From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
Carbohydrates
of Human
By
T
HE
LACK
monomers
study
on the
E.
E.
on the
platelets
elements
of blood.
The
chemically
fractionated
a glycoprotein
fraction2
determination
components
and
the
soluble
and
)
carbohydrate
KOCHOLATY
has
Platelets
and
their
has prompted
units
and their
formed
various
fraction,’
fractions
\V.
polysaccharides
and bovine
monosaccharide
-insoluble
Bovine
AND
WOOD5IDE
of information
in human
total
bound
and
of
the
acid
cent
( an
5 per
revealed
the
corresponding
this comparative
derivatives
in these
monosaccharides
mucopolysaccharide
trichloroacetic
exiStellce
of
in
acid-
numerous
moieties.
METHODS
Source
and
Human
tional
ml.
Preparation
blood
Red
of
an
number
to the
laboratory.
veins
of
The
In
blood
were
suspended
centrifugation
of
platelets
10
o
per
(
A
immediately
or
at the
abattoir
Na-
110
B.
packed
the
In
to
a
of
from
the
120
limited
in ice
transportation
so as to
the
blood
slashed
neck
mg.
per
were
following
were
arabinose,
N. Y., and
the
used
rhamnose,
gaiactosamine-HC1
Ohio;
galacturonic
distilled
at
in
and
iumman
2.0
ml.
of
the
100
C.
in
tared
was
cells
and
6
for
the
2
the
suspended
desired
time
x
3
to
100
suspensions
the
at
37
from
platelets
were
The
total
the
ranged
dishes.
from
cell
after
solutions,
aluminum
original
medium
hours
of
stock
platelet
subtracted
the
centrifugation
within
from
stock
from
preservation
aliquots
as
and
obtained
differential
out
to
Kocholaty
was
platelet
incubated
for
with
by
the
carried
referred
by
plasma
experiments,
tubes
milliliter)
the
glucuronolactone,
Biochemistry
continuous
5 Gm.
NaCl
carbohydrates,
for
both
mannose,
were
acid
obtained
the
analytical
gaiactose,
dried
theoretical
dry
weight,
and
counts.
C. P., from
accepted
to the
and
generous
0.85
Gm.,
U.
Mann
S.
from
Pfanstiehl
technic
and
paper
levulose
and
glucose,
obtained
from
Nutritional
and
giucosamine-HC1,
Dicision,
Knox,
Ky.
Submitted
Oct. 23, 1959;
#{149}Theauthors
are grateful
fGlucose
)
correlated
C. P. grade
Ill.,
for
or
outlined
Standards
Waukegan,
From
mg.
or
be
subsequently
One
that
obtained
gelatin,
could
n-milliliter
±0.2
values
Platelets
cent
appropriate
cells.
( 8.5
NaCl
corrected
xylose,
into
bovine
Carbohydrate
ml.
American
contained
formula
platelet-rich
incubation
suspensions,
weight
amount
2 per
prolonged
platelets
for
constant
Cross
customary
(luring
was
bovine
determinations
For
platelet
x
2
glycogen
used
and
collection.
saline,
distributed
The
milliliter
ter,
was
collected
procedure
after
either
blood.
were
4 C.
and
land,
of
bottles
N.I.H.
blood
was
use
glass
constituents
human
2 hours
in
Thus,
collection
The
collected
blood
experiments,
within
Tullis.4f
these
evacuated
solution,
platelet
bovine
the
with
standard
freshly
of
Fresh
a few
whole
to
the
breakdown
differential
1 to
phlebotomy
cattle.
Horn.3
per
by
The
anticoagulant
of experiments,
enzymatic
or
collected
equipment.#{176}
acid-citrate-dextrose
inhibit
of
was
Cross
of Platelets
Army
Biochemicais
Eastman
Organic
Research
sodium
of
human
acetate
0.2
water.
1173
Company,
chromatography:
glycogen.
