Oksidacijski stres u toksičnosti triazinskih pesticida - pregled glavnih biomarkera

This review article provides a summary of the studies relying on oxidative stress biomarkers (lipid peroxidation and antioxidant enzymes in particular) to investigate the effects of atrazine and terbuthylazine exposure in experimental animals and humans published since 2010. In general, experimental animals showed that atrazine and terbuthylazine exposure mostly affected their antioxidant defences and, to a lesser extent, lipid peroxidation, but the effects varied by the species, sex, age, herbicide concentration, and duration of exposure. Most of the studies involved aquatic organisms as useful and sensitive bio-indicators of environmental pollution and important part of the food chain. In laboratory mice and rats changes in oxidative stress markers were visible only with exposure to high doses of atrazine. Recently, our group reported that low-dose terbuthylazine could also induce oxidative stress in Wistar rats. It is evident that any experimental assessment of pesticide toxic effects should take into account a combination of several oxidative stress and antioxidant defence biomarkers in various tissues and cell compartments. The identified effects in experimental models should then be complemented and validated by epidemiological studies. This is important if we wish to understand the impact of pesticides on human health and to establish safe limits.Ovaj pregledni članak daje sažetak studija objavljenih od 2010. godine o djelovanju atrazina i terbutilazina na parametre oksidacijskog stresa (osobito lipidnu peroksidaciju i antioksidacijske enzime) u eksperimentalnih životinja i ljudi. Općenito je utvrđeno da izloženost eksperimentalnih životinja atrazinu i terbutilazinu najviše utječe na njihovu antioksidacijsku obranu i u manjoj mjeri na lipidnu peroksidaciju, ali učinci variraju ovisno o vrsti, spolu, dobi, koncentraciji herbicida i trajanju izlaganja. Većina studija uključivala je vodene organizme kao korisne i osjetljive bio-pokazatelje onečišćenja okoliša i važan dio prehrambenog lanca. U laboratorijskih miševa i štakora promjene u markerima oksidacijskog stresa bile su vidljive samo nakon izloženosti visokim dozama atrazina. U svojem smo nedavnom izvješću iznijeli podatak da niske doze terbutilazina također mogu inducirati oksidacijski stres u Wistar štakora. Očigledno je da pri svakoj eksperimentalnoj procjeni toksičnih učinaka pesticida treba uzeti u obzir kombinaciju nekoliko biomarkera oksidacijskog stresa i antioksidacijske obrane u različitim tkivima i stanicama. Učinci identificirani u eksperimentalnim modelima trebaju se nadopunjavati i potvrditi epidemiološkim istraživanjima. To je važno ako želimo razumjeti utjecaj pesticida na ljudsko zdravlje i uspostaviti sigurnu granicu izloženosti

Protektivni učinak mladog in vitro češnjaka kod citotoksičnosti inducirane vodikovim peroksidom u stanicama hepatocelularnog karcinoma

