natgillin/translations-raw huggingface dataset:
Dry moisture controls over night
Količine zbrane krvi so odvisne od velikosti ribe ter so običajno v razponu od 5 do 60 μl na posameznega črnoglavega pisanca in od 5 do 15 μl na posamezno cebrico.
weigh moisture controls
Plazma se od krvi loči s centrifugiranjem in shrani z inhibitorji protaze pri – 80 °C, dokler se ne analizira glede vitelogenina.
measure pH of dried abiotic controls
Pri medaki pa se kot tkivni vir za določanje vitelogenina uporabijo jetra in pri cebrici homogenat glave/repa (Dodatek 6).
Day 3, 6, 9, 12 (approx.)
Meritev VTG mora temeljiti na validirani homologni metodi ELISA, pri čemer se uporabijo homologni standard VTG in homologna protitelesa.
Supply each replicate with sufficient amount of prey organisms
Priporoča se uporaba metode, s katero je mogoče zaznati tako nizke ravni VTG, kot je nekaj ng/ml plazme (ali ng/mg tkiva), kar je raven naravnega ozadja pri neizpostavljenih ribjih samcih.
Dry water content controls over night
Vsaka plošča ELISA, uporabljena za preskuse VTG, mora vsebovati naslednje vzorce za nadzor kakovosti:
weigh water content controls
vsaj 6 kalibracijskih standardov, ki zajemajo razpon pričakovanih koncentracij vitelogenina, in vsaj en nespecifično vezalni preskus slepega vzorca (analiziran v dveh ponovitvah).
measure pH of dried controls
Absorpcija teh slepih vzorcev mora biti manj kot 5 % največje absorpcije kalibracijskega standarda.
Day 16
Analizirana bosta vsaj dva alikvota (ponovitvi vdolbinic) posameznega razredčenja vzorca.
Terminate extraction
Ponovitve vdolbinic, ki se razlikujejo za več kot 20 %, je treba ponovno analizirati.
Day 16 +
Korelacijski koeficient (R2) za umeritvene krivulje mora biti večji od 0,99.
Record number of adults and juveniles in extracted material
Vendar visoka korelacija ni dovolj za zagotovitev ustrezne napovedi koncentracije v vseh razponih.
Report results on template tables
Poleg dovolj visoke korelacije za umeritveno krivuljo mora biti koncentracija posameznega standarda, kot je izračunana po umeritveni krivulji, med 70 in 120 % nominalne koncentracije.
Report testing procedure in test protocol sheets
Če se nominalne koncentracije oddaljujejo od regresijske premice kalibracije (npr. pri nižjih koncentracijah), bo morda treba umeritveno krivuljo razdeliti na nizke in visoke razpone ali uporabiti nelinearen model, ki bo ustrezal podatkom o absorpciji.
C.37.
Če se krivulja razdeli, mora biti R2 na obeh delih > 0,99.
AY FISH ASSAY:
Meja zaznavanja (LOD) je opredeljena kot koncentracija najnižjega analitskega standarda, meja določanja (LOQ) pa je opredeljena kot koncentracija najnižjega analitskega standarda, pomnoženega z najnižjim faktorjem razredčenja.
A SHORT-TERM SCREENING FOR OESTROGENIC AND ANDROGENIC ACTIVITY, AND AROMATASE INHIBITION
Vsak dan izvedbe preskusa vitelogenina se analizira obogateni vzorec, pripravljen z medpreskusnim referenčnim standardom (Dodatek 7).
This test method is equivalent to OECD test guideline (TG) 230 (2009).
Razmerje med pričakovano koncentracijo in izmerjeno koncentracijo se sporoči skupaj z rezultati posameznih nizov preskusov, izvedenih isti dan.
The need to develop and validate a fish assay capable of detecting certain endocrine active chemicals originates from the concerns that environmental levels of chemicals may cause adverse effects in both humans and wildlife due to the interaction of these chemicals with the endocrine system.
Vrednotenje odziva biomarkerja z analizo variance (ANOVA)
In 1998, the OECD initiated a high-priority activity to revise existing guidelines and to develop new guidelines for the screening and testing of potential endocrine disrupters.
