodah, in ribjem tkivu lahko opredelimo obremenitev okolja in človeka. Glede na izmerjene vrednosti lahko sklepamo o minimalnem tveganju za človeka ter nekoliko večjem za nižje organizme, ki so vse življenje izpostavljeni izpustom iz čistilnih naprav. Pri tem moramo dodati, da se bo s povečevanjem uporabe zdravil tveganje nenehno stopnjevalo. Doktorsko nalogo smo zaključili z razvojem naprednega postopka čiščenja odpadnih voda. Razviti postopek za razgradnjo zdravilnih učinkovin iz okolja predlagamo kot enega od možnih dodatnih postopkov čiščenja na obstoječih čistilnih napravah. Z njim bi znatno zmanjšali izpuste zdravilnih učinkovin v okolje in posledično tudi obremenitev okolja. Predstavljeni podatki so lahko osnova za postopno vpeljavo rutinskega spremljanja izpustov zdravilnih učinkovin v okolje, prav tako pa lahko služijo kot predlog za spremljanje tarčnih snovi oziroma kot dopolnitev Nadzornega seznama (Izvedbeni sklep Komisije (EU) 2015/495 o določitvi nadzornega seznama snovi za spremljanje na ravni Unije na področju vodne politike v skladu z Direktivo 2008/105/ES Evropskega parlamenta in Sveta Evrope). Pharmaceuticals have been recently classified as emerging pollutants. They enter the environment mainly because of usage in human or veterinary medicine. Wastewater treatment plants are known to be the major point source of releasing considerable amounts of pharmaceuticals into the aquatic environment. Furthermore, they continuously seep into surface water or groundwater, which could be a source of drinking water. The concentrations measured in the environment are relatively low a few μg/L in wastewaters, where the highest concentrations are generally found, while the concentrations measured in surface or drinking waters are notably lower in the range of ng/L or pg/L. Despite these low concentrations of each individual pharmaceutical, the mixture of all present multiple substances may pose a great risk to non-target organisms or even human health. Various studies have already shown the bioaccumulation of pharmaceutical compounds and their negative effects such as morphological, cytological and behavioural malfunctions. Pharmaceutical occurrences in the environment were first published in 1970. However, in recent years, there has been an increase of similar papers, most probably owing to the improvement of analytical methods which enable the analysis of large numbers of substances in extremely low concentrations. Most often, an environmental analysis consist of a combination of appropriate sample preparation and a highly sensitive analytical method. According to the currently available scientific literature, the liquid chromatography coupled with tandem mass spectrometry in combination with a sample pre-concentration step seems to be the method of choice. In this doctoral thesis, we researched the sample preparation and the instrumental analytical methods. Furthermore, we developed a liquid chromatography method coupled with tandem mass spectrometry for the determination of pharmaceutical residues in wastewaters, surface waters, drinking waters, and in fish tissue. Regarding the extraction from wastewaters, two innovative approaches were developed. Firstly, an extraction based on solid phase extraction, namely the semi-automated SPE-DEX method. The second method was the stir-bar sorptive extraction. Both methods were validated however, the first one was better suited for monitoring a large number of active substances (> 100). Afterwards, a comparison between the semi-automated and manual solid phase extraction confirmed the similarity of the validation parameters. Moreover, the semi-automated system has several advantages: automaticity, a high rate of sample processing, simultaneous filtration and extraction, better repeatability, and a lower probability of extraction disc clogging. Therefore, the semi-automated SPE-DEX method was chosen for the analysis of wastewaters from 19 wastewater treatment plants. A total of 46 wastewater samples were collected 33 effluent wastewaters and 13 influent wastewaters in the period from May to August 2016. The presence of 104 pharmaceutical residues was tested using liquid chromatography coupled with tandem mass spectrometry with a single run of 8 minutes per sample. Regarding the influent samples, a total of 90 analytes out of 104 were present in at least one sample at average concentrations between 1 ng/L and 116 μg/L. In the effluent samples, we detected a total of 98 analytes out of 104 in at least one sample. The average concentrations were between 1 ng/L and 11 μg/L. In the influent wastewater samples, salicylic acid had the highest average concentration (116 μg/L) and was present in 92% of the samples. High concentrations were also measured for paracetamol (18 μg/L), metformin (17 μg/L), and caffeine (16 μg/L). Altogether, 12 analytes with a frequency of over 85% were present in concentrations above 1 μg/L, with the exception of lacosamide (7 μg/L), which was present only in one single sample. 31 analytes were detected in all samples. Regarding the highest concentrations found in effluent samples, the following analytes were present: tramadol (11 μg/L), hydrochlorothiazide (5 μg/L), metformin (4 μg/L) and salicylic acid (4 μg/L). Altogether 10 analytes with a frequency of over 60% were present in concentrations above 1 μg/L, with the exception of fluoxetine (1 μg/L), which was present in three samples and salicylic acid in one third of the samples. 14 analytes were detected in all samples. The analysis was followed by the development of an extraction method for surface waters. A manual solid phase extraction was used because it enables a higher concentration step. Based on the previous analysis, the most frequent effluent wastewater concentrations of pharmaceuticals were used as the basis for pre-selection of target compounds in surface waters. Altogether 44 analytes were included. The optimised solid phase extraction was evaluated and it showed the suitable validation parameters. Finally, six samples from different Slovene regions (five river samples and one lake sample) were analysed. 42 of all target pharmaceuticals (n = 43, one analyte was excluded due to the presence of a high background signal) were detected in at least two samples 29 of them were detected in all samples. The concentration range of the measured pharmaceuticals was between 0.08 and 47 ng/L, the highest measured concentration was for valsartan (average 11 ng/L, median 4 ng/L). Furthermore, high concentrations were measured for tramadol, which had an average concentration of 5 ng/L (median 1.2 ng/L), and sulfamethoxazole with an average concentration of 6 ng/L (median 6.3 ng/L). The developed extraction method was adjusted for the extraction from drinking water samples. The sample volume was higher, thus increasing the concentration factor. The optimised method was evaluated and was shown to provide suitable validation parameters, especially the higher sensitivity. The method was applied to 18 drinking water samples. Among the 54 investigated substances, we confirmed the presence of 13 pharmaceuticals in at least one sample. Azithromycin was present in all samples. In general, the concentrations were very low, typically below 1 ng/L the highest concentration was determined for azithromycin (26 ng/L) and caffeine (14 ng/L). The obtained data was used to speculate the amount of the analysed pharmaceuticals that humans take in with drinking water throughout their lifespan. We postulated the life span of 81 years and the consumption of 2 L of water daily. The sum of all monitored contaminants based on their highest measured values would be 3 mg in 81 years. Therefore it can be concluded that the risks the monitored pharmaceuticals present to human health are negligible. Regarding all three types of tested samples, it can be observed that all the pharmaceuticals present in drinking waters were also detected in surface and wastewaters. From the measured concentrations, the following conclusion can be drawn: The probability of pharmaceutical occurrence in surface and drinking water is greater when a very high concentration was measured in wastewater effluents, and/or when the individual analyte in the wastewaters was detected very frequently. In the last part of the sample preparation development, we optimized a QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe) extraction protocol for extraction of drug residues from fish tissue. The optimised method was validated for 56 analytes. The method was used for monitoring target analytes in fish tissue. 13 samples of muscle, skin and gill tissues were analysed. The study showed the detection of 75% of all target analytes in at least one sample. Metoprolol was detected in all samples (< LOQ–0.7 ng/g). Furthermore, caffeine (< LOQ–3.2 ng/g), its metabolite 1,7-dimethylxanthine (< LOQ) and valsartan (0.1–0.4 ng/g) were detected in more than half of the samples. The highest average concentrations were measured for ciprofloxacin (19.2 ng/g), enalapril (16.2 ng/g) and haloperidol (14.0 ng/g) in the skin and gills. Interestingly, the concentrations measured in the muscles were lower or even below the limits of quantification. Regarding these data, human exposure via the ingestion of fish muscles is most probably low or negligible. The research of this doctoral thesis confirmed the presence of pharmaceuticals in wastewaters, surface waters, drinking waters, and fish tissue. Furthermore, the impact of wastewater on the continuous release of pharmaceuticals into the environment was confirmed. Therefore, we developed a treatment process that would effectively degrade pharmaceuticals in wastewaters. Firstly, the electrochemical degradation efficiency of the investigated compounds by boron-doped diamond electrode material was tested under laboratory conditions. The material in situ produced free radicals which effectively reacted with organic matter and provoked degradation. The treatment in an electrolytic cell with a boron-doped diamond electrode degraded more than 85% of the initial concentration for 7 out of 10 target compounds however, slightly lower degradation efficiency in real wastewaters was observed. The latter fact is tightly correlated with a shorter treatment time in order to demonstrate the sewage treatment process under real conditions and with higher amount of organic material, which also react with free radicals. Furthermore, the developed electrolytic cell was afterwards used as an additional treatment step in a hospital wastewater treatment plant within European project LIFE. Some extra optimisation was needed, such as a higher number of electrolytic cells, a higher current density, and the use of extra cleaning filter. Finally, the comparison between biological and electrochemical treatment revealed a superiority of the latter approach based on 53% overall improvement in terms of pharmaceuticals degradation. The entire treatment process of the biological treatment plant with an advanced oxidation process successfully degraded more than 86% of the initial concentration of each analyte. To summarize, the doctoral thesis highlighted the presence of pharmaceutical residue in the environment. To the best of our knowledge, this was the first demonstration of a simultaneous analysis of over 100 analytes using a QqQ mass analyser. The developed methods will greatly facilitate the detection of numerous residues of pharmaceuticals in environmental samples. Our observation of wastewaters, surface waters, drinking waters, and fish tissue represents the first research with comprehensive data for the Slovene region. Based on the found concentrations in environmental waters, with the emphasis on drinking water and fish tissue, we can estimate the environmental burden and risk for humans. We can conclude that the risk for humans is minimal and that it is somewhat higher for organisms that are exposed to wastewater discharges throughout their live cycles. Finally, the research resulted in the development of a highly efficient process for the degradation of pharmaceuticals. We can say that this new technology could be a viable and feasible option for upgrading existing wastewater treatment plants in order to achieve a significantly greater cleaning efficiency, and for lessening the ecological burden. The presented data can be the basis for further introduction of routine monitoring and might represent the preposition for an extension to the Watch List from Water Framework Directive (established in Directive 2008/105/EC of the European Parliament and of the Council of Europe).