Garlic is a valuable source material for medicines due to its known antitumor, hypolipidaemic, antioxidant, and immunomodulatory effects. This study compares the protective effects of conventionally grown (CG) and in vitro propagated garlic (PG) against hydrogen peroxide-induced cytotoxicity in HepG2 cells and their antioxidant activity. Garlic used in this study was obtained by planting garlic cloves or by planting the transplants of PG directly in the field. At the end of the vegetation period, CG and PG were sampled and extracts prepared for the experiment. Compared to conventionally grown garlic bulbs, PG leafy part yielded significantly higher content of polyphenols, flavonoids and alliin, and also showed equal or higher antioxidant activity, measured by the cell viability test, GSH and ROS level. Moreover, PG can be produced in less time (shorter vegetation period) and with significantly less material (cloves). Significantly higher content of alliin, polyphenols, and flavonoids and significantly higher yield of plant biomass in PG has a great potential to become a new production model with improved garlic properties as a medicine material.Češnjak je vrijedan izvor ljekovitih sastojaka te je i dalje vrlo raširen, posebice zbog svojih antitumorskih, hipolipidemičkih, antioksidacijskih i imunomodulatornih svojstava. Ovo istraživanje pokazuje protektivni učinak te antioksidacijsko djelovanje konvencionalno uzgojenog i češnjaka uzgojenog iz in vitro presadnica. Češnjak korišten u ovom istraživanju dobiven je sadnjom češnjeva (CG) ili in vitro presadnica (PG) izravno u tlo. Nakon završetka vegetacije, CG i PG češnjak su uzorkovani te su pripremljeni ekstrakti kako bi se usporedilo njihovo protektivno djelovanje te antioksidacijska aktivnost. Utvrđeno je kako metoda uzgoja koja je rezultirala krajnjim prinosom lukovice kod CG ili nadzemnog dijela kod PG češnjaka značajno utječe na povećanje sadržaja polifenola, flavonoida, antioksidacijski učinak i na sadržaj alina. Također, ispitano je protektivno djelovanje ekstrakata češnjaka na prethodno induciranu citotoksičnost vodikovim peroksidom kod HepG2 stanica. Na temelju testa stanične vijabilnosti, sadržaja GSH i ROS-a utvrđeno je kako PG češnjak može biti jednako ili više učinkovit u svom ljekovitom i antioksidacijskom djelovanju u odnosu na CG češnjak. Nadalje, PG u usporedbi s CG češnjakom može se proizvesti u kraćem vegetacijskom razdoblju, u kojem se ostvaruje veći prinos korištenjem manje originalnog reprodukcijskog materijala (češnjeva). Na kraju, potrebno je istaknuti kako značajno veći sadržaj alina, polifenola i flavonoida, uz istovremeni značajno veći prinos biomase, utvrđen kod PG češnjaka, eventualno donosi novi uzgojni model češnjaka s povećanim sadržajem bioaktivnih komponenti, što poboljšava funkcionalnost češnjaka kao izvora ljekovitih sastojaka

Environmental Exposure to Metals, Parameters of Oxidative Stress in Blood and Prostate Cancer: Results from Two Cohorts

We studied the potential role of exposure to various metal(oid)s (As, Cd, Cr, Hg, Ni, and Pb) in prostate cancer. Two cohorts were established: the Croatian cohort, consisting of 62 cases and 30 controls, and the Serbian cohort, consisting of 41 cases and 61 controls. Blood/serum samples were collected. Levels of investigated metal(oid)s, various parameters of oxidative stress, and prostate-specific antigen (PSA) were determined in collected samples. A comparison of the measured parameters between 103 prostate cancer patients and 91 control men from both Croatian and Serbian cohorts showed significantly higher blood Hg, SOD, and GPx levels and significantly lower serum SH levels in prostate cancer patients than in controls. Correlation analyses revealed the significant relationship between certain parameters of oxidative stress and the concentrations of the measured metal(loid)s, pointing to the possible role of metal(oid)-induced oxidative stress imbalance. Furthermore, a significant inverse relationship was found between the blood Pb and the serum PSA in prostate cancer patients, but when the model was adjusted for the impacts of remaining parameters, no significant association between the serum PSA and the measured parameters was found. The results of the overall study indicate a substantial contribution of the measured metal(loid)s to the imbalance of the oxidant/antioxidant system. Although somewhat conflicting, the results of the present study point to the possible role of investigated metal(oid)s in prostate cancer, especially for Hg, since the obtained relationship was observed for both cohorts, followed by the disturbances in oxidative stress status, which were found to be correlated with Hg levels. Nevertheless, further studies in larger cohorts are warranted to explain and confirm the obtained results

Multi-elemental composition and antioxidant properties of strawberry tree (Arbutus unedo L.) honey from the coastal region of Croatia: Risk-benefit analysis