Fucose
Corporation,
Chemicals,
Laboratories,
ZtIedical
for publication
Jan.
Louisville
Chapter
supply
Chemical
Research
New
CieveRoches-
York,
N. Y.
Laboratory,
28, 1960.
of the American
Fort
National
Red
blood.
Gm.
and
gelatin
2.0
Gm.
per
100
From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
1 174
WOODSIDE
Chew
ical
iOt(1l
Deterni
motions
IlexOses,
taneous
)(‘fltOs(’5
OS
of
glucose
The
individual
the
corresponding
hexoses
0.5
was
pentoses
from
paper
Itfl(l
of
\lokrascha
and
as
nlethylpclltoses
are
chromatograms
total
reported
of
tile
5jul01-
arc
equivalents.4
as
equivalents
and
of
methylpentoses)
platelet
stock
the
glycogen
rihose
( pentosc
pentose
(lilutiOll
for
Aldohexoses
used.
and
total ilexoSe
afl(i
1.0 mlll. of 1 /20
For
1111.
technic
pentoses
and
elided
sugars.
detenninations,
an(l
equivalents
sugars
KOCHOLATY
Carbohydrates
?Hethylk’fltO.Se.S.-Tlle
(111(1
detenllination
reported
of
AND
solutions
were
used.
Glucosanzine
all(1
glucosamine
and
solutions
was
neutralized
galactosaminc
determination
galactosamine
used
with
for
of
the
3 N
Tracey7
3 N HC1
NaOH
and
-Tile
was
hydrolysis.
diluted
One
distilled
detern#{252}nation
1111.
hydrolyzed
Tile
with
silllLlltaneous
elllployed.
of
to
a
total
Only
0.2
1111.
of
the
platelet
stock
or
fractions
volume
One and 2.0 ml. aliquots
were
removed
for
the
chemical
determinations.
Sialic
acid determination-The
diphenylamine
reaction
as modified
used.
The
alpha-i
glycoprotein#{176} containing
10 per
cent
sialic
acid
standard.
stock
platelets
water
suspensions
were
were
of
by
\Vinzler2
was
used
needed
for
of
platelet
tile
6.0
ml.
was
as
these
the
deter-
mninations.
Glucuronic
color
mannose,
of
as
platelet
determinations-Only
(Wj(l
reaction
glucuronic
described
Fractionation
wit/i
rpm
employed.
sulfuric
Aliquots
of
test
was
acid
in
0.25
employed.
the
1111. afl(1
The
presence
0.50
of
ml.
stock
Procedures
.5 per
cent
to
triclmloroacetie
a(ided
was
stand
g )
for
42
filter
at
5
room
per
\Vhatman
with
agitation
temperature
for
minutes
at
paper
( TC4).-’l’o
(lei(l
(lropwise
4
and
C.
tile
1.0
10
The
I .0
mill.
ml.
of
tile
sonoratedi
per
cent
and
minutes
supernatant
precipitate
of
10
then
was
solution
resuspended
TCA.
centrifuged
in
2.0
filtered
of 5
TCA and recentrifuged.
This
procedure
was
repeated
two
more
timnes
with
the
use of 3.0 ml. of TCA,
tile final volume
of the TCA soluble
filtrate
being
10 ml. Aliquots
of 0.5 mlii. and
1 .0 ml.
of the
TCA
soluble
filtrate
were
analyzed
for total hexose
and
pentose.
Tile remainder
of the filtrate
was
lyophilized
and
hydrolyzed
according
to tile
procedure
of Glegg
and Eidinger’
for paper
chromatography.
The
TCA
insoluble
residue
was
extracted
twice
with
10 ml. portions
of
etilyl
ether
( to remove
residual
TCA ) and
ilydrolyzed
with 2 N H.,S04
at 100 C. for 4 hours
in sealed
tubes
prior
to paper
chromathrough
no.
and
used.
suspension
was allowed
( 510 x
chemical
thioglycolic
Va5
Fractionation
stock platelet
The mixture
2000
with
Dische,5
were
Polysaccharide
at
by
suspensions
a qualitative
acid
ml.
cent
tography.
