natgillin/translations-raw huggingface dataset:
I vandig opløsning spaltes brintperoxid spontant, men kun meget langsomt, til ilt og vand.
In an aqueous solution, hydrogen peroxide will spontaneously split into oxygen and water, but the process is very slow.
Ved tilstedeværelse af katalase øges reaktionshastigheden imidlertid betydeligt.
If catalase is present, however, the rate of reaction is increased significantly.
Reaktionen er som følger:
The reaction is as follows:
I det tilfælde hvor f.eks. ureaperoxid (CAS nr. 124-43-6) indgår, ser ligevægten imellem ureaperoxid og frit brintperoxid således ud:
In cases where, for instance, urea peroxide (CAS no. 124-43-6) is contained in the product, the equilibrium between urea peroxide and the free hydrogen peroxide is as follows:
Den frigjorte brintperoxid vil herefter løbende omdannes af katalasen til vand og ilt efter formel (1) ovenfor, der gør at ligevægten forskydes mod højre.
The released hydrogen peroxide is cleaved by the catalase into water and oxygen according to formula (1) above, shifting the equilibrium towards the right.
For de øvrige releasere, som er testet i denne rapport, findes nedenfor en gennemgang af de kemiske ligevægte som resulterer i en frigivelse af brintperoxid.
For the remaining releasers tested in this report, an outline of their chemical equilibrium as a result of the release of hydrogen peroxide is given below.
Calciumperoxids (CAS nr. 1305-79-9) opløselighed er stærkt pH-afhængig, ligesom ligevægten der indstilles i henhold til nedenstående reaktion afhænger af pH:
The solubility of calcium peroxide (CAS no. 1305-79-9) is highly dependent on pH, as is the equilibrium created according to the reaction below:
Natriumperborate-monohydrat (CAS nr. 10332-33-9) er en dimer, der er bundet sammen af ægte peroxygen-bindinger.
Sodium perborate monohydrate (CAS no. 10332-33-9) is a dimer that is bonded together by genuine peroxygen bonds.
I vandig opløsning ved pH 7, dannes brintperoxid og natriumborat, og der indstilles følgende ligevægt:
In an aqueous solution at pH7, hydrogen peroxide and sodium borate are created and the following equilibrium is created:
Natriumpercarbonat (CAS nr. 15630-89-4) har en stor vandopløselighed og frigiver i vand carbonat og brintperoxid efter følgende reaktion
Sodium percarbonate (CAS no. 15630-89-4) is highly water soluble and when dissolved in water releases carbonate and hydrogen peroxide according to the following reaction:
I produkter indeholdende en til flere ”releasere”, vil assayet fremme omdannelsen af den bundne fraktion af peroxider til frit brintperoxid.
In products containing one or more releasers, the assay will promote the conversion of the bound fraction of peroxides to free hydrogen peroxide.
Derved opnås en bestemmelse af den totale mængde brintperoxid i produktet.
In this way, determination of the total quantity of hydrogen peroxide in the product is achieved.
Enzymet findes naturligt intracellulært i alle aerobt respirerende organismer, hvor dets funktion er at nedbryde reaktive ilt-radikaler, der dannes som et biprodukt i åndingsprocessen.
The enzyme is naturally present intracellularly in all aerobically respirating organisms, where its function is to break down the reactive oxygen radicals created as a by-product of the respiratory process.
Indholdet af peroxider i tandblegeprodukter opgives normalt i vægt-procent.
The content of peroxides in tooth-whitening products is usually stated as a percentage of weight.
Det vil sige, at et tandblegemiddel som f.eks. indeholder ureaperoxid udgør selve brintperoxiden kun ca. 35 % af molekylets vægt.
That is, in a tooth-whitening agent which contains, for instance, urea peroxide, the hydrogen peroxide represents only about 35% of the weight of the molecule.
Dermed vil blegeeffekten af et produkt indeholdende f.eks. 18 % ureaperoxid omtrent svare til et produkt indeholdende 6 % brintperoxid.
The bleaching effect of a product containing e.g. 18% urea peroxide will therefore approximately correspond to a product containing about 6% hydrogen peroxide.
Tilsvarende forskelle i blegeeffekt vil gøre sig gældende også for de øvrige releasere.