The concentration of 23 major and trace elements, total phenolic content (TPC) and 1, 1- diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity were determined in nine samples of strawberry tree honey and compared to other types of unifloral honeys. The most abundant elements in strawberry tree honey were potassium, calcium, magnesium and sodium, ranging between 1276 and 2367, 95.2-154, 14.4- 74.4 and 13.4-64.3 mg/kg, respectively. Strawberry tree honey had generally higher TPC (range: 0.314-0.522 g GA/kg) and DPPH (1.94- 4.45 mM TE/kg) compared to other analysed unifloral honeys. A strong positive relationship was found between TPC and DPPH, TPC and concentration of homogentisic acid (HGA), chemical marker of strawberry tree honey, and between DPPH and HGA. Regarding daily intake of essential elements, strawberry tree honey can be considered nutritionally richer than the majority of unifloral honeys available in Croatia, while contribution to tolerable intake set for potentially toxic elements was very low, corresponding to pristine areas

Oksidacijski stres, aktivnost kolinesteraza i primarna oštećenja u jetri, krvi i plazmi Wistar štakora nakon 28-dnevnog izlaganja glifosatu

In this 28 day-study, we evaluated the effects of herbicide glyphosate administered by gavage to Wistar rats at daily doses equivalent to 0.1 of the acceptable operator exposure level (AOEL), 0.5 of the consumer acceptable daily intake (ADI), 1.75 (corresponding to the chronic population-adjusted dose, cPAD), and 10 mg kg-1 body weight (bw) (corresponding to 100 times the AOEL). At the end of each treatment, the body and liver weights were measured and compared with their baseline values. DNA damage in leukocytes and liver tissue was estimated with the alkaline comet assay. Oxidative stress was evaluated using a battery of endpoints to establish lipid peroxidation via thiobarbituric reactive substances (TBARS) level, level of reactive oxygen species (ROS), glutathione (GSH) level, and the activity of glutathione peroxidase (GSH-Px). Total cholinesterase activity and the activities of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) were also measured. The exposed animals gained less weight than control. Treatment resulted in significantly higher primary DNA damage in the liver cells and leukocytes. Glyphosate exposure significantly lowered TBARS in the liver of the AOEL, ADI, and cPAD groups, and in plasma in the AOEL and cPAD group. AChE was inhibited with all treatments, but the AOEL and ADI groups significantly differed from control. Total ChE and plasma/liver ROS/GSH levels did not significantly differ from control, except for the 35 % decrease in ChE in the AOEL and ADI groups and a significant drop in liver GSH in the cPAD and 100xAOEL groups. AOEL and ADI blood GSH-Px activity dropped significantly, but in the liver it significantly increased in the ADI, cPAD, and 100xAOEL groups vs. control. All these findings show that even exposure to low glyphosate levels can have serious adverse effects and points to a need to change the approach to risk assessment of low-level chronic/sub-chronic glyphosate exposure, where oxidative stress is not necessarily related to the genetic damage and AChE inhibition.U okviru 28-dnevnog pokusa istražili smo učinke herbicida glifosata na modelu odraslih mužjaka Wistar štakora koji su oralno dobivali testirani spoj u subletalnim dnevnim dozama: 0,1 od prihvatljive razine izloženosti operatera (0,1xAOEL), 0,5 od prihvatljivog dnevnog unosa za potrošače (0,5xADI), 1,75 (odgovara kroničnoj populacijskoj prilagođenoj dozi, cPAD) i 10 mg kg-1 tjelesne težine na dan (odgovara 100xAOEL). Tijekom pokusa praćeni su sistemski toksični učinci. Nakon završetka svih tretmana svakoj je pokusnoj životinji izmjerena tjelesna težina i težina jetre te su uspoređene s polazišnim vrijednostima. Alkalnim komet-testom izmjerena je razina primarnih oštećenja DNA u leukocitima i jetrenim stanicama. Primjenom metoda za procjenu oksidacijskog stresa izmjerene su razine lipidne peroksidacije (TBARs), reaktivnih kisikovih vrsta (ROS) i glutationa (GSH) te aktivnost enzima glutation peroksidaze (GSH-Px). Izmjerene su i aktivnosti ukupnih kolinesteraza (ChE), acetilkolinesteraze (AChE) i butirilkolinesteraze (BChE). Izloženi štakori imali su manje priraste težine od kontrolnih. Izloženost glifosatu uzrokovala je značajne poraste razine primarnih oštećenja DNA u jetrenim stanicama te malo manje u leukocitima. U svim izloženim skupinama izmjerene su niže vrijednosti TBARs u odnosu na kontrolu, sa značajno nižim vrijednostima u AOEL, ADI i cPAD skupinama u uzorcima jetre te u AOEL i cPAD skupinama u uzorcima plazme. Aktivnost AChE bila je smanjena u svim tretmanima, s najnižom stopom nakon izlaganja dozi ADI. Aktivnost BChE blago je smanjena nakon izlaganja ADI, a povećana nakon izlaganja dozama cPAD i 100xAOEL. Ukupna aktivnost ChE te razine ROS/GSH u plazmi / jetri nisu se značajno razlikovale od kontrole, osim značajnog smanjenja jetrenog GSH nakon izlaganja dozama cPAD i 100xAOEL te 35-postotnog smanjenja aktivnosti ChE nakon izlaganja dozama AOEL i ADI. Aktivnost GSH-Px u krvi značajno je smanjena u AOEL i ADI tretmanu, a aktivnost GSH-Px u uzorcima jetre značajno je povećana u skupinama ADI, cPAD i 100xAOEL prema kontroli. Dobiveni rezultati pokazuju da čak i izloženost vrlo niskim dozama glifosata može izazvati mjerljive toksične učinke te upućuje na potrebu za promjenom pristupa procjeni rizika zbog kronične/subkronične izloženosti niskim dozama glifosata gdje oksidacijski stres ne mora nužno korelirati s razinom oštećenja DNA i inhibicijom acetilkolinesteraz