Za določitev potencialnega endokrinega delovanja kemikalije se z analizo variance (ANOVA) primerjajo odzivi med tretiranji in kontrolnimi skupinami.
One element of the activity was to develop a Test Guideline for the screening of chemicals active on the endocrine system of fish species.
Kadar se uporabi kontrola s topilom, je treba za vsako končno točko izvesti primeren statistični preskus med vodo za redčenje in kontrolami s topilom.
The 21-day Fish Endocrine Screening Assay underwent an extensive validation programme consisting of inter-laboratory studies with selected chemicals to demonstrate the relevance and reliability of the assay for the detection of oestrogenic and aromatase inhibiting chemicals (1, 2, 3, 4, 5) in the three fish species investigated (the fathead minnow, the Japanese medaka and the zebrafish);
Smernice za obravnavanje podatkov o vodi za redčenje in kontroli s topilom pri nadaljnji statistični analizi so navedene v OECD, 2006c (27).
the detection of androgenic activity is possible in the fathead minnow and the medaka, but not in the zebrafish.
Vse podatke o biološkem odzivu je treba analizirati in sporočiti ločeno po spolu.
This test method does not allow the detection of anti-androgenic chemicals.
Če niso izpolnjene zahtevane domneve za parametrske metode – nenormalna porazdelitev (npr. Shapiro-Wilkov preskus) ali heterogena varianca (Bartlettov ali Levenejev preskus), je potreben razmislek o preoblikovanju podatkov, da se poenotijo variance pred izvedbo ANOVA ali pred izvedbo tehtane ANOVA.
The validation work has been peer-reviewed by a panel of experts nominated by the National Coordinators of the Test Guideline Programme (6).
Za nemonotoni odziv na odmerek se lahko uporabi Dunnettov preskus (parametrski) večkratnih primerjav med pari ali Mann-Whitneyjev preskus z Bonferroni-Holmovo prilagoditvijo (neparametrski).
The assay is not designed to identify specific mechanisms of hormonal disruption because the test animals possess an intact hypothalamic-pituitary-gonadal (HPG) axis, which may respond to chemicals that impact on the HPG axis at different levels.
Če je odziv na odmerek približno monoton, se lahko uporabijo drugi statistični preskusi (npr. preskus po Jonckheere-Terpstraju ali Williamsov preskus).
The Fish Short Term Reproduction assay (OECD TG 229) includes fecundity and, as appropriate, gonadal histopathology for the fathead minnow, as well as all endpoints included in this test method.
V pomoč pri odločitvi glede izbire najprimernejšega statističnega preskusa, ki naj se uporabi, je v Dodatku 8 zagotovljena statistična tabela poteka.
OECD TG 229 provides a screening of chemicals which affect reproduction through various mechanisms including endocrine modalities.
Dodatne informacije so navedene v dokumentu OECD o sedanjih pristopih k statistični analizi podatkov o ekotoksičnosti (27).
This should be considered prior to selecting the most appropriate test method.
Poročanje o rezultatih preskusa
This test method describes an in vivo screening assay where sexually mature male and spawning female fish are held together and exposed to a chemical during a limited part of their life-cycle (21 days).
Podatki študije morajo obsegati:
At termination of the 21-day exposure period, depending on the species used, one or two biomarker endpoint(s) are measured in males and females as indicators of oestrogenic, aromatase inhibition or androgenic activity of the test chemical;
Preskusna infrastruktura:
these endpoints are vitellogenin and secondary sexual characteristics.
odgovorno osebje in njihove obveznosti v zvezi s študijo,
Vitellogenin is measured in fathead minnow, Japanese medaka and zebrafish, whereas secondary sex characteristics are measured in fathead minnow and Japanese medaka only.
vsak laboratorij mora s sklopom reprezentativnih kemikalij dokazati usposobljenost.