Similar differences in bleaching effect will apply for the remaining releasers.
3 Udvikling og test af katalase bioassay til bestemmelse af peroxider
3 Development and testing of catalase bioassay to determine peroxides
Dernæst er der blevet gennemført en metodeindkøring, hvor
the optimal quantity of active substance (peroxide) was determined;
den optimale mængde aktiv stof (peroxid) er fastlagt
the optimal quantity of enzyme was determined;
den hensigtsmæssige reaktionstid er kortlagt
the optimal pH was identified.
Protokollen er efterfølgende blevet afprøvet på forskellige tandblegeprodukter indkøbt i detailhandlen.
This protocol was subsequently tested on different tooth-whitening products purchased on the Danish retail market.
3.1 Teknisk design af målekammer, elektrode og dataopsamling
3.1 Technical design of measuring chamber, probe and data collection
Der er benyttet et iltmåle- og regulatorsystem designet og fremstillet i samarbejde med Loligo Systems, Tjele, Danmark.
An oxygen measurement and regulator system was used, designed and developed in cooperation with Loligo Systems, Tjele, Denmark.
I målekammeret findes en glascoatet magnet som ved omrøring (1000 rpm) sikrer en ensartet opblanding af alle komponenterne under reaktionen og iltmålingen.
Inside the measuring chamber is a glass-coated magnet, which, when rotated (1,000 rpm) ensures uniform mixture of all the components during the reaction and oxygen measurement.
Figur 1: Opstilling til katalase bioassay.
Figure 1: Set-up for catalase bioassay
Elektroden har en maksimal måleusikkerhed i følge specifikationerne på +/- 1 % af den målte værdi.
According to the specifications the probe has a maximum measurement uncertainty of +/- 1% of the measured value.
Data opsamles hvert sekund ved hjælp af datalogger (fra Measurement Computing, MA, USA) og overføres til Windows-baseret software (TracerDAQ) til online måling af iltudviklingen.
Data is collected every second using a data logger (from Measurement Computing, MA, USA) and transferred to a Windows-based software (TracerDAQ.
Elektrodens specifikationer er vedlagt i Bilag 1 og ordre-oversigt fra Loligo Systems findes i Bilag 2.
Specifications for the probe are included in Annex 1 and an order overview from Loligo Systems is in Annex 2.
Ved målingens start fyldes glaskammeret med kvælstof gennemboblet autoklaveret fosfatbuffer (pH = 7), hvorefter akryllåget med ilt-elektroden monteres.
Before measuring commences, the glass chamber is filled with autoclaved phosphate buffer (pH = 7) purged with nitrogen, after which the acrylic lid with the oxygen probe is mounted.
Fjernelse af ilten i bufferen ved kvælstof-gennembobling sikrer, at iltudviklingen kan følges uden risiko for bobledannelse p.gr.a. overmætning i kammeret.
Removal of oxygen in the buffer by nitrogen gassing ensures that the changes in oxygen concentrations can be monitored without bubbles forming due to over saturation in the chamber.
Bufferen steriliseres ved autoklavering ligesom udstyret steriliseres ved afspritning for at forhindre at biologisk iltforbrug under kørslen påvirker måleresultatet.
The buffer is sterilised using autoclaving, while the equipment is sterilised with alcohol in order to prevent biological oxygen consumption during the run from affecting the measurements.
Enzymet katalase (Sigma-Aldrich, katalognummer C100) tilsættes nu i kendt mængde.
Catalase (Sigma-Aldrich, cat. no. C100) is now added in a known quantity.
I alle forsøgene er det tilsat ca. 5000 units katalase til forsøgskammeret (1 unit nedbryder 1 µmol H2O2 pr. min. ved 25 °C).
In all of the tests, about 5,000 units of catalase were added to the test chamber (1 unit degrades 1 µmol H2O2 per minute at 25°C).
Umiddelbart inden målingen påbegyndes, tilsættes en kendt mængde peroxid eller tandblegemiddel.
Immediately prior to analysis, a known quantity of peroxide or tooth-whitening agent is added.
Da ilts opløselighed i væske er temperaturafhængig er alle målinger foretaget ved samme temperatur (22 °C) i et airconditioneret rum.
