免费文献传递   相关文献

Lipid metabolomics in serum of hyperuricemic rats induced by fructose based on UPC2-Q/TOF-MS

基于UPC2-Q/TOF-MS技术的果糖诱导高尿酸血症大鼠血清脂质代谢组学研究



全 文 :2016
y

z à
41
áà

}
Vol41,No.6 March,2016
Ô°
UPC2Q/TOFMS
()ƒÄÅæçÆ]
¡Ç‡ˆ¡ÈW´P¶•„…
+7±
1,
;þk
2,
89ê
2,
:;
1,
+<
1,
‰M
1
(1`
aVWz?Z VzZY
,`
a
100102;2`
a…»?Z {ZY
,`
a
100102)

lm
] 
ÁiW´P¶•()

„…ÄÅæçÆ]¡Ç‡ˆ†Ê¸‡ˆ¡È$WdP¶‹ƒ¯_

§¨$†Æ]¡Ç
²ôƒÓ²°‹BV‹

@ò2‚=²KL

ƒ÷Ò

9nOs´L›i
(UPC2Q/TOFMS)
()‘›Æ]¡Ç‡ˆ

5
­•
)¨
ʸ‡ˆ

­\•

¡ÈP¶Lq

µi>~â;‘›€úÂÃ

•ƒP¶L±

§¨$±–P¶‹

<Äãÿ

5­•¨­\•ƒP¶Lqý²{㏱
,`
§¨$
11
±P¶‹

ÁiÀ‚´ö=<=8^´Lq¹H‚vúÌ°ƒà
]

£¤]

ö]

Fö]ö

?Ó²°‹BV‹

ñ^¿RúÔ°
UPC2Q/TOFMS
()—nÄÅæçÆ]¡Ç‡ˆ¡È
W´P¶•„…ƒ€ú

{uW¨]P¶±¸œÒ†Æ]¡Çƒm¢fùBô

òÆ]¡Çƒv}„´¨:T÷v&
Ôä


nop
] 
ÄÅ

Æ]¡Ç

W´P¶•

°‹BV‹

qr)(
] 20150803

vwAx
] 
‰M

]^s

ot„uVz?z°2bcÓ]^
,Tel:(010)86768292,Email:maqun99@163com

Ax€
] 
+7±

‚A]^7
,Email:1090361658@qqcom
Lipidmetabolomicsinserumofhyperuricemicratsinducedby
fructosebasedonUPC2Q/TOFMS
ZHANGShuli1,WANGYiping2,DUZhenxia2,XUEMeng1,ZHANGJiao1,MAQun1
(1SchoolofChineseMateriaMedica,BeijingUniversityofChineseMedicine,Beijing100102,China;
(2SchoolofScience,BeijingUniversityofChemicalTechnology,Beijing100102,China)
[Abstract] Toexplorethediferencesinlipidmetabolitesinserumofhyperuricemicratsinducedbyfructoseandnormalratsby
usinglipidmetabolomicstechnology,andscreenthepotentialbiomarkersrelatedtohyperuricemiaThemetabolicfingerprint
spectrumoftheseruminhyperuricemicrats(modelgroup)andnormalrats(controlgroup)wasobtainedandanalyzedbyusing
ultraperformanceconvergencechromatographytandemQtimeofflightmassspectrometry(UPC2Q/TOFMS)methodandthe
diferencesofmetabolicspectrabetweentwogroupswerecomparedviathemultivariatestatisticalmethodstoscreendiferential
metabolitesTheresultsindicatedthattherewassignificantdiferenceinmetabolicspectrabetweenmodelgroupandcontrol
group,and11diferentialmetaboliteswerescreenedTheneightpotentialbiomarkerssuchasarachidonicacid,palmiticacid,
oleicacidandlinoleicacidweretentativelyidentifiedbyusingtheexactmassnumberandsecondarymassspectrometry(MS/MS
spectrum)Therefore,anewresearchmethodforlipidmetabolomicsinserumofhyperuricemicratsinducedbyfructosewases
tablishedsuccessfulybasedonUPC2Q/TOFMSWhat′smore,itwasspeculatedthattheabnormalmetabolismoffatyacid
mightbeassociatedwiththepathogenesisofhyperuricemia,whichwouldprovidescientificbasisforearlydetectionandprevention
ofhyperuricemia
[Keywords] fructose;hyperuricemia;lipidmetabolomics;biomarkers
doi:10.4268/cjcmm20160627
·5311·
2016
y

z à
41
áà

}
Vol41,No.6 March,2016
  
EyìFG™Ð°•ªÝ¨ûÍ<^ƒÐ¯

Æ]¡Ç’“²ô»¢ƒm¢¬ˆ?_‰
[1]。
hòÇvOP8[U™@$«¢¬ÃƒP¶–»
¢

Æ]¡Ç²Mî`b2Z>‘Ö‡
[2]。

¢”„…Ì

WP¶‡d†P¶–»¢”ÅÆ
¢

#¢öDB²ô–
[34],
ó”