Graphical abstract, Highlights • Design improvement of pinned disc rotational generator of hydrodynamic cavitation. • Evaluation of cavitation intensity by flow visualization and pressure measurements. • Rotor speed and pin spacing are most decisive parameters for cavitation efficiency. • Pin diameter and surface roughness are less significant design parameters. • Under best conditions COD was reduced by 31% in 15 liquid passes., In this study, the hydrodynamic cavitation and wastewater treatment performance of a rotary generator with pin disk for hydrodynamic cavitation are investigated. Various geometrical features and arrangements of rotor and stator pins were evaluated to improve the configuration of the cavitation device. The pilot device used to perform the experiments was upgraded with a transparent cover that allows visualization of the hydrodynamic cavitation in the rotor-stator region with high-speed camera and simultaneous measurement of pressure fluctuations. Based on the hydrodynamic characteristics, three arrangements were selected and evaluated with respect to the chemical effects of cavitation on a 200-liter wastewater influent sample. The experimental results show that the rotational speed and the spacing of the rotor pins have the most significant effect on the cavitation intensity and effectiveness, while the pin diameter and the surface roughness are less significant design parameters. Cavitation intensity increases with pin velocity, but can be inhibited if the pins are arranged too close together. At best configuration, COD was reduced by 31% in 15 liquid passes, consuming 8.2 kWh/kg COD. The number of liquid passes also proved to be an important process parameter for improving the energy efficiency.

Benzotriazoli predstavljajo eno izmed skupin novih onesnaževal. Polarne spojine iz te skupine zaradi protikorozivnih lastnosti uporabljamo kot dodatke k različnim industrijskim tekočinam, pa tudi k detergentom za strojno pomivanje posode, ki jih uporabljamo v gospodinjstvu. Njihov glavni vir vnosa v okolje je odpadna voda iz čistilnih naprav, kjer se nepopolno odstranijo. Relativno stabilni so tudi v okolju, ker pa so vodotopni, nazadnje pritečejo iz naših pip. V okolju se nahajajo v koncentracijah reda velikosti od ng/L do μg/L. Toksični so predvsem za vodne organizme, predstavljajo pa tudi potencialne karcinogene spojine za človeka. Razvijanje terciarnih metod za njihovo odstranjevanje je zato zelo pomembno. Ena od metod za obdelavo odpadnih vod je elektrooksidacija (EO), ki pa je za razliko od te študije v literaturi preizkušena predvsem v kombinaciji z drugimi tehnikami. Spremljala sem razgradnjo 1H-benzotriazola (BTA) v vzorcih sintetičnih odpadnih vod z EO. Pomemben del priprave tovrstnih vzorcev je ekstrakcija na trdno fazo (SPE), ki sem jo tudi sama uporabljala. Prav zato sem raziskala tudi, kako na izkoristek ekstrakcije BTA iz vzorcev vplivajo različni parametri. pH sam po sebi ni vplival na izkoristek ekstrakcije, vendar pa so se razlike med izkoristki ekstrakcij pri različnih pH vrednostih pokazale ob prisotnosti pomožnih elektrolitov. Preizkusili smo štiri različne elektrolite: NaCl, H2SO4, Na2SO4 in Na2CO3. Vrsta pomožnega elektrolita je pomembno vplivala na izkoristek ekstrakcije, ki je segal od 59 % pri Na2SO4 do 86 % pri Na2CO3. Ugotovila sem še, da se izkoristki ekstrakcij znatno znižajo pri koncentracijah BTA, višjih od 20 mg/L. Pri monitoringu razgradnje BTA sem opazovala vpliv anodnega materiala, pomožnega elektrolita in gostote toka na EO ter primerjala hitrost razgradnje pri visokih in nizkih koncentracijah BTA. Razgradnja je bila hitrejša pri višjih gostotah toka in nižjih koncentracijah BTA. Med anodnimi materiali smo preizkusili elektrodo iz mešanih kovinskih oksidov (MMO) in iz z borom dopiranega diamanta (BDD), pri čemer je bila razgradnja z BDD anodo hitrejša in je povzročila manj pretvorbenih produktov. Razgradnje so bile pri vseh pomožnih elektrolitih psevdo prvega reda. Produkte razgradnje sem spremljala z LC-MS/MS in jih opazila pri uporabi NaCl, ki je povzročil nastanek toksičnih kloriranih organskih spojin, ter H2SO4, ki je povzročila nastanek različnih hidroksiliranih benzotriazolov. Razgradnja je bila najhitrejša z uporabo H2SO4 – po 2,5 h se je od približno 1 mM začetne koncentracije razgradilo 97 % BTA. Benzotriazoles are a group of emerging organic pollutants. Due to their anticorrosive properties, the polar ones are used as additives in various industrial liquids, as well as in dishwashing detergents that we use in our households. Their main source of entry into the environment is wastewater from wastewater treatment plants, where they are insufficiently removed. They are also relatively stable in the environment and in the end, due to their water solubility, travel to our faucets. In the environment, they are found in concentrations ranging from ng/L to μg/L. They are toxic for aquatic organisms but are also suspected human carcinogens. Development of tertiary methods for their removal is hence of great importance. One of the methods of wastewater treatment is electrooxidation (EO) which is, in contrast to the present study, mainly tested in combination with other techniques. I monitored the degradation of 1H-benzotriazole (BTA) in synthetic wastewater samples with EO. An important part of analysis is sample preparation by solid-phase extraction (SPE) which I also used within my work. Hence, I also researched the influence of different parameters on BTA recovery via SPE. pH value of samples did not directly influence the recoveries, however, differences among recoveries at different pH values arose when supporting electrolytes were present. We tested four different electrolytes: NaCl, H2SO4, Na2SO4 and Na2CO3. The type of electrolyte had a great influence on SPE recoveries which ranged from 59% with Na2SO4 to 86% with Na2CO3. I also found that SPE recoveries lowered notably at BTA concentrations higher than 20 mg/L. During BTA degradation monitoring, I observed the influence of anodic material, supporting electrolyte and current density on EO and compared the rate of degradation at high and low BTA concentrations. The rate of degradation was higher at higher current densities and lower BTA concentrations. Among anodic materials we tested mixed metal oxide (MMO) and boron-doped diamond (BDD) electrodes and found that the rate of degradation was higher and accompanied by fewer transformation products at BDD electrode. Degradations with all tested electrolytes gave pseudo-first order kinetics. Transformation products were monitored by LC-MS/MS and were noticed mainly with the use of NaCl, which caused the emergence of toxic chlorinated organic compounds, and with the use of H2SO4, which caused the emergence of various hydroxylated benzotriazoles. The degradation rate was the highest when H2SO4 was used – after 2.5 h, 97% BTA was degraded from approximately 1 mM initial concentration.