This bioassay serves as an in vivo screening assay for certain endocrine modes of action and its application should be seen in the context of the “OECD Conceptual Framework for the Testing and Assessment of Endocrine Disrupting Chemicals” (28).
opredelitev preskusne kemikalije,
INITIAL CONSIDERATIONS AND LIMITATIONS
fizikalno stanje in ustrezne fizikalno-kemijske lastnosti,
Vitellogenin is normally produced by the liver of female oviparous vertebrates in response to circulating endogenous oestrogen.
metodo in pogostost priprave preskusnih koncentracij,
It is a precursor of egg yolk proteins and, once produced in the liver, travels in the bloodstream to the ovary, where it is taken up and modified by developing eggs.
informacije o stabilnosti in biološki razgradljivosti.
Vitellogenin is almost undetectable in the plasma of immature female and male fish because they lack sufficient circulating oestrogen;
Topilo:
however, the liver is capable of synthesizing and secreting vitellogenin in response to exogenous oestrogen stimulation.
opredelitev topila (vrsta, uporabljena koncentracija),
The measurement of vitellogenin serves for the detection of chemicals with various oestrogenic modes of action.
utemeljitev izbire topila (če to ni voda).
The detection of oestrogenic chemicals is possible via the measurement of vitellogenin induction in male fish, and it has been abundantly documented in the scientific peer-reviewed literature (e.g. (7)).
Preskusne živali:
Vitellogenin induction has also been demonstrated following exposure to aromatizable androgens (8, 9).
vrsto in sev,
A reduction in the circulating level of oestrogen in females, for instance through the inhibition of the aromatase converting the endogenous androgen to the natural oestrogen 17β-estradiol, causes a decrease in the vitellogenin level, which is used to detect chemicals having aromatase inhibiting properties (10, 11).
dobavitelja in specifično infrastrukturo dobavitelja,
The biological relevance of the vitellogenin response following oestrogenic/aromatase inhibition is established and has been broadly documented.
starost rib na začetku preskusa in razmnoževalno stanje/stanje drstenja,
However, it is possible that production of VTG in females can also be affected by general toxicity and non-endocrine toxic modes of action, e.g. hepatotoxicity.
podrobnosti o postopku aklimatizacije živali,
Several measurement methods have been successfully developed and standardised for routine use.
telesno težo rib na začetku izpostavljenosti (iz podvzorca staleža rib).
This is the case of species-specific Enzyme-Linked Immunosorbent Assay (ELISA) methods using immunochemistry for the quantification of vitellogenin produced in small blood or liver samples collected from individual fish (12, 13, 14, 15, 16, 17, 18).
Preskusni pogoji:
Fathead minnow blood, zebrafish blood or head/tail homogenate, and medaka liver are sampled for VTG measurement.
uporabljeni preskusni postopek (vrsta preskusa, stopnja obremenitve, gostota rib itd.),
In medaka, there is a good correlation between VTG measured from blood and from liver (19).
metodo priprave založnih raztopin in pretok,
Appendix 6 provides the recommended procedures for sample collection for vitellogenin analysis.
nominalne preskusne koncentracije, tedensko izmerjene koncentracije preskusnih raztopin in uporabljeno analitsko metodo, sredine izmerjenih vrednosti in standardnih odklonov v preskusnih posodah ter dokaze, da se meritve nanašajo na koncentracije preskusne kemikalije v dejanski raztopini,
Kits for the measurement of vitellogenin are widely available;
značilnosti vode za redčenje (vključno s pH, trdoto, alkalnostjo, temperaturo, koncentracijo raztopljenega kisika, koncentracijo sledi klora, skupnim organskim ogljikom, suspendiranimi trdnimi snovmi in vsemi ostalimi opravljenimi meritvami),
such kits should be based on a validated species-specific ELISA method.
kakovost vode v preskusnih posodah:
Secondary sex characteristics in male fish of certain species are externally visible, quantifiable and responsive to circulating levels of endogenous androgens;
pH, trdota, temperatura in koncentracija raztopljenega kisika,
this is the case for the fathead minnow and the medaka — but not for zebrafish, which does not possess quantifiable secondary sex characteristics.
podrobne informacije o hranjenju (npr. vrsta hrane, vir, dana količina in pogostost dajanja ter analiza zadevnih onesnaževal, če je na voljo (npr. PCB, PAH in organoklorni pesticidi).
