Chang
es in salivary composition corr
elat
e with dis
eas
esusc
eptibility, dis
eas
e stat
e, or both. How
ev
er, us
e ofsaliva for diagnostic purpos
es is complicat
ed by th
e gland-sp
ecific
eff
ects of circadian rhythm or diurnal variation.W
e r
ec
ently charact
eriz
ed a suit
e of p
eptid
es in th
e![](/imag<font color=)
es/
entiti
es/l
e.gif">10-kDa fraction of human parotid saliva that includ
edmany nov
el sp
eci
es. In this study, w
e us
ed nov
el iTRAQlab
eling ch
emistry to inv
estigat
e possibl
e diurnal
eff
ectson p
eptid
e g
en
eration. W
e coll
ect
ed sampl
es produc
edby gustatory stimulation as th
e ductal s
ecr
etions at fourtim
e points und
er conditions that minimiz
ed prot
eolysis,pool
ed th
em according to coll
ection tim
e, and isolat
ed th
eLMW fractions. Sampl
es coll
ect
ed at
each coll
ection tim
ew
er
e d
erivatiz
ed with a diff
er
ent isobaric iTRAQ r
eag
ent.Th
e lab
el
ed sampl
es w
er
e combin
ed, s
eparat
ed by r
ev
ers
ed-phas
e HPLC, co-spott
ed with matrix on MALDItarg
ets, and analyz
ed by MALDI TOF/TOF mass sp
ectrom
etry. With this approach, w
e achi
ev
ed r
elativ
e quantification of th
e parotid p
eptid
es at four tim
e points. Ins
ev
eral cas
es, abundanc
e during th
e day chang
ed dramatically. iTRAQ tagging improv
ed th
e effici
ency ofMS/MS fragm
entation, which in turn allow
ed th
e id
entification of s
ev
eral nov
el p
eptid
es. Our r
esults d
emonstrat
ed both th
e utility of this m
ethod and th
e importanc
eof diurnal
eff
ects on th
e composition of th
e human parotidsaliva p
eptidom
e.