Amfifilne snovi oziroma surfaktanti so kemijske snovi, ki znižajo medfazno površinsko napetost. Njihove molekule imajo dvojno naravo, saj je en del molekule hidrofilen (hidrofilna glava), drugi pa lipofilen (hidrofoben rep). Surfaktanti so široko uporabni v različnih tehnoloških procesih, v visokih koncentracijah pa so prisotni tudi v različnih čistilih v splošni uporabi v gospodinjstvih. Zaradi tega se pojavljajo tudi v tehnoloških odpadnih vodah in odplakah, kjer predstavljajo bolj ali manj motečo komponento za vodne ekosisteme. V diplomski nalogi smo se osredotočili na vlogo surfaktantov v odpadnih vodah. Proučili bomo kakšni so lahko njihovi vplivi na okolje, posebno pozornost pa smo namenil različnim postopkom, ki jih uporabljamo za odstranjevanje takšnih snovi iz odpadnih vod. Amphiphilic compounds or surfactants are chemical compounds that lower the interfacial surface tension. Their molecules contain both hydrophilic groups (hydrophilic head) and the lipophilic molecular part (hydrophobic tail). Surfactants are used in many technological processes and are also found in high concentrations in many household products such as detergents and other cleaning agents. Therefore, they are also present in wastewaters and can represent the environmental risks to aquatic ecosystems. In my diploma thesis, I will focus on the effects of surfactants to the wastewater. In doing so, I will pay attention to their effects on the environment and especially the processes we use to remove them from wastewater.

Sidestream partial nitritation and deammonification (pN/A) of high-strength ammonia wastewater is a well- established technology. Its expansion to the mainstream is, however mainly impeded by poor retention of anaerobic ammonia oxidizing bacteria (AnAOB), insufficient repression of nitrite oxidizing bacteria (NOB) and difficult control of soluble chemical oxygen demand and nitrite levels. At the municipal wastewater treatment plant in Strass (Austria) the microbial consortium was exhaustively monitored at full-scale over one and a half year with regular transfer of sidestream DEMON® biomass and further retention and enrichment of granular anammox biomass via hydrocyclone operation. Routine process parameters were surveyed and the response and evolution of the microbiota was followed by molecular tools, ex-situ activity tests and further, AnAOB quantification through particle tracking and heme measurement. After eight months of operation, the first anaerobic, simultaneous depletion of ammonia and nitrite was observed ex-situ, together with a direction to higher nitrite generation (68% of total NOx-N) as compared to nitrate under aerobic conditions. Our dissolved oxygen (DO) scheme allowed for transient anoxic conditions and had a strong influence on nitrite levels and the NOB community, where Nitrobacter eventually dominated Nitrospira. The establishment of a minor but stable AnAOB biomass was accompanied by the rise of Chloroflexi and distinct emergence of Chlorobi, a trend not seen in the sidestream system. Interestingly, the most pronounced switch in the microbial community and noticeable NOB repression occurred during unfavorable conditions, i.e. the cold winter season and high organic load. Further abatement of NOB was achieved through bioaugmentation of aerobic ammonia oxidizing bacteria (AerAOB) from the sidestream-DEMON® tank. Performance of the sidestream pN/A was not impaired by this operational scheme and the average volumetric nitrogen removal rate of the mainstream even doubled in the second half of the monitoring campaign. We conclude that a combination of both, regular sidestream-DEMON® biomass transfer and granular SRT increase via hydrocyclone operation was crucial for AnAOB establishment within the mainstream.

Graphical abstract, Highlights • Performance evaluation of pinned disc hydrodynamic cavitator. • Improved cavitation performance compared to the serrated disc. • High cavitation intensity and relatively low energy consumption. • Confirmed oxidation potential and particle size reduction., This study investigates hydrodynamic performance of a novel pinned disc rotating generator of hydrodynamic cavitation in comparison with a serrated disc variant on a pilot-scale. Experimental results show that at a given rotational speed and liquid flow rate, the pinned disc generates more intense cavitation (i.e. lower cavitation number, higher volume fraction of vapor and higher amplitude of pressure fluctuations) than the serrated disc, while also consuming less energy per liquid pass (i.e., higher flow rate and pumping pressure difference of water at similar power consumption). Additionally, mechanical and chemical wastewater treatment performance of the novel cavitator was evaluated on an 800 L influent sample from a wastewater treatment plant. Mechanical effects resulted in a reduction of average particle size from 148 to 38 µm and increase of specific surface area, while the oxidation potential was confirmed by reduction of COD, TOC, and BOD up to 27, 23 and 30% in 60 cavitation passes, respectively. At optimal operating conditions and 30 cavitation passes, pinned disc cavitator had a 310% higher COD removal capacity while consuming 65% less energy per kg of COD removed than the serrated disc cavitator. Furthermore, the specific COD-reduction energy consumption of the pinned disc cavitator on the pilot scale is comparable to the best cases of lab-scale orifice and venturi devices operating at much lower wastewater processing capacity.

Ne samo izločene zdravilne učinkovine, temveč tudi njihovi metaboliti lahko predstavljajo tveganje za okolje, saj lahko s svojo ohranjeno farmakološko aktivnostjo negativno vplivajo na zdravje ljudi in drugih izpostavljenih organizmov. Zato je bil cilj magistrske naloge razviti hitro in visoko občutljivo metodo za merjenje vsebnosti izbranih metabolitov zdravilnih učinkovin, pogosto prisotnih v slovenskih odpadnih vodah. Vzorce odpadnih vod smo pripravili z metodo ekstrakcije na trdnem nosilcu s pol-avtomatskim sistemom SPE-DEX in jih analizirali z LC-MS/MS. Koncentracije analitov smo izračunali s pomočjo metode standardnega dodatka, s katero smo hkrati odpravili vpliv ozadja na točnost kvantifikacije. Metodo smo uspešno optimizirali in ovrednotili glede točnosti, ponovljivosti, delovnega območja, meje določitve, izkoristka in vpliva ozadja. Potrdili smo njeno zanesljivost in visoko občutljivost ob sočasni analizi 26 analitov (11 zdravilnih učinkovin iz več terapevtskih skupin in 15 njihovih metabolitov), ki smo jih izbrali na podlagi pregleda literature in predhodnih analiz. Metodo smo uporabili na dvanajstih realnih vzorcih odpadnih vod (treh vtočnih in devetih iztočnih), vzorčenih v devetih komunalnih čistilnih napravah iz več slovenskih regij. Od skupno 26 analitov smo jih 12 zaznali v vseh vzorcih. Če upoštevamo samo metabolite, smo jih 60 % zaznali v vseh vzorcih, 80 % pa vsaj v enem vzorcu. V največjih povprečnih koncentracijah smo v vtočnih in iztočnih vzorcih izmerili kofein (4,1 in 4,6 µg/L), 1,7-dimetilksantin (3,6 in 2,7 µg/L) in salicilno kislino (1,1 in 1,7 µg/L). Karbamazepin smo izmerili v vseh vzorcih, in sicer v povprečni koncentraciji 165,9 ng/L v vtočnih in 142,2 ng/L v iztočnih vzorcih. Diazepama, fluoksetina, norfluoksetina, paliperidona in endoksifena nismo zaznali v nobenem vzorcu. Na splošno so bile izmerjene koncentracije manjše ali primerljive z navedenimi v podobnih študijah po svetu. Za čistilne naprave, iz katerih smo dobili tako vtočni kot iztočni vzorec, smo ocenili učinkovitost odstranjevanja analitov. Koncentracije v iztočnih vzorcih so se v primerjavi z vtočnimi zmanjšale, razen pri diklofenaku in karbamazepinu, za katera smo ugotovili slabo ali celo negativno učinkovitost odstranjevanja. Predvidevamo, da izmerjene koncentracije v iztočnih vzorcih ne predstavljajo neposrednega tveganja za človeka, vseeno pa moramo v bodoče ekotoksikološke študije vključiti tudi metabolite, zlasti tiste z ohranjeno farmakološko aktivnostjo, zaradi njihovega vpliva na vodne organizme. Not only eliminated pharmaceuticals, but also their metabolites may pose a risk to the environment because of their preserved pharmacological activity which may adversely affect the human health and other exposed organisms. Therefore, the aim of this Master's thesis was to develop a rapid and highly sensitive method for quantification of drugs' metabolites frequently found in Slovenian wastewaters. Wastewater samples were prepared with solid-phase extraction method using a semi-automated system SPE-DEX and analysed by LC-MS/MS. Analyte concentrations were calculated using the standard addition method, which also eliminated the influence of matrix effect on the accuracy of quantification. The method was successfully optimised and validated in terms of accuracy, precision, working range, limit of quantification, recovery and matrix effect. We confirmed its reliability and high sensitivity for the simultaneous analysis of 26 analytes (11 pharmaceuticals from multiple therapeutic groups and 15 of their metabolites), which were selected based on a literature review and preliminary analyses. The method was applied to twelve real wastewater samples (three influent and nine effluent) that were collected from nine municipal wastewater treatment plants from several Slovenian regions. Out of the total 26 analytes, 12 were detected in all samples. Considering only metabolites, 60 % were detected in all samples and 80 % in at least one sample. The highest average concentrations in influent and effluent samples were measured for caffeine (4.1 and 4.6 µg/L), 1,7-dimethylxanthine (3.6 and 2.7 µg/L) and salicylic acid (1.1 and 1.7 µg/L), respectively. Carbamazepine was measured in all samples in average concentrations of 165.9 ng/L in influent and 142.2 ng/L in effluent samples, respectively. Diazepam, fluoxetine, norfluoxetine, paliperidone and endoxifen were not detected in any sample. In general, the measured concentrations were lower or comparable to those reported in similar studies worldwide. For the wastewater treatment plants from which both influent and effluent samples were collected, the removal efficiencies of the analytes were evaluated. Concentrations in effluents decreased compared to influents, except for diclofenac and carbamazepine, which were found to have low or even negative removal efficiencies. In conclusion, the measured concentrations in effluents presumably do not pose a direct risk to humans. However, the metabolites, especially those with preserved pharmacological activity, should be included in future ecotoxicological studies due to their impact on aquatic organisms.