Æ]†¡WP¶‡
dðsý²§%ôõ
[58]。
W´P¶•
(lipido
mics)
hò7ÖüƒP¶•‘[

²W´P¶»¢
ƒ:T¨„´

W´°‹B.‹

,‹çèƒVvì
’,‹„mö€÷€q=úÖü—‡
[910]。
Æ]¡Ç¨¨@«¢¬ƒ‰†sÄÅû¸
ƒ©ø‰Bô
[1112]。
/‹(„…ˆÜ{

†“
|Åd£¤

ÄÅû͜2ZÌÍd/‹¡Æ]
ªÝRQø
[1316],
Õ1N(µi^Þ5€
ú
[17]。
N^ói¿Rƒ
UPC2Q/TOFMS
W´P¶
•€ú„…ÄÅæçÆ]¡Ç‡ˆ¡ÈP¶‹
ƒÐ¯

‚v“Ó²ƒ°‹BV‹

ì}òÆ]¡
ǃv}„´¨:Tp)Š.

1 
ʸ†€ú
11 
Áj†¥1
 
2‚=²KL

ƒ÷Ò9nO
s´L›iÁ
(UPC2Q/TOFMS,
†%
Waters
D
E
);ACQUITYUPC2TM HSSC18 SBKLN(30
mm×100mm,18μm);H-Q•Sf(GL21M,
bH¬ÎÁDE
)。
D
ÄÅ
(AMRESCO);
¡Æ]¥1µ

93¿
°‹gû„…³

o<
20150227);
à]

À]Ô

±
®Ê

KLZ

†%
ThermoFisherScientific
DE
);
»H

àÊ

²P:ò‘›Z

2Zª
(MiliQ

fÕù

†%
Milipore
DE
)。
12 
/‹‘•†5
 SD
‡ˆ



Ö
(240±
20)g,´
Õ23\@Á¹(/‹$S

/‹=þ
Ì<
SCXK(
3
)20120001。
SD
7–‡ˆ
16
÷

¬ó–Û¥
5d
~

[´ö
Ff‘ò­\•¨5­•



÷

Û·E@·
¸

¤O­\•û]•3ª

5­•û
10%
ÄŪ


•/‹:Õ7ûÍ

ûª

·¸¨ª|Ný©

]
,`
V\ûªö

5}s

|
7d
ɛœq¡[
]

ðÌ
21d。
13 
°_q5
 
q›œ¡
1mL,
[Æ]¥1µ
ù{ryh

14 
‚Nƒ:I”†
UPC2Q/TOFMS
‘›‹Œ
 
›œ¡
3mL,
°
4℃
Û
4000r·min-1
•S
15min,
qbȢ-
-20℃
ÝZ¥i

q
80μL¡È‰

15mLEP
$

‰
320μL»HàÊ(3∶1)³
ª

]

|]
15s,10000r·min-1
¸-•S
10
min,
qbȢ
10μL ¢´›iÁ—n‘›。
KL‹Œ
:ACQUITYUPC2TM HSSC18SBKLN
(30mm×100mm,18μm);®/² Aò CO2,®


ò
02%
à]
[
±®Ê

àÊ
(6∶1)

],
Â
šß±
,0~05min,99%A,05~9min,99% ~
60%A,9~10min,60%A,10~11min,60% ~99%
A,11~13min,99%A,
®Q
15mL·min-1;
N-
40℃。
´L‹Œ

µi
XevoG2SQTOFMS
´Lõ
â

ÒwOP
m/z50~1200,
•3_5£UwxU
•
(ESI),
µi¬•3

5£qu

ßÄUï
2500V,
•3>-š
120℃,
x_Û-š
450℃,
b
?Û®Q
50L·h-1。
15 
=.I”
 
Ái
MarkerlynxV41
錭Ð
l=.—n Ê¨ö[_I”