Since the solubility of oxygen in a liquid is temperature-dependent, all measurements were carried out at the same temperature (22°C) in an air-conditioned room.
Ved de endelige udregninger er der korrigeret for de små variationer i kammervolumen.
The final calculations include adjustments for the small variations in chamber volume.
3.1.2 Test af elektrodens stabilitet
3.1.2 Testing of the stability of the probe
En sammenligning mellem elektrodens udslag målt i volt (V) ved uafhængige kalibreringer foretaget over 20 dage demonstrerer en stor elektrode-stabilitet, idet afvigelsen er mindre end 1 % af gennemsnitsværdien (Tabel 1).
A comparison of probe outputs measured in volts (V) in independent calibrations conducted over a 20-day period demonstrated great probe stability, as the deviation was less than 1% of the average value (Table 1).
Stabilitet af ilt-elektroden testet ved at måle output (Volt) ved 0 og 100 % iltmætning (n = 20).
Stability of the oxygen probe tested by measuring output (volts) at 0 and 100% oxygen saturation (n = 20).
3.1.3 Test af reaktionskammerets tæthed
3.1.3 Testing of the tightness of the reaction chamber
For at forhindre tab af ilt under målingen, og modsvarende forhindre luft i at trænge ind i reaktionskammeret, er det vigtigt at reaktionskammer og elektrode udgør en tætsluttende enhed.
In order to prevent the loss of oxygen during measuring, and correspondingly, to prevent air from entering the reaction chamber, the reaction chamber and the probe must comprise a tightly sealed unit.
Figur 2: Reaktionskammerets tæthed og egetforbrug af ilt.
Figure 2: Reaction chamber tightness and consumption of oxygen
Dette skyldes sandsynligvis et mikrobielt forbrug af ilt, da målingen i dette tilfælde ikke blev gennemført under sterile forhold.
This is probably due to microbial consumption of oxygen, as the measurement in this case was not conducted under sterile conditions.
I de efterfølgende måleserier (22 og 75 %) og i alle efterfølgende målinger er væsken forinden steriliseret og kamrene aftørret med sprit for at minimere denne usikkerhedsfaktor.
In the subsequent series of measurements (22% and 75%) and in all subsequent measurements, the liquid was sterilised and the chambers wiped off with alcohol beforehand to minimise this factor of uncertainty.
3.2 Fastlæggelse af optimale betingelser for katalase bioassay
3.2 Determination of the optimal conditions for the catalase bioassay
Indkøring af metoden er sket ved anvendelse af enten brintperoxid og ureaperoxid i kendte mængder og i ren form.
The method was tested using either hydrogen peroxide and urea peroxide in known quantities or in pure forms.
Disse to stoffer er i dag de hyppigst anvendte peroxider i tandblegemidler.
These two substances are the most commonly used peroxides in tooth-whitening products.
Det drejer sig om natriumperborat-monohydrat, natriumperborat-tetrahydrat og natriumpercarbonat, der i reaktionsmønster minder om brintperoxid og ureaperoxid, samt calciumperoxid der er lidt anderledes, idet dets reaktion med vand er meget pH-afhængig.
The method was moreover tested on four pure substances, which are also used in tooth-whitening products, namely sodium perborate monohydrate, sodium perborate tetrahydrate and sodium percarbonate.
3.2.1 Test af metodens optimale måleområde og nedre detektionsgrænse.
3.2.1 Testing the optimal measurement scope and lower detection limit of the method
Der er lavet en forsøgsserie med varierende koncentrationer af både brintperoxid (50, 150 og 170 µl af en 1 % opløsning tilsat reaktionskammeret) og ureaperoxid (100, 250 og 500 µl af en 1 % opløsning), for at fastslå metodens optimale måleområde (Figur 3 og 4).
A series of tests were conducted with varying concentrations of both hydrogen peroxide (50, 150 and 170 µl of a 1% solution added to the reaction chamber) and urea peroxide (100, 250 and 500 µl of a 1% solution) in order to determine the method's optimal measurement range (Figs. 3 and 4).
Figur 3: Iltudvikling i reaktionskammeret ved tilsætning af henholdsvis 50, 150 og 170 µl af en 1 % brintperoxidopløsning.
Figure 3: Changes in oxygen concentration in the reaction chamber after addition of 50, 150 and 170 µl, respectively, of a 1% hydrogen peroxide solution.