Females maintain the capacity to develop male secondary sex characteristics, when they are exposed to androgenic chemicals in water.
dokaze, da kontrole izpolnjujejo merila za sprejemljivost preskusa,
Several studies are available in the scientific literature to document this type of response in fathead minnow (20) and medaka (21).
podatke o smrtnosti za posamezne preskusne koncentracije in kontrole,
A decrease in secondary sex characteristics in males should be interpreted with caution because of low statistical power, and should be based on expert judgement and weight of evidence.
uporabljene tehnike statistične analize, obdelavo podatkov in utemeljitev uporabljenih tehnik,
There are limitations to the use of zebrafish in this assay, due to the absence of quantifiable secondary sex characteristics responsive to androgenic acting chemicals.
podatke o bioloških opažanjih splošne morfologije, vključno s sekundarnimi spolnimi značilnostmi in vitelogeninom,
In the fathead minnow, the main indicator of exogenous androgenic exposure is the number of nuptial tubercles located on the snout of the female fish.
rezultate analize podatkov, po možnosti v obliki tabele in grafičnega prikaza,
In the medaka, the number of papillary processes constitutes the main marker of exogenous exposure to androgenic chemicals in female fish.
pojav kakršnih koli neobičajnih reakcij rib in kakršnih koli vidnih učinkov, ki jih povzroči preskusna kemikalija.
Appendix 5A and Appendix 5B indicate the recommended procedures to follow for the evaluation of sex characteristics in fathead minnow and in medaka, respectively.
SMERNICA ZA RAZLAGO IN SPREJEMLJIVOST REZULTATOV PRESKUSA
Definitions used in this test method are given in Appendix 1.
V tem razdelku je navedenih nekaj pomislekov, ki jih je treba upoštevati pri razlagi rezultatov preskusa za različne izmerjene končne točke.
In the assay, male and female fish in a reproductive status are exposed together in test vessels.
Rezultate je treba previdno razlagati, kadar se zdi, da preskusna kemikalija povzroča očitno strupenost ali vpliva na splošno stanje preskusne živali.
Their adult and reproductive status enables a clear differentiation of each sex, and thus a sex-related analysis of each endpoint, and ensures their sensitivity towards exogenous chemicals.
Pri določanju razpona preskusnih koncentracij je treba paziti, da se ne preseže najvišja tolerančna koncentracija, da bo mogoča smiselna razlaga podatkov.
At test termination, sex is confirmed by macroscopic examination of the gonads following ventral opening of the abdomen with scissors.
Kadar ni znakov strupenih učinkov, je pomembno, da se izvede vsaj eno tretiranje.
An overview of the relevant bioassay conditions is provided in Appendix 2.
Znake bolezni in znake strupenih učinkov je treba temeljito oceniti in o njih poročati.
The assay is normally initiated with fish sampled from a population that is in spawning condition;
Možno je na primer, da na nastajanje VTG pri samicah vplivajo tudi splošna strupenost in neendokrini strupeni načini delovanja, npr. hepatotoksičnost.
senescent animals should not be used.
Vendar se lahko razlaga učinkov podkrepi z drugimi stopnjami tretiranja, na katere ne vpliva sistemska strupenost.
Guidance on the age of fish and on the reproductive status is provided in the section on Selection of fish.
V zvezi s sprejemljivostjo rezultatov preskusa je treba upoštevati nekaj vidikov.
The assay is conducted using three chemical exposure concentrations as well as a water control, and a solvent control if necessary.
Ravni VTG v kontrolnih skupinah samcev in samic morajo biti kot smernica izrazite ter ločene za približno tri rede velikosti pri črnoglavem pisancu in cebrici ter približno en red velikosti pri medaki.
Two vessels or replicates per treatment are used (each vessel containing 5 males and 5 females) in medaka and zebrafish, whereas four vessels or replicates per treatment are used (each vessel containing 2 males and 4 females) in fathead minnow.
Primeri razpona vrednosti v kontrolnih in tretiranih skupinah so navedene v validacijskih poročilih (1) (2) (3) (4).