ói
SIMCAP
1303
éŒ
(UmctricsSweden)
­‚³—n‘•¨

Ê>

G7:8¥ú®_‘›
(OPLSDA)。
‘›
<Äì8\¨L\=‘q¨
Splot
qÜÿ

ì
VIP>1
ƒ‘hò­5­DB{ã2¦ÞƒBV
‹

•sÂõi

q(

§¨
P<005
ƒ=.=
rBdƒ¯ö

2 

21 
°_AB
 


NO
,2
•¡ÈÆ]2Ø
l$ë{㏱
(P<001)。
5
14,21d


•¡ÈÆ]2Ë{㏱
(P<005),
ù{²
(}s5­/v
,¨
O5
7d
ƒ=.—nW´P
¶•‘›

<ĝÜ
1。
Ü
1 
£¤5Os‡ˆ¡È
UA
¯_<ÄÂÃ
(珋x±s,
n=8)
Table1 ComparisonofUAchangesinserumofratswithdifer
entmodelingtime(珋x±s,n=8)
•_
7d 14d 21d
­13838±2354 14715±1142 12596±057

16964±25132) 17256±8492) 15678±21081)
  
 

†­\•²Â
1)P<005,2)P<001(
Ü

¤
)。
22 
¡È‚³ƒ
LCMS
‘›
 
Ái
ESI
¥35
£‘_µÀ­\•¨5­•‡ˆ¡È‚N=.

=
·6311·
!„Ùö

Ô°
UPC2Q/TOFMS
()ƒÄÅæçÆ]¡Ç‡ˆ¡ÈW´P¶•„…
r¡È‚Nƒ
BPI
q

q
1。
³BP¶‹²
13
min
8OÒ=r?$ƒ‘•

9

•ƒ{ã£
¤

òÌ=.ƒœ`–

²=.µÀòû$

|s
Ù

‚³

Xi´ø‚³—‚

]

ìGuµÀõ
âƒ/v–

q
1 
­\•
(A)、
5­•
(B)
²¬•35£Ûƒ
BPI
q
Fig1 BPIchromatogramsofthemodelandcontrolofserum
sampleobtainedfromthenegativeion
23 
=.‘›
 
µi
OPLSDA
­
Markerlynx
ç$
ƒ=.—n‘­

<ĝq
2,3。
x•¡È=.‘
›ƒ
Splot
qq
4,
5­•†­\•‘•{ã
,S
­ÐÈ`uƒ=.­‘_PÜ|‚N•œvš7
ƒ./•3

oÓ²°‹BV‹

q
2 2
•‡ˆ¡Èƒ
OPLSDA
8\=‘q
Fig2 2DscoreplotsofOPLSDAofratserumsamplefrom
diferentgroup
24 
°‹BV‹ƒmë†Vv
 
?
OPLSDA
‘
›
,¨
q
VIP>1
ƒ±¯ö

µi

q(­§¨ƒ
±¯ö²­\•†5­•—n(Ì

÷B
P<
005
ƒ¯öœhò†Æ]¡Ç‡ˆ²ôƒÓ²
°‹BV‹

*\Bô^Þ`î.[^

8^´Lv


Ái
HMDB(htp://wwwhmdbca/)¨ LIPID
MAPS(htp://wwwlipidmapsorg/)
ö=.4

ì
    
q
3 2
•‡ˆ¡Èƒ
OPLSDA
L\=‘q
Fig3 3DscoreplotsofOPLSDAofratserumsamplefrom
diferentgroup
q
4 
¬•35£Ûƒ
Splot
q
Fig4 SplotofOPLSDAfromthenegativeion
m/z3032774
ƒ•3òó

XiÀ‚²­‘3´ö
=.qJ

ñ@ò
ESI
¬5£Ûƒ8^Çπ£{
uP¶‹<^

Çπ£ò
:m/z3032774[M-
H]-,25956[M - CH2 O3 - H]
-,5893
[C2H3O2]
-,
q
5。

LIPIDMAPS
=.4
MS/
MS
Lq¨Ì°ƒà]­\³­Â
,m/z3032774
ƒ•3uVvòÌ°ƒà]