Volatile compounds and amino acids in the urine of testicular tumour patients

Tumor testisa najčešće je maligno oboljenje u muškaraca mlađe životne dobi. Razvojem tumora nastaju promjene u metabolizmu stanica koje se mogu pratiti analizom metabolita u biološkim uzorcima ispitanika što može biti koristan alat za procjenu stanja organizma i daljnje liječenje bolesti. Cilj ovog istraživanja bio je kreirati kvantitativni profil slobodnih aminokiselina i elemenata u urinu ispitanika s tumorom testisa i odrediti vrijednosti parametara oksidacijsko-redukcijske ravnoteže u krvi. U ispitanika s tumorom testisa izmjerene su značajno povišene koncentracije asparaginske kiseline, mangana i cinka te aktivnost superoksid dismutaze dok su koncentracija treonina, serina, histidina, kobalta i glutationa te ukupni oksidacijski status, bili značajno niži u odnosu na kontrolne ispitanike. Nadalje, provedena je linearna diskriminantna analiza kako bi se ispitalo razdvaja li kreirani profil aminokiselina ispitanike s tumorom testisa od kontrolnih ispitanika, a uspješnost diskriminacije iznosila je 86 %. U urinu ispitanika s tumorom testisa kreiran je i kvalitativni profil hlapljivih organskih metabolita kako bi detektirali koje se vrste, ovisno o funkcionalnim skupinama, javljaju u urinu. Analizom urina u kromatogramu je detektirano oko 200 pikova hlapljivih organskih metabolita te su identificirana 123 metabolita. Procijenjen je učinak kemoterapije cisplatinom na koncentraciju aminokiselina i elemenata te na parametre oksidacijsko-redukcijske ravnoteže. Nakon primljene kemoterapije, u ispitanika s tumorom testisa izmjerene su povišene koncentracije većine aminokiselina i elemenata u urinu te glutationa u krvnoj plazmi u odnosu na vrijednosti prije kemoterapije. Godinu dana nakon kemoterapije rezultati su pokazali slične vrijednosti za većinu aminokiselina osim za treonin, serin i živu koji su bili značajno sniženi, te mangan i platinu koji su i dalje bili značajno povišeni kod ispitanika koji su bolovali od tumora testisa u odnosu na kontrolne ispitanike. Rezultati upućuju na postupni oporavak metabolizma nakon prestanka kemoterapije. Ovim istraživanjem dobiveni su rezultati koji upućuju na to da razvojem tumora u testisima dolazi do poremećaja u ciklusu limunske kiseline i ciklusu uree. Dobiveni rezultati mogu poslužiti kao temelj za daljnja istraživanja o ulozi međuprodukata navedenih ciklusa u tumorigenezi što može pridonijeti eventualnom otkrivanju novih bioloških biljega tumora testisa.Testicular cancer is the most common malignancy in young men and in recent decades its occurrence has been growing rapidly. Tumour development induces changes in cell metabolism that can be monitored by analysing the metabolites in biological samples of the subject. The aim of this study was to create a quantitative profile of free amino acids and elements in urine and levels of oxidative stress parameters in blood of men with testicular cancer and compare them with control subjects. In subjects with testicular cancer, significantly higher concentrations of aspartic acid, manganese and zinc were measured, as well as a higher activity of superoxide dismutase. On the other hand, concentrations of threonine, serine, histidine, cobalt and glutathione and total oxidant status were significantly lower in comparison to control subjects. Additionally, linear discriminant analysis (LDA) was performed to examine whether the created amino acid profile discriminated subjects with testicular cancer from control subjects and the results showed that the discrimination rate was 86%. In the urine of subjects with testicular cancer a qualitative profile of volatile organic metabolites was also created to detect which species, depending on their functional groups, can be detected in urine. GCMS analysis of urine detected about 200 peaks of volatile organic metabolites, and 123 of them were identified. The effect of cisplatin chemotherapy