Glycogen
determination-Initially,
giycogeu
extraction
technic
described
by the 30 per cent KOH
cedure
extracted
initial
into
approximately
volume
tubes
Sero,nucoid
to
of
Harvard
Platelet
cent.
platelet
37
Treatment
glycoprotein
Medical
were
for
5
according
Glycogen
Scllaffer
of
glycogen
or(Ier
to
also
and
Kirk.’#{176} The
and
required
only
half
aliquots
were
pipetted
experiments,
in
tile
TCA
determined
to
was
avOi(1
clumping
latter
pro-
associated
the
witil
C.
by
outlined
was
Winzler2
applied
to
1.0
011.
suspensions.
procedure
suspensions
alpha-i
suspensions
water.
determined
Roe.”
incubation
incubation
or
( F-I ) fraction
Medicine,
amount
technic
stock
platelet
#{176}A
sanlple
95 per
4
fraction-The
stock
mnucopolysaccharide
tap
0 hours
either
tile
prolonged
determination-The
Mucopolysaccharide
of
the
at
of sonorated
applied
In the
at
incubation
portions
twice
of platelets.
centrifuge
prolonged
was
by Carroll,
Longley
and
extraction
technic
of Fong,
(200
and
the
was
School,
outlined
to
900
84
treate(i
with
niinutes
at
per
kindly
Boston,
a
mg.
by
dry
cent
Giegg
and
weight
of
Eidinger1
platelets).
ethanol-insoluble
supplied
by
Dr.
K.
fraction
Schmid,
was
The
acid
( F-I!)
Department
Mass.
Raytheon
resonance
10
resulted
KC
oscillator
in
a
platelet
cooled
by
(lisintegration
running
of
From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
C1RBOHYDIlATES
were
OF
hydrolyzed
and
the
HUMAN
i)y
doted
tile
ANI)
cation
BOVINE
excilange
hexosamines
were
1175
PLArELI;1s
resin
(lrie(i
1 The
technic.
in
40
tt
VdclI()
hydrolyzed
C.
for
nlonosaccharides
paper
cilrOmatographic
analysis.
Paper
Chroinatographic
The
ascending
employed.
it
Due
was
to
necessary
charides,
tile
by
(leproteinized
to be
fractions
material
obtained
was
tilen
i)e
tile
spray
by
prepare(l
for
the
cation
for
4 hours
correlation
of the
dry
values
for both human
of 21
separate
12.82
1 x
±
10
Witil
their
dry
standard
approximately
basis.
The
mental
work
Although
of the
been
Campbell,
to hydrolysis.
It is possible
but
complex,
diffusioml
to
platelets
were
glycogen
extracted
of the
total
chromatograpily
tions
)
,
the
the
above
lets.
acids
10’’
hydrolyzed
Partridge.’’
Ofl
bovine
Platelets
was
weight
on
the
in
for
given
would
Platelets
humall
(Iry
a
average
dry
Tile average
platelets
preseilted
Lopilato
and
appear
a dry
weight
following
basis
from
Danlesilek’4
levulose
tilat
as
tile
this
experi-
tile
average
does
not
washings.
the
findings
of
\Vagner,’
human
found
to contain
by the KOH
content
aldohexose
monomers,
would
in mammalian
human
glycogen
identified
bovine
and
chromatography
were
glucose,
sialic
acid
further
tissues.
of
identified
mannose,
was
demonstrate
also
fucose
found
the
and
in both
as
rihose.
human
fact,
cent
hydrolysis
extracindicate
contain
such
experimental
bovine
platelets,
galactosamine,
In
and
(IistribiItion’2#{176}
re-
bovine
in
per
would
and
acid,
be
be
KOH
finding
human
and
wi(Ie
well
which
could
under
the
: glucuronic
may
Upon
TCA
This
both
as
platelets.