“|
10
?P¶‹Xi
dõ€úV_

Ü
2。
3 
õã†<ã
UPC2Q/TOFMS
()²‘•Ò@

‘›Os
’ÞCš€÷øý¢´›i
(LCMS)
B?‡
8>

‘›VvÒ@Š

ϑRQԥ

Ò÷=
àsý>=íƒP¶Lq
[18]。
“xÿ­W´‚
³—n8°_I”
[19],
6èBf²#×~—‚
‘›

™_úgh®û

¤OœRQכ$W´
ƒ<^

Õ1N(¨O
UPC2QTOF/MS
ƒW
´P¶•()

N(mëW¨]
FA(C16∶1),FA(C16∶0),
FA(C18∶1)
sö5­•°­\•
,FA(C20∶4),
FA(C24∶0),FA(C24∶1)
söH°­\•

·7311·
2016
y

z à
41
áà

}
Vol41,No.6 March,2016
    
q
5 
Ì°ƒà][^´Lq
(A)、
8^´Lq
(B)¨
­³8^´Lq
(C)
Fig5 MS(A)andMS/MSspectrum(B)ofarachidonicacid
andMS/MSspectrumofstandard(C)
W¨]ƒ=†Ái

‰Q
ATP
ƒ‘×

X
HUA
°
‰

˜êÆ°Æ]¡Ç
[20]。
]¨W¨]í
ò>éç÷—O¡ÇƒrùÕO
[21],
E“é£
¤]
(C16∶0)[6]。
r,£]¨W¨]Ì°ƒà]

dÌ°]d‹´

W´òt_‹ƒRQÆ°¨Ï¦

êÌ°ƒà]é8 Lఋ=ƒî‹´

 Là†ú>†ÇBô

àB¨@
[22]。
B„…Ü{
¡ÈW¨]²ÒÓfWP¶€÷Â
TG,TC
CD

²—ZÆ]¡ÇnJ

ÿ dqu¡ÈW¨]Ë
šƒ¯_

!v¶Ê¡ÈW¨]’WP¶‡d

Œ4
/œÐ‚‹_ƒm°
[23]。
N(<ÄÜ{W¨]
P¶±¸œÒ†Æ]¡Çƒm¢fùBô

˜W
´P¶•™šòÆ]¡Ç†P¶å=/ƒ²ô
–p)úŠ.

N„…ói
UPC2Q/TOFMS
ƒW´P¶•
€ú‘›ÄÅæçÆ]¡Ç‡ˆ†Ê¸‡
ˆ¡ÈƒWdP¶‹¯_
,`
­“—n>~â
;‘›

mëÆ]¡Ç‡ˆ5­¡È$W¨
]d‘m°¯_

ñ€úxÿ­‚³—n8
°_I”

gh®û™_

ÝrRQJ‘Æ]
¡Ç‡ˆ¨­\•‡ˆƒíƒ
,`
mëú1‘
Ó²°‹BV‹

ò˜P¶@Nƒ™š#$“
m¢fù’Æ]¡Çƒv}„´¨:T÷v
úÔä

Ü
2 
±P¶‹ƒV_<Ä
Table2 Identificationresultsofdiferentmetabolites
t/min m/z
‘3£ _£
VIP
P¶‹ &>

24361 2532366 FA(16∶1) C16H30O2 3275 £¤ö]1) ↑
24935 2792681 FA(18∶2) C18H32O2 7314 Fö]1) ↑
24938 3032774 FA(20∶4) C20H32O2 3804 Ì°ƒà]1) ↓
25493 2812822 FA(18∶1) C18H34O2 5890 ö]1) ↑
30719 3653908 FA(24∶1) C24H46O2 1247 83ƒà]1) ↓
30868 4294345 Unknown Unknown 2147 Unknown1) ↓
31698 3674101 FA(24∶0) C24H48O2 1827 83ƒ]2) ↓
35390 2552635 FA(16∶0) C16H32O2 3870 £¤]1) ↑
36554 2832939 FA(18∶0) C18H36O2 3122 ‹W]1) ↓
45999 3363651 Unknown Unknown 3090 Unknown1) ↓
54591 2010526 FA(11∶0) C11H22O3 3141 Unknown2) ↓

yg?z

[1] 
I.C

˱w

Æ]¡ÇTœ$óô ƒôSøù
[J]
$¹8‘%P¶-.
,2011,27(7):543
[2] GoncalvesJP,OliveiraA,SeveroM,etalCrosssectionaland
longitudinalassociationsbetweenserumuricacidandmetabolic
syndrome[J]Endocrine,2012,41(3):450
[3] NicholsonJK,ConnelyJ,LindonJC,etalMetabonomics:a
platformforstudyingdrugtoxicityandgenefunction[J]NatRev
DrugDiscov,2002,2:153
[4] FernandisAZ,WenkMR,ChromatogrJ,etalTargetinglipid
·8311·
!„Ùö