on the concentration of amino acids and elements in urine and on the parameters of oxidative stress was also estimated. After receiving chemotherapy, concentrations of the majority of amino acids and elements in urine and glutathione in blood plasma increased in comparison to values obtained before chemotherapy. One year after receiving chemotherapy the results showed that the concentrations of most amino acids and elements were similar to those measured in the control subjects, except for threonine, serine and mercury, which had significantly higher values, and manganese and platinum, which had significantly lower values than the control subjects. Results indicated a gradual metabolism recover following chemotherapy. The results of this research suggest that the development of a tumour in the testicles leads to a disorder in the citric acid cycle and the urea cycle. The obtained results can be used as a basis for further research on the role of the intermediates of the above-mentioned cycles in tumorigenesis, which could contribute to the possible discovery of new biomarkers for testicular tumours

Oxidative stress in triazine pesticide toxicity: a review of the main biomarker findings

This review article provides a summary of the studies relying on oxidative stress biomarkers (lipid peroxidation and antioxidant enzymes in particular) to investigate the effects of atrazine and terbuthylazine exposure in experimental animals and humans published since 2010. In general, experimental animals showed that atrazine and terbuthylazine exposure mostly affected their antioxidant defences and, to a lesser extent, lipid peroxidation, but the effects varied by the species, sex, age, herbicide concentration, and duration of exposure. Most of the studies involved aquatic organisms as useful and sensitive bio-indicators of environmental pollution and important part of the food chain. In laboratory mice and rats changes in oxidative stress markers were visible only with exposure to high doses of atrazine. Recently, our group reported that low-dose terbuthylazine could also induce oxidative stress in Wistar rats. It is evident that any experimental assessment of pesticide toxic effects should take into account a combination of several oxidative stress and antioxidant defence biomarkers in various tissues and cell compartments. The identified effects in experimental models should then be complemented and validated by epidemiological studies. This is important if we wish to understand the impact of pesticides on human health and to establish safe limits.Ovaj pregledni članak daje sažetak studija objavljenih od 2010. godine o djelovanju atrazina i terbutilazina na parametre oksidacijskog stresa (osobito lipidnu peroksidaciju i antioksidacijske enzime) u eksperimentalnih životinja i ljudi. Općenito je utvrđeno da izloženost eksperimentalnih životinja atrazinu i terbutilazinu najviše utječe na njihovu antioksidacijsku obranu i u manjoj mjeri na lipidnu peroksidaciju, ali učinci variraju ovisno o vrsti, spolu, dobi, koncentraciji herbicida i trajanju izlaganja. Većina studija uključivala je vodene organizme kao korisne i osjetljive bio-pokazatelje onečišćenja okoliša i važan dio prehrambenog lanca. U laboratorijskih miševa i štakora promjene u markerima oksidacijskog stresa bile su vidljive samo nakon izloženosti visokim dozama atrazina. U svojem smo nedavnom izvješću iznijeli podatak da niske doze terbutilazina također mogu inducirati oksidacijski stres u Wistar štakora. Očigledno je da pri svakoj eksperimentalnoj procjeni toksičnih učinaka pesticida treba uzeti u obzir kombinaciju nekoliko biomarkera oksidacijskog stresa i antioksidacijske obrane u različitim tkivima i stanicama. Učinci identificirani u eksperimentalnim modelima trebaju se nadopunjavati i potvrditi epidemiološkim istraživanjima. To je važno ako želimo razumjeti utjecaj pesticida na ljudsko zdravlje i uspostaviti sigurnu granicu izloženosti