V magistrskem delu, smo obravnavali problematiko obremenjenosti realnih odpadnih tekstilnih vod z adsorbljivimi organskimi halogeni (AOX). V prvi fazi raziskav smo na podlagi seznama pomožnih sredstev in barvil, katera se uporabljajo v procesu barvanja, poiskali pripadajoče kemijske strukture spojin in zbrali podatke o recepturah, katere se bile uporabljene takrat, kadar so bile vrednosti AOX presežene. Glede na podane recepture, smo poiskali pripadajoče kemijske strukture uporabljenih spojin in njihove lastnosti. Osredotočili smo se na tiste spojine, katere imajo v svoji strukturi vezane halogene elemente, predvsem klor in brom, katera v pralni vodi tvorita halogenirane organske spojine ter v največji meri pripomoreta k tvorjenju AOX. Pri eksperimentalnem delu magistrske naloge, smo od podjetja pridobili 20 vzorcev odpadne obarvane vode in fotometrično določili vrednost adsorbljivih organskih halogenov v odpadni tekstilni vodi, t.j. po procesu barvanja. Pred tem, smo test preverili s standardno raztopino 4-klorobenzaldehida. Tako smo sestavili seznam barvil in pomožnih sredstev za katera sklepamo, da pripomorejo k povečanim vrednostim AOX in bi jih bilo smotrno zamenjati z ekološko primernejšimi substituenti. V okviru analize smo izvedli še hitri test za določanje kloridnih ionov. Master thesis presents the problem of the load of real textile wastewater with adsorbable organic halogens (AOX). In the first phase of the research, on the basis of the list of auxiliaries and dyes used in the coloring process, we found the corresponding chemical structures of the compounds and collected data on the recipes that were used when the AOX values were exceeded. Based on the given recipes we searched for the corresponding chemical structures of the compounds used and their properties. We focused on those compounds that have halogen-based elements in their structure, especially chlorine and bromine, which in the washing water form halogenated organic compounds and largely contribute to the formation of AOX. In the experimental part of the master's thesis we obtained 20 samples of waste water from the company and photometrically determined the value of adsorbable organic halogens in textile wastewater, i.e. after the dyeing process. Prior to this the test was checked with a standard solution of 4-chlorobenzaldehyde. Thus, we have drawn up a list of colorants and auxiliary agents that we believe are contributing to the increased AOX values and it would be sensible to replace them with ecologically more suitable substituents. In the framework of the analysis fast test for the determination of chloride ions was also carried out.

Vzpostavitev obtoka hranil in iskanje alternativnih virov fosforja je bistveno za ohranjanje visoke produktivnosti kmetijstva in prehranske varnosti v prihodnosti, ko bo človeštvo izčrpalo lahko dostopne zaloge fosforne rude. Fosfor, ki ga vsebujejo človeški iztrebki se trenutno v veliki večini zavrže kot odpadek. Človeške fekalije so lahko, kot so že bile v preteklosti, pomemben vir fosforja in ostalih hranil v rastlinski pridelavi. S poskusom vermikompostiranja črne vode smo hoteli z deževniki pospešiti mineralizacijo hranil in povečati dostopnost fosforja rastlinam v substratu iz zrelega komposta in žaganja iglavcev, ob ustrezni higienizaciji za nadaljnjo varno uporabo v kmetijstvu. V poskusu smo ugotovili, da deževniki pospešijo mineralizacijo hranil v substratu, vendar je za pridelavo kakovostnega vermikomposta potrebna daljša obdelava kot 21 dni oz. večja koncentracija deževnikov v substratu. Z dodajanjem črne vode se je v 21 dneh vermikompostiranja povečala koncentracija dostopnega fosforja za 11 – 24 %, medtem ko nismo dosegli zadostnega zmanjšanja števila patogenov mikroorganizmov in predelan substrat ni ustrezal standardom za uporabo v rastlinski pridelavi. Ob izvedbi preliminarnega poskusa smo ugotovili, da je foto dokumentiranje poskusa v obliki videa formata »time lapse«, dobra metoda predstavitve poskusa, ki nam je podala vpogled na potek procesa vermikompostiranja. Establishing a nutrient flow and finding alternative sources of phosphorus is essential to maintaining high agricultural productivity and food security in the future, when humanity could deplete readily available phosphorus ore reserves. Phosphorus and other nutrients contained in human excrements are currently being largely discarded as waste although they can be a good source for crop production. By 21-day trial of vermicomposting of black water mixed into a matrix of compost and sawdust, we found that earthworms accelerate the mineralization of nutrients, but for the production of quality vermicompost longer processing than 21 days or higher concentration of earthworms in the substrate is required. Addition of black water increases the concentration of available phosphorus in the finished vermikompost. Earthworms increase the concentration of available phosphorus by an average of 11-24 %. Vermicomposting within 21 days does not lead to sufficient reduction of pathogens to meet the standards for use in crop production. In carrying out the preliminary experiment, we found that photo documentation of the experiment in the form of time lapse video is a good method of presenting the experiment, which gave us an insight into the progress of the vermicomposting process.

Voda je vir življenja in nosilec vseh bioloških procesov, ki se odvijajo na našem planetu. Nekatera škodljiva sredstva povzročajo prekomerno razraščanje alg, ki onemogočajo svetlobi, da prodre v vodo. Barvila v vodi (predvsem temna) onemogočajo prodor svetlobe v vodo, kar povzroči, da koristne bakterije ne morejo opravljati svojega dela – čiščenja odpadne vode. Skoraj vsa sintetična barvila se razgrajujejo zelo počasi, kar je ekološko nezdravo in okoljsko nevarno. V diplomskem delu smo z adsorpcijo na zeolit ter granulirano oglje prikazali razbarvanje vod. Obarvanost se lahko pojavi na odpadnih vodah, ki pridejo v čistilno napravo Rajnkovec, Rogaška Slatina. Z adsorpcijo smo želeli zmanjšati vrednosti parametrov, ki so lahko občasno tudi preseženi. Najučinkovitejše razbarvanje smo dosegli po adsorpciji na granulirano aktivno oglje. V diplomsko nalogo pa smo vključili tudi shematski prikaz alarmnega sistema. Če pride do vdora padavinskih voda, lahko z njim to preprečimo oziroma hitreje odpravimo. Water is the source of life and the carrier of all biological processes that take place on our planet. Some harmful agents cause excessive overgrowth of algaes, that are preventing light to penetrate the water. The dyes in water (especially dark ones) are preventing penetration of light to the water and consequently, benefitial bacterias can not do their work – clean waste water. In this diploma thesis we have displayed the discoloration of water by adsorbtion on zeolite and granulated chorcoal. Coloration can appear on the waste wathers, that come to waste water treatment plant Rajnkovec, Rogaška Slatina. With adsorption we wanted to decrease the values of parameters that could sometimes be exceeded. We have reached the most efficient discoloration after adsorption on granulated active charcoal. In this diploma thesis we have also included shematic display of alarm system. If rainwater intrusion occurs, we can use the system to prevent or eliminate it more quickly.