Ô°
UPC2Q/TOFMS
()ƒÄÅæçÆ]¡Ç‡ˆ¡ÈW´P¶•„…
metabolismtostarveandstopcancercels[J]EurJLipidSci
Technol,2009,877:905
[5] FiehnOCombininggenomics,metabolomeanalysis,andbio
chemicalmodelingtounderstandmetabolicnetworks[J]Comp
FunctGenom,2001,2(3):155
[6] 
s^©

s.$

Í+Ù

ö

Ô°¢²KL
´
L()ƒ
Æ]¡Ç¡¥P¶•„…
[J]
Ö|A
,2013(2):176
[7] 
ø,

$Ïy™@Æ]¡Ç’¸´d­ƒP¶•
./
[D]
4x

4x$A,‡
,2013
[8] 
²ÇE

Æ]¡Ç¨¡W‡d­P¶å=/ƒµ¶
[J]
59%TA,
,2006,8(3):194
[9] 
6yÑ

7

~K—

W´•²A,„…$ƒói
[J]
ÄÅ°‹
,2007,29(2):169
[10] HanA,GrossRWGlobalanalysesofcelularlipidomesdirectly
fromcrudeextractsofbiologicalsamplesbyESImassspectrome
try:abridgetolipidomics[J]JLipidRes,2003,44(6):
1071
[11] ChoiHK,MountDB,ReginatioAM,etalPathogenesisof
gout[J]AnnInternMed,2005,143:499
[12] ChoiJW,FordES,GaoX,etalSugarsweetenedsoftdrinks,
dietsoftdrinks,andserumuricacidlevel,thethirdnational
healthandnutritionexaminationsurvey[J]ArthritisRheum,
2008,59(1):109
[13] NakagawaT,TutleKR,ShortRA,etalHypothesis:fruc
toseincludedhyperuricemiaasacausalmechanismfortheepi
demicofthemetabolicsyndrome[J]NatClinPractNephrol,
2005,1:80
[14] TappyL,LeKAMetabolicefectsoffructoseandtheworldwide
increaseinobesity[J]PhysiolRev,2010,90(1):23
[15] PerezPozoSE,ScholdJ,NakagawaT,etalExcessivefructose
intakeinducesthefeaturesofmetabolicsyndromeinhealthyadult
men:roleofuricacidinthehypertensiveresponse[J]IntJ
Obes,2010,34(3):454
[16] NakagawaT,HuH,ZharikovS,etalAcausalroleforuric
acidinfructoseinducedmetabolicsyndrome[J]AmJPhysiol
RenalPhysiol,2006,290(3):625
[17] MachmoundMF,EiNagarM,EiBassossyHMAntiinflam
matoryefectofatorvastatinonvascularreactivityandinsulinre
sistanceinfructosefedrats[J]ArchPharmRes,2012,35
(1):155
[18] 
²¡œ

˛

é|L

ö

2‚=²KL()²$,„…
¨´öøù$ƒói
[J]
Çv&()
———
$A,ëP
_
,2014,16(3):534
[19] RaoGVN,GnanadevG,BellamR,etalSupercriticalfluid
(carbondioxide)basedultraperformanceconvergencechroma
tographyfortheseparationanddeterminationoffulvestrantdiaste
reomers[J]AnalMethods,2013,18(5):4832
[20] 
£ÒÑ

°Ä

i8&
[M]
23

™4°$()

2009:2766
[21] DandonaP,AljadaA,BandyopadhyayAInflammation:thelink
betweeninsulinresistence,obesityanddiabetes[J]TrendsImmu
nol,2004,25(1):47
[22] HeH,RenX,WangX,etalTherapeuticefectofYunnan
Baiyaoonrheumatoidarthritiswaspartialyduetoregulating
arachidonicacidmetabolisminosteoblasts[J]JPharmBiomed
Anal,2012,59:30
[23] 
ڕ

ÍÊ,

ÍÁ†

¡ÈO•W¨]­Æ]¡ÇP¶
ƒµ¶
[J]
Ù5A,
,2006,46(31):53

{|-0
 
}~~

·9311·