Figur 4: Iltudvikling i reaktionskammeret ved tilsætning af 100, 250 og 500 µl af en 1 % ureaperoxidopløsning.
Figure 4: Changes in oxygen concentration in the reaction chamber after addition of 100, 250 and 500 µl, respectively, of a 1% urea peroxide solution.
Ved tilsætning af katalase efter 110 sek. ses en meget hurtig iltudvikling på grund af enzymets spaltning af brintperoxid (Figur 3).
When catalase is added after 110 sec., oxygen is rapidly released because the enzyme splits the hydrogen peroxide (Fig. 3).
Det bemærkes, at tilsætning af 170 µl brintperoxid resulterer i et ustabilt signal, hvilket sandsynligvis skyldes dannelse af bobler omkring elektrodens overflade som forstyrrer målingen.
Note that addition of 170 µl hydrogen peroxide results in an unstable signal, probably due to the formation of bubbles on the surface of the probe.
Bobledannelsen skyldes formentlig at ligevægten mellem opløst og frit ilt ikke kan opretholdes under den voldsomme momentane reaktion og iltudvikling.
The formation of bubbles is presumably because the equilibrium between dissolved and free oxygen cannot be maintained during the extreme, momentary reaction and oxygen release.
Da målemetoden forudsætter at ilten forbliver opløst i væsken, er det vigtigt at tage højde for dette - for eksempel ved at tilstræbe et forsøgsdesign, hvor den udviklede mængde ilt svarer til mindre end 80 % af iltmætningen.
Since the method requires the oxygen to remain dissolved in the liquid, it is important to take this into account, for instance by aiming for a test design in which the quantity of oxygen released corresponds to less than 80% of the oxygen saturation.
Alternativt kan mindre mængde enzym tilsættes, hvorved reaktionen vil forløbe langsommere.
Alternatively, a smaller quantity of enzyme can be added, whereby the reaction will run more slowly.
Det ses også, at problemet med ustabilt signal ved høj iltudvikling er mindre, sandsynligvis fordi iltudviklingen foregår over et længere tidsrum.
Here, the problem of an unstable signal when the release of oxygen is high is less significant, probably because the release takes place over a longer period of time.
Som det ses af Fig. 4, er der en direkte proportionalitet mellem tilsat mængde ureaperoxid og iltudvikling.
As can be seen in Fig. 4, there is direct proportionality between the quantity of urea peroxide added and the development of oxygen.
Dette skyldes, at brintperoxid kun udgør ca. 35 % af vægten i ureaperoxid-molekylet.
This is because hydrogen peroxide makes up only about 35% of the weight of the urea peroxide molecule.
Det vil sige, at der fra en ureaperoxid-opløsning kun kan frigives ca. 35 % af den iltmængde, der kan frigives fra en brintperoxid-opløsning af samme koncentration.
In other words, the quantity of oxygen that can be released from urea peroxide solution is only about 35% of the quantity of oxygen that can be released from a hydrogen peroxide solution in the same concentration.
3.2.1.1 Vurdering af den nedre detektionsgrænse for katalase bioassayet.
3.2.1.1 Assessing the lower detection limit for the catalase bioassay
Katalase bioassayet er meget følsomt, og det er muligt at måle præcist indhold af peroxid i en prøve, når iltudviklingen holdes mellem 1og 80 % af kammerets iltmætning.
The catalase bioassay is very sensitive, and it is possible to measure the precise quantity of peroxide in a sample, when oxygen development is kept between 1 and 80% of the chamber's oxygen saturation.
På baggrund af de testede tandblegeprodukter (afsnit 4) vides, at der maksimalt vil kunne opløses 10 g produkt i reaktionskammeret (100 ml).
On the basis of the tooth-whitening products tested (section 4), it was determined that a maximum of 10 grams of a product can be dissolved in the reaction chamber (100 ml).
Da ureaperoxid indeholder den laveste peroxid-mængde (vægtprocent), benyttes dette stof som baggrund for vurdering af nedre detektionsgrænse.
Since urea peroxide contains the lowest quantity of peroxide (% by weight), this substance was used as the basis of an assessment of the lower detection limit.