Antibiotike vedno več uporabljamo v humani medicini, veterini in kmetijstvu. Čiščenje antibiotičnih odpadnih voda je zelo zahtevno, saj te odpadne vode poleg antibiotikov vsebujejo tako različne anorganske kot organske spojine, kar vse skupaj pogosto zavira delovanje mikroorganizmov, ki delujejo v čistilnih napravah. Zaviralni učinek na mikroorganizme v čistilnih napravah je še posbej izrazit pri konvencionalnih čistilnih napravah, ki uporabljajo procese aktivnega blata, biološke filtre in membranske biorektorje za čiščenje odpadnih voda. Posledično narašča onesnaženje odpadnih in izcednih voda z antibiotiki in njihovimi ostanki, ki predstavljajo skupino mikroonesnažil. Koncentracije antibiotikov v odpadnih vodah so od nekaj ng/L do nekaj mg/L, v naravnih vodah so koncentracije zaradi redčenja manjše, od nekaj ng/L do nekaj 100 µg/L. Ker so antibiotiki na čistilnih napravah in v okolju nasploh slabo razgradljivi, ima njihovo kopičenje v vodi lahko resne okoljske posledice. Tudi majhne koncentracije vodijo v širjenje antibiotske rezistence, spodbujajo nastanek biofilma pri patogenih bakterijah in imajo lahko toksičen učinek na vodne organizme. Trenutni postopek kvantifikacije antibiotikov v vodnih sistemih zajema ekstrakcijo na trdni fazi, kateri sledi tekočinska kromatografija sklopljena s tandemsko masno spektrometrijo. Taka analiza je zelo primerna za detekcijo večine antibiotikov, tudi ko so ti prisotni v zelo majhnih koncentracijah. Zato je v prihodnosti nujno, da bolj natančno determiniramo kritične koncentracije antibiotikov, ter tako zavarujemo naravno okolje in človeško zdravje pred njihovimi negativnimi učinki. Antibiotics application is increasing in human medicine, veterinary medicine and agriculture. Treatment of antibiotic wastewater is very difficult since these wastewaters do not only contain antibiotics but also inorganic and organic compounds that all together can slow down the activity of microorganisms operating in wastewater treatment plants (WWTP). The inhibitory effect on micro-organisms in WWTPs is especially seen in conventional treatment plants that use active sludge process, biological filters or membrane biorectors for the treatment of wastewaters. Contamination of wastewaters and leachates with antibiotics and their residues, represents a group of emerging micro-pollutans. Concentrations of antibiotics range from ng/l to mg/L in wastewaters, to a few ng/L or 100 μg/L in natural waters due to dilution effect. Because antibiotics in the environment are as poorly degradable as in wastewater treatments, their accumulation in the water can have serious environmental consequences. Even low concentrations can cause dissemination of antibiotic resistance, biofilm formation in pathogenic bacteria and toxic effects on aquatic organisms. Quantification of antibiotics in aqueous systems is carried out by solid phase extraction followed by liquid chromatography and tandem mass spectrometry. This analytical system is quite efficient for detecting all major classes of antibiotics, even when they are present at very low concentrations. Therefore, it is necessary to take decisive steps in the future, aimed at the determination of critical concentrations of antibiotics, which will make it possible to maintain safe values that will not exert a negative impact on the natural environment and human health.

Zagotavljanje čiste in kvalitetne vode postaja problem sodobne družbe. Težke kovine so eno izmed najnevarnejših onesnaževal, saj niso biološko razgradljive, se kopičijo v naravi in toksično učinkujejo na žive organizme že pri zelo nizkih koncentracijah. V primerjavi z ostalimi metodami za odstranjevanje težkih kovin iz odpadnih vod, se je adsorpcija na različne adsorbente izkazala kot bolj ekonomična in učinkovita. Z razvojem nanotehnologije so bili magnetni nanodelci deležni pozornosti mnogih raziskav za iskanje najprimernejši adsorbentov težkih kovin. Namen diplomske naloge je bila sinteza stabilnih maghemitnih nanodelcev ter njihova funkcionalizacija s hitozanom po treh različnih metodah. Ugotoviti je bilo treba, kateri delci so najprimernejši za odstranjevanje Cr(VI) ionov iz raztopine in pri kateri pH vrednosti je adsorpcija najučinkovitejša. Maghemitne delce smo sintetizirali z metodo koprecipitacije in jih nato funkcionalizirali s hitozanom po treh različnih metodah (metoda kovalentne vezave, mikroemulzijska metoda in suspenzijska zamreževalna metoda). Delce smo dodali v raztopino z znano koncentracijo Cr(VI) ionov, da je potekla adsorpcija. Adsorpcijo smo izvedli pri treh različnih pH vrednostih (pH 3, 4 in 5) ter določili optimalno vrednost za adsorpcijo težkih kovin na posamezne magnetne nanodelce. Kvantitativno analizo Cr(VI) v raztopini po adsorpciji smo izvedli spektrofotometrijsko z 1,5- difenilkarbazidom. Po opravljeni analizi smo primerjali uspešnost adsorpcije in določili delce, ki so dosegli najvišje odstotke adsorpcije. Ugotovili smo, da so najprimernejši delci za odstranjevanje Cr(VI) iz vode maghemitni delci funkcionalizirani z mikroemulzijsko metodo. Izkazalo se je tudi, da so najboljšo adsorpcijo vsi delci dosegli pri vrednosti pH 3. Providing clean and quality water is one of the major challenges of the modern world. Heavy metals are one of the most dangerous contaminants due to their non- degradability and persistence in the environment. Heavy metals accumulate through the food chain, moreover, they can be toxic and harmful to living organisms, even in small amounts of exposure. Among numerous technologies that have been developed for the removal of heavy metals from wastewater, adsorption has been proved as one of the most economical and efficient methods. As nanotechnology is progressing rapidly, magnetic nanoparticles have been gaining increasing interest as adsorbents for heavy metals. The aim of the diploma thesis was preparation of stable maghemite nanoparticles and their functionalization with chitosan, using three different methods. The main objectives were determining which magnetic nanoparticles are the most suitable for removal of Cr(VI) and evaluating removal efficiency at different pH. Maghemite nanoparticles were synthesized by co-precipitation method and then functionalized with chitosan by three different methods (covalent binding method, microemulsion method and suspension cross-linking technique). Different nanoparticles were used as magnetic adsorbents for removal of Cr(VI) from aqueous solution. In order to determine the effect of pH on the process od adsorption, we conducted experiments at different pH value (pH 3, 4, and 5). For quantitative determination of Cr(VI) concentrations in aqueous solutions after adsorption process, we used spectrophotometric method with 1,5-diphenylcarbazide. After analysis we ascertained which magnetic nanoparticles have achieved efficient removal rates of Cr(VI). The study reveals that the maghemite nanoparticles functionalized with chitosan by microemulsion method, are most effective adsorbents for removal of Cr(VI) from aqueous solutions. All off the tested maghemite nanoparticles achieved higher removal rates with a pH value decrease.

Diplomsko delo se osredotoča na dejanski primer sanacije rastlinske čistilne naprave v kraju Motovilci. Predstavljen bo nastanek odpadnih vod, njihove značilnosti in čiščenje le-teh. Podrobno bom predstavil tudi delovanje rastlinske čistilne naprave v odvisnosti od toka vode skozi njih ter bolj podrobno horizontalni, vertikalni ter kombinirani sistem podpovršinskega toka vode, ki se nanaša na dejanski primer nadgradnje horizontalnega toka z vertikalnim na rastlinski čistilni napravi Motovilci. V sami sanaciji si bomo ogledali problematiko dosedanjega stanja in rešitve, vezane na kanalizacijsko omrežje, kot tudi samo rastlinsko čistilno napravo. S slikovnim gradivom bomo dodobra prikazali način gradnje grede z vertikalnim tokom, z rezultati prvih meritev pa izsledke sposobnosti sanirane rastlinske čistilne naprave. The diploma focuses on the factual case of remediating the constructed wetland for waste water treatment in Motovilci, Slovenia. The production of waste water, its characteristics and the procedure of purifying it, is the core focus in the presentation. The operation of the constructed wetland is presented independently of the flow of water passing through the treatment facility. The horizontal, vertical and combined subsurface flow system described, refers to the upgrade of the horizontal and vertical system flow of the constructed wetland in question. In the remediation process, the challenges in the current state and the solutions connected to the sewage system are examined along with the entire constructed wetland as a whole. Using visual material, we demonstrate how to construct a vertical flow bed. The results of the initial measurements, the findings of the performance, pre and post the sanation of the constructed wetland are to be presented in the final case study.

V bolnišnicah je higiena ključnega pomena, zato se v njihovih pralnicah dnevno opere več ton umazanega perila. Zaradi večjih količin dodanih pralnih sredstev, prisotnih organskih in anorganskih snovi ter drugih nečistoč, je treba odpadno vodo pred odvajanjem v vodotoke ali kanalizacijo, ustrezno očistiti. V diplomskem delu smo z adsorpcijo na zeolita ZAG–CWK in ZAG–4A–2–3 ter granulirano aktivno oglje, čistili odpadno vodo iz pralnice Splošne bolnišnice Celje. Z metodo adsorpcije smo želeli z adsorbenti zmanjšati vrednosti občasno preseženih parametrov (pH, električna prevodnost, motnost, KPK, vsebnost tenzidov). Vpliv vzorcev odpadnih vod pred in po adsorpciji smo raziskali tudi s pomočjo čebulnega testa. Najučinkovitejše očiščenje odpadne vode in posledično znižanje vrednosti parametrov smo dosegli po adsorpciji na granulirano aktivno oglje. In hospitals, hygiene is of key importance, therefore their laundry services launder several tons of dirty linen every day. Due to large amounts of added laundry detergents, presence of organic and inorganic substances and other impurities, the waste water must be properly treated before it is discharged into watercourses or sewage systems. In the diploma thesis, we treated waste water from the General Hospital Celje laundry service with adsorption on zeolites ZAG–CWK and ZAG–4A–2–3 and granular activated charcoal. By using adsorbents, we wanted to reduce values of occasionally exceeded parameters (pH, electrical conductivity, turbidity, COD, tensides content) with the adsorption method. Waste water samples before and after adsorption were also examined with an onion test. Most efficient waste water treatment and, consequently, the reduction of parameters was achieved with adsorption on granular activated charcoal.