This is to accommodate the territorial behaviour of male fathead minnow while maintaining sufficient power of the assay.
Visoke vrednosti VTG pri samcih iz kontrol bi lahko ogrozile odzivnost preskusa in njegovo zmožnost za zaznavanje šibkih agonistov estrogena.
The exposure is conducted for 21 days and sampling of fish is performed at day 21 of exposure.
Nizke vrednosti VTG pri samicah iz kontrol bi lahko ogrozile odzivnost preskusa ter njegovo zmožnost za zaznavanje inhibitorjev aromataze in antagonistov estrogena.
On sampling at day 21, all animals are killed humanely.
Pri pripravi te smernice so bile uporabljene validacijske študije.
Secondary sex characteristics are measured in fathead minnow and medaka (see Appendix 5A and Appendix 5B);
Če laboratorij še ni izvedel preskusa ali je prišlo do znatnih sprememb (npr. sprememba seva ali dobavitelja rib), se priporoča izvedba študije tehnične usposobljenosti.
blood samples are collected for determination of vitellogenin in zebrafish and fathead minnow, alternatively head/tail can be collected for the determination of vitellogenin in zebrafish (Appendix 6);
Priporočljivo je, da se uporabijo kemikalije, ki zajemajo različne načine delovanja ali vplive na številne preskusne končne točke.
liver is collected for VTG analysis in medaka (Appendix 6).
V praksi se vsak laboratorij spodbuja k vzpostavitvi lastnih podatkov o kontroli iz preteklih preskusov za samce in samice ter izvedbi pozitivne kontrole za estrogeno delovanje (npr. 17β-estradiol pri100 ng/l ali znan šibek agonist), ki povzroči povečanje VTG pri ribjih samcih, pozitivne kontrole za zaviranje aromataze (npr. fadrozol ali prokloraz pri 300 μg/l), ki povzroči zmanjšanje VTG pri ribjih samicah, in pozitivno kontrolo za androgeno delovanje (npr. 17β-trenbolon pri 5 μg/l), ki povzroči indukcijo sekundarnih spolnih značilnosti pri samicah črnoglavega pisanca in medake.
TEST ACCEPTANCE CRITERIA
Vsi ti podatki se lahko primerjajo z razpoložljivimi podatki iz validacijskih študij (1) (2) (3), da se zagotovi usposobljenost laboratorija.
For the test results to be acceptable the following conditions apply:
Na splošno je treba meritve vitelogenina šteti za pozitivne, če se VTG pri samcih statistično značilno poveča (p < 0,05) ali pri samicah statistično značilno zmanjša (p < 0,05) vsaj pri najvišjem preskušenem odmerku v primerjavi s kontrolno skupino ter ob odsotnosti znakov splošne strupenosti.
the mortality in the water (or solvent) controls should not exceed 10 % at the end of the exposure period;
Pozitiven rezultat je nadalje podprt s prikazom biološko verjetnega razmerja med odmerkom in krivuljo odziva.
the dissolved oxygen concentration should be at least 60 % of the air saturation value (ASV) throughout the exposure period;
Kot je bilo navedeno, zmanjšanje vitelogenina morda ni v celoti endokrinega izvora;
the water temperature should not differ by more than ± 1,5 °C between test vessels at any one time during the exposure period and be maintained within a range of 2 °C within the temperature ranges specified for the test species (Appendix 2);
vendar je treba pozitiven rezultat na splošno razlagati kot dokaz endokrinega delovanja in vivo ter si prizadevati za nadaljnjo pojasnitev.
evidence should be available to demonstrate that the concentrations of the test chemical in solution have been satisfactorily maintained within ± 20 % of the mean measured values.
Screening and testing for endocrine disruption in fish-biomarkers as ‚signposts,‘ not ‚traffic lights,‘ in risk assessment.