Som vist på Figur 4 vil tilsætning af 100 µl 1 % ureaperoxid til reaktionskammeret resultere i en iltudvikling svarende til 17,5 % af reaktionskammerets iltmætning.
As shown in Fig. 4, addition of 100 µl of 1% urea peroxide to the reaction chamber will result in an oxygen development in the reaction chamber corresponding to 17.5% of the chamber's oxygen.
Tilsætning af 100 µl 0,1 % vil derfor resultere i ca. 1,75 % af kammerets iltmætning.
Addition of 100 µl of 0.1% urea peroxide will therefore result in approximately 1.75% of the chamber's oxygen saturation.
Et nøjagtigt resultat kan imidlertid opnås ned til en iltmætning i reaktionskammeret på 1 %, hvilket vil svare til tilsætning af 10 g af et tandblegeprodukt med et indhold på 0,00057 % ureaperoxid.
An accurate result can however be achieved with an oxygen saturation in the reaction chamber as low as 1%, which would correspond to adding 10 grams of a tooth-whitening product with a content of 0.00057% urea peroxide.
For målinger af tandblegeprodukter er den nedre detektionsgrænse for katalase bioassayet således 0,0006 % ureaperoxid svarende til 0,0002 % brintperoxid.
For measurements of tooth-whitening products, the lower detection limit for the catalase bioassay is therefore 0.0006% urea peroxide, corresponding to 0.0002% hydrogen peroxide.
3.2.2 Test af katalasens stabilitet
3.2.2 Testing of the stability of the catalase
Enzymets stabilitet, pH-følsomhed og reaktionsevne blev kortlagt gennem en række forsøg.
The stability of the catalase, its pH sensitivity and reactivity were analysed in a series of tests.
I et første forsøg blev peroxid og katalase gentagne gange tilsat det samme reaktionskammer (Figur 5).
In the first test, peroxide and catalase were added repeatedly to the same reaction chamber (figure 5).
Figur 5: Gentagen tilsætning af H2O2 og ureaperoxid til reaktionskammeret.
Figure 5: Repeated addition of H2O2 and urea peroxide to the reaction chamber.
Pil 3 angiver tilsætning af 100 µl 1 % ureaperoxid, mens pil 4 angiver tilsætning af 50 µl 1 % brintperoxid.
Arrow 3 points to where 100 µl 1% urea peroxide were added, whereas arrow 4 shows where 50 µl 1% hydrogen peroxide were added.
Ved pil 5 afsluttes forsøget og elektroden overføres til en udgasset phosphat-buffer.
Arrow 5 shows where the test stopped and the probe was transferred to an out-gassed phosphate buffer.
Dette viser, at den første reaktion er forløbet til ende, og at der derfor har været tilstrækkelig mængde aktivt enzym i systemet.
This shows that the first reaction has proceeded to the end and that an adequate initial quantity of active enzyme was added to the system.
Ved tilsætning af 100 µl 1 % ureaperoxid (pil 3 + 4) stiger iltniveauet straks som forventet.
When 100 µl 1% urea peroxide (arrows 3 + 4) are added, the level of oxygen rises immediately as expected.
Dette demonstrerer, at den tilsatte mængde enzym fortsat er aktivt.
This shows that the quantity of enzyme added is still active.
Efter 9 min (1750 s) ses et kraftigt udslag i signalet, idet elektroden tages op af testkammeret og overføres til den udgassede buffer-opløsning, hvorefter signalet igen stabiliserer sig på ca. 1,5 % iltmætning.
After 9 min. (1,750 sec.), there is a marked change in the signal when the probe is removed from the test chamber and transferred to the out-gassed buffer solution, after which the signal stabilises once more at about 1.5% oxygen saturation
3.2.2.1 Test af varierende katalase koncentration
3.2.2.1 Testing of varying concentrations of catalase
Tilsætning af stigende mængder katalase øger hastigheden hvormed peroxiden spaltes til ilt og vand, men har ikke betydning for bestemmelsen af peroxidmængden (Figur 6).
Increasing concentrations of catalase, increases the speed at which the peroxide is split into oxygen and water but has no bearing on the determination of the content of peroxide (Fig. 6).
Figur 6: Katalase-koncentrationens betydning for reaktionshastigheden.