Objekt raziskovanja te naloge je bila odpadna industrijska voda iz umetnega zbirnega jezera, ki se nahaja v urbanem okolju. Prebivalstvo v tem okolju zlasti v poletnih mesecih moti pogosto penjenje in motnost vode v jezeru. Z namenom preiskave mikrobnih združb tega jezera smo poleti 2013 odvzeli vzorce vod in sedimentov iz petih, geografsko različnih vzorčnih mest jezera. Vzorcem smo ob odvzemu preiskali tudi izbrane fizikalno-kemijske lastnosti. Na hranilnem agarju smo iz vzorcev osamili 71 morfološko različnih izolatov. Vse izolate smo opisali s fenotipskimi in molekularno-biološkimi pristopi. S preiskavo nukleotidnih zaporedij genov za 16S rRNA smo identificirali 20 različnih bakterijskih rodov iz 16 družin. Sekvenca gena za 16S rRNA enega izmed izolatov se je z najbolj podobno sekvenco referenčnih sevov ujemala v le 96,2 %, zato smo ta izolat v nadaljevanju natančno preiskali s fenotipskimi pristopi in analizo njegovega genomskega zaporedja. Na osnovi teh rezultatov smo izolat opisali in objavili kot novo bakterijsko vrsto Paenibacillus aquistagni 11T. Preiskovanje odpornosti proti klinično pomembnim antibiotikom pri izolatih iz družine Enterobacteriaceae in iz rodov Acinetobacter ter Pseudomonas je odkrilo odpornost proti ampicilinu, pri 14,3 % sevov iz družine Enterobacteriaceae pa tudi proti kloramfenikolu. Izolati, identificirani kot Acinetobacter sp., Pseudomonas otitidis in Bacillus sp., ki smo jih izolirali iz vzorčnega mesta kjer je bilo penjenje najbolj obilno, so tudi v pogojih laboratorijskega namnoževanja tvorili stabilno peno na površini kompleksnega gojišča. Po analizi fizikalno-kemijskih lastnosti vode smo ugotovili najvišjo koncentracijo kovin v vzorčnem mestu 5, zato smo izolatom tega vzorčnega mesta preiskali odpornost proti železu in manganu. Za vse izolate smo ugotovili rast ob prisotnosti 300 mg/L Fe v kompleksnem gojišču, za 73 % ob prisotnosti 900 mg/L Fe, en izolat pa je toleriral 1500 mg/L Fe. Odpornost proti 1500 mg/L Mn smo potrdili pri 37 % preiskanih sevih, pri koncentraciji 300 mg/L Mn pa je rastlo kar 91 % izolatov. S tehnologijo Ion Torrent smo na osnovi preiskovanja sekvenc sedmih variabilnih regij genov za 16S rRNA tudi brez predhodne osamitve izolatov preiskali sestavo mikrobne združbe iz istih petih vzorčnih mest jezera odpadne vode kot smo pridobili izolate. V vzorcih Voda 1, Voda 3 in Voda 4 so prevladovale operacijske taksonomske enote iz družine Prochlorococcacea, v vzorcu Voda 2 iz družine Sphingomonadaceae, v vzorcu voda 5 pa iz družine Gallionellacea. V vzorcu Sediment 1 so prevladovale enote OTE iz družine Rhodobacteraceae, v vzorcu Sediment 2 iz Sphingomonadaceae, v vzorcu Sediment 3 iz Desulfobacteraceae, v vzorcu Sediment 4 iz Pseudomonadaceae in v vzorcu Sediment 5 iz družine Nostocaceae. Primerjava med bakterijskimi družinami, identificiranimi po osamitvi izolatov, ter bakterijskimi družinami identificiranimi neposredno, je odkrila najboljše ujemanje med rezultati za vzorec Voda 2 (57,2 %), nekoliko slabše za vzorec Voda 1 (21,8 %), Voda 3 (20,0 %) in Voda 4 (14,5 %), najslabše pa za vzorec Voda 5 (1,2 %). Pri sedimentih je bilo najboljše ujemanje med bakterijskimi družinami, identificiranimi z obema metodološkima pristopoma, v vzorcu Sediment 4 (74,0 %) in Sediment 2 (57,4 %), slabše pri Sediment 3 (10,2 %) in Sediment 1 (14,0 %), najslabše ujemanje pa je bilo ponovno v sedimentu iz vzorčnega mesta 5 (2,4 %). Z analizo glavnih komponent PCA smo ugotavljali vpliv fizikalno-kemijskih lastnosti na prisotnost posameznih bakterijskih družin. V vzorcih vod smo ugotovili povezavo med pH-vrednostjo, vsebnostjo Mg, Na, Cu, Fe, Mn, Zn in skupino bakterijskih družin Gallionellaceae, Burkholderiaceae, Eubacteriaceae, Methylophilaceae in Ferrovaceae. V vzorcih sedimentov pa je bila prevladujoča povezava med vsebnostjo As, Cd, Co, Cu, Fe, Mn, Ni, Zn in skupino bakterijskih družin Nostocaceae, Ferrovaceae, Gallionellaceae, Acetobacteraceae in Acidobacteriaceae. The object of this work was the industrial wastewater from the artificial lake situated in the urban environment, where the population is concerned, especially during the summer, by frequent foaming and opacity of the lake water. With the purpose of investigating microbial community of the lake, we have taken water and sediment samples from five sampling points representing different geographical areas of the lake. The sampling was performed in summer of 2013. Selected physicochemical characteristics have been also analyzed. On nutrient agar, we have isolated 71 morphologically different isolates. We have described all the isolates with phenotypic and molecular-biological approaches. After analysis of 16S rRNA gene sequences, we have identified 20 different bacterial genera from 16 families. The 16S rRNA gene sequence of one isolate had only 96.2 % identity to its closest reference species, which is why we have further investigated this isolate with phenotypic approaches and by analysis of the entire genome sequence. Based on the results, the strain was described and published as a novel bacterial species Paenibacillus aquistagni 11T. By analyzing resistance to clinically important antibiotics among isolates from Enterobacteriaceae family, and genera Acinetobacter and Pseudomonas, we have uncovered resistance to ampicillin, with 14.3 % of the Enterobacteriaceae strains also being resistant to chloramphenicol. Isolates obtained from the most foam-prone sampling site and identified as Acinetobacter sp., Pseudomonas otitidis or Bacillus sp. have also created a stable foam on the surface of a complex medium under laboratory conditions. After the analysis of physico-chemical properties of water, we have concluded that the highest metal concentration is present in sampling site 5. The analysis identified the capability of isolates from this point for growing in medium with 300 mg/L Fe, for 73 % of them for growing in the presence of 900 mg/L Fe and for one of them even with the presence of 1500 mg/L Fe. Tolerance to 300 mg/L Mn was confirmed for 91 % of strains and to 1500 mg/L Mn for 37 % of tested strains. Using the Ion Torrent technology and analysis based on seven variable regions of 16S rRNA gene sequences, we have also directly analyzed the composition of microbial community from the same five sampling points of the lake as we performed the growing and isolation of strains. In water samples 1, 3 and 4 operational taxonomic units (OTU) from the Prochlorococcacea family represented the highest number, in sample 2 from Sphingomonadaceae and in sample 5 from Gallionellacea. In sediment 1, OTU from the Rhodobacteraceae had the highest number, in sediment 2 from Sphingomonadaceae, in sediment 3 from Desulfobacteraceae, in sediment 4 from Pseudomonadaceae, and in sample sediment 5 from Nostocaceae. A comparison of identified bacterial families by indirect approach and next generation sequencing technology showed the highest coverage for water sample 2 (57.2 %), a bit lower for water sample 1 (21.8 %), water 3 (20.0 %) and water 4 (14.5 %) and the lowest for water sample 5 (1.2 %). The same type of comparison showed the highest coverage for samples sediment 4 (74.0 %) and sediment 2 (57.4 %), substantially lower for samples sediment 3 (10.2 %) and sediment 1 (14.0 %) and the lowest as in the case of water samples for the samples from sampling point 5 (2.4 %). Using the analysis of principal components, we have analyzed in the structure of reduced space of three principal factors the influence of physicochemical properties on the presence of bacterial families in samples. We have detected an important link between the pH, the content of Mg, Na, Cu, Fe, Mn, Zn in the water and families Gallionellaceae, Burkholderiaceae, Eubacteriaceae, Methylophilaceae, Ferrovaceae. In sediments, the prevailing link was between As, Cd, Co, Cu, Fe, Mn, Ni, Zn, and families Nostocaceae, Ferrovaceae, Gallionellaceae, Acetobacteraceae, Acidobacteriaceae.