Normal laboratory equipment and especially the following:
Environmental Health Perspectives;
oxygen and pH meters;
Okrajšave in opredelitev pojmov
equipment for determination of water hardness and alkalinity;
Kemikalija
adequate apparatus for temperature control and preferably continuous monitoring;
snov ali zmes.
tanks made of chemically inert material and of a suitable capacity in relation to the recommended loading and stocking density (see Appendix 2);
koeficient variacije.
spawning substrate for fathead minnow and zebrafish, Appendix 4 gives the necessary details;
encimski imunski test.
suitably accurate balance (i.e. accurate to ± 0,5 mg).
Stopnja obremenitve
Any water in which the test species shows suitable long-term survival and growth may be used as test water.
mokra teža rib na količino vode.
It should be of constant quality during the period of the test.
Gostota rib
The pH of the water should be within the range 6,5 to 8,5, but during a given test it should be within a range of ± 0,5 pH units.
število rib na količino vode.
In order to ensure that the dilution water will not unduly influence the test result (for example by complexion of test chemical), samples should be taken at intervals for analysis.
VTG (vitelogenin)
Measurements of heavy metals (e.g. Cu, Pb, Zn, Hg, Cd, and Ni), major anions and cations (e.g. Ca2+, Mg2+, Na+, K+, Cl-, and SO42-), pesticides (e.g. total organophosphorus and total organochlorine pesticides), total organic carbon and suspended solids should be made, for example, every three months where dilution water is known to be relatively constant in quality.
fosfoglikoprotein in predstopnja rumenjakovih beljakovin, ki se običajno pojavi pri spolno aktivnih samicah vseh oviparnih vrst.
If water quality has been demonstrated to be constant over at least one year, determinations can be less frequent and intervals extended (e.g. every six months).
Os HPG
Some chemical characteristics of acceptable dilution water are listed in Appendix 3.
os hipotalamus – hipofiza – spolne žleze.
Test solutions of the chosen concentrations are prepared by dilution of a stock solution.
najvišja tolerančna koncentracija, ki predstavlja približno 10 % LC50.
The stock solution should preferably be prepared by simply mixing or agitating the test chemical in dilution water by using mechanical means (e.g. stirring or ultrasonication).
Poskusni pogoji za endokrini presejalni preskus rib
Saturation columns (solubility columns) can be used for achieving a suitable concentrated stock solution.
Priporočene vrste
For difficult test chemicals, a solvent may be technically the best solution;
Vrsta preskusa
the OECD Guidance Document on aquatic toxicity testing of difficult substances and mixtures should be consulted (22).
pretočni
The choice of solvent will be determined by the chemical properties of the chemical.
Temperatura vode
The OECD Guidance Document recommends a maximum of 100 μl/l, which should be observed.
Kakovost osvetljenosti
However a recent review (23) highlighted additional concerns when using solvents for endocrine activity testing.
fluorescenčne sijalke (široki spekter)
Therefore it is recommended that the solvent concentration, if necessary, is minimised wherever technically feasible (dependent on the physical-chemical properties of the test chemical).
10–20 μE/m2/s, 540–1000 luksov ali 50–100 ft-c (ravni v laboratoriju)
A flow-through test system will be used.
Obdobje osvetljenosti (svitanje in mrak sta neobvezna, vendar se ne štejeta za potrebna)
Such a system continually dispenses and dilutes a stock solution of the test chemical (e.g. metering pump, proportional diluter, saturator system) in order to deliver a series of concentrations to the test chambers.
16 ur svetlobe, 8 ur teme
The flow rates of stock solutions and dilution water should be checked at intervals, preferably daily, during the test and should not vary by more than 10 % throughout the test.
12–16 ur svetlobe, 12-8 ur teme
Care should be taken to avoid the use of low-grade plastic tubing or other materials that may contain biologically active chemicals.
< 5 g/l
When selecting the material for the flow-through system, possible adsorption of the test chemical to this material should be considered.
Velikost preskusne komore
Holding of fish
10 l (minimum)
Test fish should be selected from a laboratory population, preferably from a single stock, which has been acclimated for at least two weeks prior to the test under conditions of water quality and illumination similar to those used in the test.
2 l (minimum)
It is important that the loading rate and stocking density (for definitions, see Appendix 1) be appropriate for the test species used (see Appendix 2).
5 l (minimum)