Valorization potential of Plantago major L. solid waste remaining after industrial tincture production: Insight into the chemical composition and bioactive properties

Herbal residues from the production and processing of medicinal plants are usually discarded as waste material. Plantago major is an edible plant, traditionally used for medicinal purposes, having wide application in the pharmaceutical and cosmetics industry, usually in the form of liquid extracts and tinctures. In this work, extracts of P. major leaves waste remaining after industrial tincture production and dried leaves used initially for tincture production were investigated and compared. Methods: The chemical composition was obtained by FTIR analysis, whereas polyphenolic profile was assessed by HPLC. Antioxidant activity, sun protection factor (SPF), cytotoxic activity against colon carcinoma (HCT116) and melanoma (Hs294T) human cell lines as well as antistaphylococcal activity against S. aureus ATCC strains and one clinical isolate were also evaluated. Results: FTIR analysis revealed wider chemical diversity in waste samples than in initial plant material. Among detected phenolics, chlorogenic acid, luteolin, and rutin were the most abundant in all extracts, whereas luteolin was even higher in the waste. Waste extracts had a significantly lower sun protection factor (SPF) when compared to initial dried leaves. On the contrary, cytotoxic activity of waste extracts against tested human cell lines were more efficient when compared to initial dried leaves, which can be attributed to the higher luteolin content in tincture residues. Both waste and initial dried leaves extracts exhibited antibacterial activity against all tested S. aureus strains at higher tested concentrations. Conclusion: P. major waste remaining after industrial tincture production represents high-value material with great valorization potential. Graphic abstract: [Figure not available: see fulltext.

Efficient and Green Isolation of Keratin from Poultry Feathers by Subcritical Water

The isolation of keratin from poultry feathers using subcritical water was studied in a batch reactor at temperatures (120–250 °C) and reaction times (5–75 min). The hydrolyzed product was characterized by FTIR and elemental analysis, while the molecular weight of the isolated product was determined by SDS-PAGE electrophoresis. To determine whether disulfide bond cleavage was followed by depolymerization of protein molecules to amino acids, the concentration of 27 amino acids in the hydrolysate was analyzed by GC/MS. The optimal operating parameters for obtaining a high molecular weight protein hydrolysate from poultry feathers were 180 °C and 60 min. The molecular weight of the protein hydrolysate obtained under optimal conditions ranged from 4.5 to 12 kDa, and the content of amino acids in the dried product was low (2.53% w/w). Elemental and FTIR analyses of unprocessed feathers and dried hydrolysate obtained under optimal conditions showed no significant differences in protein content and structure. Obtained hydrolysate is a colloidal solution with a tendency for particle agglomeration. Finally, a positive influence on skin fibroblast viability was observed for the hydrolysate obtained under optimal processing conditions for concentrations below 6.25 mg/mL, which makes the product interesting for various biomedical applications