The use of rare earth elements is a growing trend in diverse industrial activities, leading to the need for eco-friendly approaches to their efficient recovery and reuse. The aim of this work is the development of an environmentally friendly and competitive technology for the recovery of those elements from wastewater. Kinetic and equilibria batch assays were performed with zeolite, with and without bacterial biofilm, to entrap rare earth ions from aqueous solution. Continuous assays were also performed in column setups. Over 90% removal of lanthanum and cerium was achieved using zeolite as sorbent, with and without biofilm, decreasing to 70% and 80%, respectively, when suspended Bacillus cereus was used. Desorption from the zeolite reached over 60%, regardless of the tested conditions. When in continuous flow in columns, the removal yield was similar for all of the rare earth elements tested. Lanthanum and cerium were the elements most easily removed by all tested sorbents when tested in single- or multi-solute solutions, in batch and column assays. Rare earth removal from wastewater in open setups is possible, as well as their recovery by desorption processes, allowing a continuous mode of operation., This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the research project PTDC/AAG-TEC/5269/2014, the strategic funding of UID/BIO/04469/2013 unit and BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte, Portugal., info:eu-repo/semantics/publishedVersion

V raziskavi smo preverjali, če je možno z anaerobno razgradnjo surovega blata z dodatkom sirotke z laktozo proizvesti več bioplina kot iz samega surovega blata. Surovo blato je mešanica primarnega in sekundarnega blata vzetega iz čistilne naprave Domžale - Kamnik. Želeli smo ugotoviti način doziranja in maksimalno količino dodane sirotke z laktozo v anaerobne bioreaktorje, da ostane proces stabilen in brez negativnega vpliva na strojno zgoščanje. Preverili smo tudi ali se ob povečani proizvodnji bioplina na račun sirotke z laktozo tudi poveča delež metana v bioplinu. Raziskavo smo izvedli z dvema pilotnima bioreaktorjema, katerih volumen je 197 L. Ugotovili smo, da je anaerobna razgradnja sirotke z laktozo potekala stabilno in brez večjih težav. V eksperimentalnem reaktorju se je povečala proizvodnja bioplina, in sicer v prvih petih obdobjih dodajanja po 1 L sirotke na dan za 172 % in v naslednjih petih obdobjih dodajanja po 2 L sirotke na dan za 200 %. S povečano proizvodnjo bioplina v eksperimentalnem reaktorju se ni povečal delež metana v bioplinu, ampak je ta celo upadel. Ob dodajanju 1 L sirotke na dan je delež metana v bioplinu upadel za 2 % in ob dodajanju 2 L sirotke na dan za 11 %. Bioplinska produktivnost je tako znašala v eksperimentalnem reaktorju ob dodajanju 1 L sirotke na dan 19 m3/m3/dan in ob dodajanju 2 L sirotke na dan 21 m3/m3/dan, medtem ko je v kontrolnem reaktorju znašala 14 m3/m3/dan. Rezultati so pokazali, da je bil na račun dodane sirotke izplen bioplina večji v eksperimentalnem reaktorju. Ugotovili smo, da se količina suhe snovi in delež organske snovi ne razlikujeta med bioreaktorjema in da sirotka nima vpliva na učinkovitost centrifugiranja dehidriranega blata. In the study, we examined if anaerobic degradation of raw sludge (RS) with addition of whey produces more biogas then anaerobic degradation of RS. The RS is a mixture of primary and secondary sludge taken from a wastewater treatment plant Domžale -¬ Kamnik. We wanted to find out the method of dosing and the maximum amount of added whey to the anaerobic bioreactors to keep the process stable and without adversely affecting machine densification. We checked whether the increased biogas production at the expense of whey increases the proportion of methane present in biogas. The study was carried out with two pilot bioreactors, the volume of which was 197 L. We found that anaerobic decomposition of the whey was stable. In the experimental reactor (R2), the production of biogas increased in the first five periods of adding 1 L/day of whey by 172 % and in the next five periods of adding 2 L/day of whey by 200 %. With the increased production of biogas in the R2, the proportion of methane in biogas did not increase, but it even declined. When adding 1 L/day of whey, the methane content decreased by 2 % and added 2 L/day of whey it decreased by 11 %. Biogas productivity was thus found to be in the R2 while adding 1 L/day of whey 19 m3/m3/day and adding 2 L/day of whey 21 m3/m3/day while 14 m3/m3/day was produced in the control reactor. The results showed that the extraction of the biogas was higher in the R2 where we used whey. It has been found that the amount of dry matter and the proportion of organic matter does not differ between bioreactors and that whey has no influence on the efficiency of centrifugation of dehydrated sludge.

V Sloveniji je več kot 70 % odpadne vode iz kanalizacijskih sistemov prečiščene, kljub temu pa je skoraj vsa izpuščena v naravo neuporabljena. V letu 2017 so Združeni narodi posebno pozornost namenili ravno ponovni uporabi odpadne vode, na ravni Evropske unije pa so v zadnjih leti pripravili več smernic in predlog za ponovno uporabo odpadne vode. V Sloveniji je zakonodaja o upravljanju z odpadno vodo v smislu ponovne uporabe žal še v povojih. Zavedanje, da so naravne dobrine končne, nas spodbuja k usmeritvi k trajnostnemu razvoju in krožnemu gospodarstvu. V Sloveniji spada krožno gospodarstvo med strateške razvojne prioritete, katerih osrednji cilj je zagotavljanje kakovosti življenja za vse. Ponovna uporaba odpadne vode tako predstavlja pomemben element naše sedanjosti in prihodnosti, tako iz gospodarskega kakor naravovarstvenega vidika. V diplomski nalogi bom teoretično predstavila izgradnjo sistema ponovne uporabe prečiščene odpadne vode na primeru Centralne čistilne naprave Šentjur. Namen naloge je prikazati smiselnost izgradnje takšnega sistema iz vidika okoljevarstva, varčevanja z naravnimi viri in stroškovnega vidika. Na podlagi izračunov se naložba povrne v devetnajstih mesecih, hkrati pa prispevamo k manjši obremenjenosti naravnih virov vode. In Slovenia, more than 70% of wastewater from sewerage systems is cleaned at wastewater treatment plants, but almost all of it is left further unused. In 2017, the United Nations paid special attention to the re-use of wastewater and several guidelines and proposals for the re-use of wastewater have also been drawn up at European Union level in recent years. In Slovenia, the legislation on wastewater management in terms of re-use is still in its infancy. The awareness that natural resources are finite, encourages us to move towards sustainable development and circular economy. In Slovenia, the circular economy is one of the strategic development priorities whose main goal is to ensure the quality of life for all. The reuse of wastewater thus represents an important element of our present and future, both from an economic and environmental point of view. In the following text we will theoretically present the construction of a system for the re-use of purified waste water on the example of the Central Wastewater Treatment Plant Šentjur. The purpose is to demonstrate the building of such system from the point of view of environmental protection, saving with natural resources and the cost perspective. On the basis of the calculations, the investment is reimbursed in the course of nineteen months, while at the same time we contribute to a lesser burden on natural water resources.

V skladu z določili splošne emisijske uredbe je v industrijski odpadni vodi potrebno določiti izbrane kemijske parametre in tudi strupenost za vodni organizem Daphnia magna, ki spada v skupino nižjih rakov. V primeru, kadar so v odpadni vodi prisotni herbicidi, bi za realnejšo oceno okoljskega tveganja bilo primerneje testirati strupenost na drugih vrstah organizmov, kot so zelene alge. V magistrski nalogi smo obravnavali učinek različnih odpadnih vod in herbicida terbutilazina na vodni ekosistem. Naš namen je bil raziskati in utemeljiti uvedbo testa strupenosti z zeleno algo Pseudokirchneriella subcapitata v slovensko zakonodajo. Teste strupenosti z vodno bolho smo izvedli v skladu s standardno metodo SIST EN ISO 6341:2013, test zaviranja rasti sladkovodnih alg pa s standardno metodo SIST EN ISO 8692:2012. Testiranim vzorcem smo s standardizirano metodo določili EC50 v primeru testa z vodno bolho Daphnia magna in ErC50 v primeru testa z zeleno algo Pseudokirchneriella subcapitata. Ugotovili smo, da odpadne vode različno učinkujejo na mobilnost vodne bolhe in na rast zelenih alg. Herbicid terbutilazin tudi pri najvišjih koncentracijah ni izkazal strupenih učinkov na vodno bolho, na alge pa je deloval zaviralno pri vseh uporabljenih koncentracijah. In accordance with the provisions of the general emission regulation, it is necessary to determine concentrations of selected chemical parameters as well as the toxicity for the aquatic organism Daphnia magna (small planktonic crustacean, water flea) in industrial waste water. In the case where herbicides are present in wastewater, it would be more appropriate to test the toxicity of organisms from first trophic level such as green algae for a more realistic environmental risk assessment. In this research we discussed the effect of various waste waters and herbicide terbutylazin on the aquatic ecosystem. Our intention was to investigate and justify the introduction of the toxicity test with the green alga Pseudokirchneriella subcapitata into Slovenian legislation. Aquatic toxicity test with water flea was carried out in accordance with the standard method SIST EN ISO 6341: 2013, and the freshwater algae growth control inhibition test was carried out using the standard method SIST EN ISO 8692: 2012. In tests endpoint EC50 was determined in the case of Daphnia magna and ErC50 in the case of a green algae test Pseudokirchneriella subcapitata. We have determined that waste waters have a different effect on the water flea mobility and on the growth of green algae. Herbicide terbutilazin, even at the highest concentrations used, did not exhibit toxic effects on daphnia, whereas in the case of algae concentrations were inhibitory at all levels used in tests.

