Environmental Engineering Reference
In-Depth Information
were.measured.using.a.weight.and.height.scale.(RGZ-120-RT,.Mike.Balance.
Instrument. Factory,. Nanjing,. China). with. maximum. weight. of. 120.kg. and.
maximum.height.of.190.cm..Children.were.required.to.urinate.and.take.off.
shoes..All.work.was.carried.out.by.trained.study.members.
Collection and Analysis of Samples
Biologic. samples. were. collected. by. trained. nurses. from. both. groups. after.
they.had.fasted.for.12.h..Blood.samples.(2.mL).were.drawn.from.a.forearm.
vein.into.heparinized.glass.tubes.(with.polyethylene.stoppers).and.mixed.by.
gentle.inversion..Freshly.voided,.irst-morning.urine.samples.were.collected.
in. 15.mL. polyethylene. bottles. with. screw. caps.. The. urine. samples. were.
centrifuged.at.1000.rpm.for.5.min..Blood.and.urine.samples.were.stored.at.
−20°C.until.analysis.
All. the. glassware. and. plasticware. were. soaked. in. 20%. (v/v). HNO
3
. for.
24.h,. rinsed. with. ultrapure. water,. dried. and. stored. in. a. closed. polypro-
pylene. container.. High-quality. water,. obtained. using. a. Milli-Q. system.
(Millipore),.was.used.exclusively..Before.analysis,.the.frozen.blood.samples.
were.slowly.thawed.at.ambient.temperature,.homogenized.in.a.vortex.mixer.
and. diluted.1:9.with.0.5%.(v/v).HNO
3
.. Gloves.were. used.for.all. biological.
manipulations.. The. concentrations. of. Mn. in. blood. samples. were. deter-
mined. by. graphite. furnace. atomic. absorption. spectrometry. with. Zeeman.
background.correction.(Zeenit-650,.Analytik.Jena,.Germany)..The.method.
was.validated.before.use.by.standard.evaluation..A.correlation.coeficient.
of. at. least. 0.99. for. the. calibration. curve. was. considered. satisfactory.. The.
main. parameters. used. for. blood. Mn. determination. were. a. wavelength. of.
279.5.nm;. a. lamp. current. of. 4.mA;. a. slit. width. of. 0.2.nm;. drying. at. 90°C,.
105°C,.and.120°C;.ashing.at.850°C;.and.atomization.at.1700°C..The.detection.
limit.was.0.15.μg/L.
All. analyses. of. renal. biomarkers. were. performed. under. similar. experi-
mental. condition. in. the. same. laboratory.. Urinary. RBP. and. β
2
-MG. were.
measured.with.ELISA.(kit.from.Sun.Biotech.Company.of.Shanghai,.China).
methods.(Cui.et.al..2005)..Creatinine.concentrations.of.urine.were.measured.
using. the. picric. acid. method. (kit. from. Jiancheng. Bioengineering. Institute,.
Nanjing,.China)..All.urinary.parameter.assays.were.adjusted.for.creatinine.
in.urine..Urine.samples.with.creatinine.level.<0.3.or.>3.0.g/L.were.excluded.
from.the.analyses.(N.=.6).
Statistical Analysis
Nonparametric.analyses.were.used.for.data.with.skew.distributions,.and.chi-
square. tests. were. used. for. categorical. data.. Spearman. correlation. analysis.
was.conducted.to.evaluate.the.relationship.between.RBP,.β
2
-MG,.and.MnB..
The.level.of.signiicance.was.set.at.
P
.<.0.05.(two.tailed)..All.statistical.analyses.
were.performed.using.SPSS.version.13.0.software.(SPSS,.Chicago,.IL).
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