Figure 6: Effect of increasing concentrations of catalase on the reaction rate.
Der er tilsat henholdsvis 50 (1); 100 (2); 500 (3); 1250 (4); 5000 (5) og 10.000 (6)units katalase.
50 (1); 100 (2); 500 (3); 1,250 (4); 5,000 (5); and 10,000 (6) units of catalase were added.
Forsøgene vist i Figur 6 er udført i triplikat ved pH 7.
The tests shown in Fig. 6 were conducted in triplicate at pH 7.
Der er tilsat samme mængde (250 µl) 1 % ureaperoxid i alle forsøg.
The same quantity (250 µl) 1% urea peroxide was added in all tests.
Ved tilsætning af 10.000 katalase-units sker reaktionen omgående og al peroxid spaltes på mindre end ½ minut.
When 10,000 units of catalase are added, the reaction is immediate and all of the peroxide is split in less than half a minute.
Mindskes katalase-tilsætningen falder hastigheden.
If less catalase is added, the rate of the reaction drops.
Ved tilsætning af 1250 katalase-units spaltes al peroxid indenfor 1½ minut.
When 1,250 units of catalase are added, all of the peroxide is split within 1½ minutes.
Selv ved meget lave katalase-koncentrationer i kammeret forløber reaktionen dog til ende inden for 60 minutter og den endelige ilt-produktion er som forventet ens i alle forsøg.
Even at very low concentrations of catalase in the chamber, however, the reaction concludes within 60 minutes, and, as expected, the final oxygen production is the same in all of the tests.
Tilsvarende forløb er set for brintperoxid (data ikke vist).
The same was seen for hydrogen peroxide (data not shown).
Udfra disse data anbefales tilsætning af 5000 units pr. reaktion.
Based on this data, it is recommended that 5,000 units are added per reaction.
Som brugsopløsning anvendes en fortynding på 140.000 units/ml i 50 mM phosphat-buffer.
A stock solution of 140,000 units/ml was prepared by dilution in 50 mM phosphate buffer.
I et forsøg på at undersøge katalasen holdbarhed, blev en brugsopløsningen delt i to, hvoraf den ene halvdel blev opbevaret i køleskab ved 4 °C, mens den anden halvdel blev anbragt ved ca. 20 °C (stuetemperatur).
In an attempt to examine the shelf life of the catalase, a stock solution was divided in two parts, of which one was stored in a refrigerator at 4°C, while the other was stored at around 20°C (room temperature).
Efter 0, 1, 2, 3, 7, 50 døgn blev opsat standardforsøg ved pH = 7.
After 0, 1, 2, 3, 7, and 50 days, a standard test was set up at pH7.
Til kammeret blev tilsat katalase (5000 units) og 100 µl brintperoxid (1 %) svarende til en teoretisk iltudvikling på 0,5377 mg.
Catalase (5,000 units) was added to the chamber as well as 100 µl hydrogen peroxide (1%), corresponding to a theoretical oxygen development of 0.5377 mg.
Tabel 2: Katalasens holdbarhed i opløsning ved 4 °C og 20 °C
Table 2: Assessment of the shelf life of the catalase in a solution at 4 °C and 20 °C.
Holdbarheden af brugsopløsningen under opbevaring ved 4 °C og 20 °C er meget lang.
The shelf life of the solution when stored at 4°C and 20°C is very long.
Som det fremgår af Tabel 2, var katalase-aktiviteten således efter 50 døgns opbevaring ved 20 °C stadig på samme niveau som umiddelbart efter fremstilling af brugsopløsningen.
As can be seen from Table 2, the catalase's activity after 50 days storage at 20°C remained at the same level as just after the solution was prepared.
Selve reaktionshastigheden var heller ikke påvirket af den lange opbevaring (data ikke vist).
Furthermore, the reaction rate was not affected by the long storage time (data not shown).
3.2.2.3 Katalase-aktivitetens afhængighed af pH i bufferen
3.2.2.3 Effects of pH on the activity of the catalase
Frigivelse af peroxid fra releaseren calciumoxid er pH-afhængig.
The release of peroxide from the releaser calcium oxide is pH-dependent.
Det er derfor relevant at teste metodens pH følsomhed.
It is therefore relevant to test the method's sensitivity to changes in pH.