Cilj diplomskega dela je poiskati najprimernejši način odvajanja in čiščenja odpadnih vod na območjih z razpršeno poselitvijo. Opisali smo različne vzorce poselitve in statistiko poselitve države in občine Gornja Radgona. Nazorno je predstavljeno, da v celotni državi, vključno z občino Gornja Radgona, večji del prebivalstva živi na podeželju. Opisani bodo štirje načini odvajanja vod na podeželju, prav tako pa tudi primeri čiščenja z uporabo osrednje in male čistilne naprave. Podane bodo delitve čistilnih naprav po tipologiji čiščenja. V zadnjem poglavju bodo obravnavane rešitve odvajanja in čiščenja odpadnih vod na podeželju v primeru razpršene poselitve. The aim of the graduation thesis is to find the most suitable way of drainage and wastewater treatment in the areas of dispersed settlement. We described different settlement patterns and the statistics of the settlement in the country and the Municipality of Gornja Radgona. It is clearly presented that in the entire country, including the Municipality of Gornja Radgona, the larger part of the population lives in the countryside. Four ways of water drainage in the countryside will be described, as well as the examples of treatment with the use of a medium or small treatment plant. The division of treatment plants will be introduced according to the treatment typology. The solutions of the drainage and wastewater treatment in the countryside in the event of dispersed settlement will be discussed in the last chapter.

V raziskavi smo se osredotočili na biološko odstranjevanje fosforja na Centralni čistilni napravi Domžale-Kamnik (CČNDK). Ta je bila v letu 2016 nadgrajena s štirimi sekvenčnimi šaržnimi reaktorji (SBR), ki omogočajo tudi terciarno čiščenje odpadne vode. Namen raziskave je bil ugotoviti ali CČNDK po nadgradnji zadosti novim, strožjim mejnim vrednostim za izpust očiščene vode v odvodnike, ki vstopajo v veljavo z novo Uredbo o odvajanju in čiščenju komunalne odpadne vode (2015). Z ustreznim delovanjem reaktorjev SBR, naj bi CČNDK v prihodnjih letih zagotovila koncentracije skupnega fosforja v iztoku iz čistilne naprave nižje od 1 mg/L. V drugem delu raziskave smo se osredotočili na dokazovanje prisotnosti polifosfat akumulirajočih bakterij (PAO) v aktivnem blatu. V aktivnem blatu iz CČNDK smo prisotnost PAO na različnih stopnjah čiščenja dokazovali preko sledenja funkcionalnega genskega markerja ppk1, ki kodira encim polifosfat kinazo 1, ključno za sintezo polifosfata iz molekule ATP. Poleg tega smo z oligonukleotidnimi začetniki, ki nalegajo na specifično regijo gena za 16S rDNK posebej sledili tudi domnevno ključnega predstavnika PAO - še neizolirano bakterijsko vrsto Candidatus Accumulibacter phosphatis (CAP). S kemijskimi analizami odpadne vode smo ugotovili, da je CČNDK z nadgradnjo v letu 2016 zadostila novim, strožjim mejnim vrednostim. Ugotovili smo, da je bila v zagonski fazi mikrobiološka učinkovitost čiščenja fosforja iz odpadne vode 35 %. Z molekularnimi metodami smo dokazali prisotnost PAO v aktivnem blatu odvzem na različnih merilnih mestih. In this research we focused on biological phosphorus removal in Wastewater treatment plant Domzale-Kamnik (WWTPDK). In 2016 WWTPDK was upgraded with four sequence batch reactors (SBR), which enable tertiary treatment of wastewater. The aim of our research was to find out, if the upgrade of WWTPDK reduces the amounts of phosphorous in wastewaters below the values established in a new regulatory limits. With appropriate functioning, SBR reactors should reduce the total phosphorus concentrations in WWTPDK effluent below 1 mg/L. In second part of the research, we tried to detect phosphate accumulating organisms (PAO) in the active sludge. PAO presence in active sludge from different parts of WWTPDK was confirmed via the presence of functional genetic marker ppk1. The latter encodes the enzyme crucial for synthesis of polyphosphate from ATP - polyphosphate kinase 1. Specific 16S rDNA primers were also used to identify important yet unisolated representative of PAO Candidatus Accumulibacter phosphatis (CAP). Chemical analysis of wastewater proved that the upgraded WWTPDK can satisfactory reduce the amounts of phosphorous below the limiting values established in a new regulation. Microbiological phosphorus removal efficiency in wastewater was 35 %. We also detected PAO in the active sludge from different sampling points.

Water treatment at Seierstad drinking water treatment plant is done by chemical coagulation using aluminum-based coagulant. The quality of the drinking water source – Lake Farris, has decreased throughout the years, that is why more and more particles, organic matter and dissolved minerals must be removed. That contributes to the volume of sludge being produced at the end of the process. The main objective of the assignment is to reduce sludge volume by coagulation and flocculation in the sedimentation basins using a coagulant/flocculent. In order to choose the right coagulant tests have been performed in the laboratory and on the actual process. The laboratory analyses showed great potential and one of the coagulants has been chosen and tested on the actual process. The testing was followed up for the entire summer of 2016 and then compared to the year before. We have been following the pH and Al3+ content in the water being released back to the environment and visibility in the top water layer in the sedimentation basin. Cost analysis, including the cost of chemicals and electricity usage, has been done. The sludge reduction was successful, and it was approximately 49% less than the year before. Oskrba s pitno vodo je v sodobni družbi zmeraj pomembnejša dobrina. Pri procesu čiščenja pitne vode uporabljamo fizikalne, kemijske in biološke metode. Kakovost vodnega vira, iz katerega Vestfold Vann zajema vodo, se je v zadnjih letih poslabšala, zato je potrebno odstraniti vse več delcev, organskih snovi in mineralov. V podjetju Vestfold Vann (postaja Seierstad) čiščenje vode temelji na kemijskem čiščenju, pri katerem s pomočjo koagulacije, flokulacije in filtracije skozi dvo medijski filter, očistijo vodo pred izpustom v vodovodni sistem. Koagulant, ki je v uporabi je t. i. polialuminijev klorid, ter flokulant Magnafloc LT20. Koagulant in flokulant pripomoreta k združevanju delcev v večje kosme, za lažjo filtracijo. Filtraciji sledi izpiranje filtrov. Odpadne vode iz filtra so preusmerjene v proces sedimentacije. V sedimentacijskem bazenu se oblikujeta dva sloja. Vrhnji sloj vode je izpuščen nazaj v naravo, medtem ko usedlino črpajo naprej v čistilno napravo odpadnih voda (Lilevik). Namen naloge je izboljšati sedimentacijo nečistoč v sedimentacijskih bazenih s pomočjo koagulacije/flokulacije, brez da bi s tem vplivali na kvaliteto čiščenja pitne vode. Z zmanjšanjem odpadnih voda bi zmanjšali stroške črpanja usedlin v čistilno napravo za čiščenje odpadnih voda, ter vpliv aluminija na okolje. Za izbiro flokulanta smo se odločili s pomočjo laboratorijskih rezulatov, reakcije flokulanta na druge kemikalije, ki so uporabljene v procesu, ter stroškov nakupa le-tega. Testirali smo dva koagulanta in izbrali enega, s katerim smo izvajali teste na dejanskem procesu. Pred izvajanem samega eksperimenta smo vgradili nov dozirni sistem, ki nam je omogočal doziranje flokulanta pred sedimentacijskimi bazeni. Spremembam smo sledili z zajemanjem vzorcev. Merili smo pH vrednost in koncentracijo Al3+ v vodi, ki jo izpuščamo nazaj v okolje. Nadzorovali smo tudi vidljivost v zgornjem sloju vode. Vidljivost in koncentracija Al3+ sta si obratno sorazmerni. Črpanje samih usedlin je bilo nadzorovano s strani operatorjev na postaji. S pomočjo doziranja flokulanta, smo v letu 2016, v sedimentacijskem bazenu uspeli zmanjšati volumen usedlin za približno 49% v prmerjavi z letom 2015. Tudi stroški obratovanja so se kljub nakupu dodatnih kemikalji zmanjšali za približno 45%.