glucose.
was
whicil
of glycogen.
for nearly
50
( both
fractions
demonstrated
a polysaccilaride-
1)lcttelets
amounts
accounted
and
in
ester
of other
polysaccharides
or galactose
had
occurred
monomers
galactose,
considerable
procedure
in both
of the
exist
phosphate
free
ilave
ketohexose
could
not
bovine
I)ltttelets
subsequent
or
repeated
hydrolysis
This
than
the
only
following
cosamine,
not
fractions
C. Tile
count,
obtained.
boville
human
platelets,
in either
humall
merely
that
no co-precipitation
aldohexoses
as mannose
conditions.
Upon
1 x
during
hexose
and
the
found
dry weight
determinations
mug. The
above
values
are
Thus,
values
levulose
in
ideiltified
by
Contrary
of
these
of
I)iatelet
were
for
smaller
Small,
of
moved
proce(lure
was
some
were
100
at
readily
DiscussioN
mean.
calculated
dimnen-
above.
the presence
chromatographically
1)rotein
AND
and
Consequently,
tubes
monosactwo
hexosanlines
fractionation’
glass
the
for
residues
resin.
seale(l
in
of 11 separate
20.64
±
.79
one-tllird
have
exchange
or
phthalate,12
used
Eidingcr
platelets,
Two
one
hydrogen
insohmi)le
i)y
carbohydrate
mentioned
and
either
m was
in
only.
by
reagent
TCA
determinations
errors
various
spray
present
dimension
aniline
weight
with
the
and bovine
platelets
.58 mg. The
average
llunlan
Platelets
was
to be
values
weight
first
Rcitc’
and
be
to
resolved
chronlatography
RESULTS
By
weight
the
reagent,
Glegg
Bartz
found
in
The
tile
Hamnerniami,
cOI1)leteiy
platelets,
in
2 N H2S04
%Vitil
not
\Vilole
hydrolyzed
hydrolyzed
of
lllOflosaccilari(leS
ionoiiiers.
two
Partridge.’
completely
were
tileSe
of
chronlatogram
could
however,
between
described
paper
ribose,
chroillatography;
technic
number
lilllited
run
and
differentiated
that
chromatographic
the
to
fucose
sional
Procedure
paper
glu-
addition
to
the
bovine
plate-
of
sialic
the
From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
1176
WOODSIDE
The
glucuronic
acid
spot
identified
spot on the paper
platelets.
Elution
analysis,8
glucuronic
for glucuronic
acid.
The
such
these
of the
as iduronic,
acids
were
overlapping
mannuronic
undertaken
components
tent
8.8
of
human
per cent
Determination
of the
fractionations
( table
lets
per
cent
of
bovine
acid,
the
approximated
present.
platelets,
The
significant
water
total
bound
was
Chromatography
in platelets
between
revealed
chemical
of only
acids,
identify
carbohydrate
calculated
human
and
TCA-soluble
fraction
of human
platelets
hexose,
whereas
it accounted
for 68 per
platelets.
and
the presence
other
uronic
content
of human
or
of the individual
carbo-
distribution
differences
another
unand bovine
No attempts
to
solvent
systems.
respectively,
( table 1).
and pentose
dry weight
of the hexose
2 ) . The
of the total
distilled
total
carbohydrate
by the addition
to be
bovine
revealed
with
KOcHOLATY
with
human
acids
revealed
would
not exclude
or guluronic
acids.21’22
in the above-mentioned
found
and
contaminated
of hydrolyzed
spots
and galacturonic
unidentified
spot
Excluding
glucuronic
bovine
platelets
can be
hydrate
was invariably
chromatograms
AND
the
con-
as
8.5 and
by
various
bovine
plate-
accounted
cent of the
presence
of
for 54
hexose
ribose
and
only.
glucose
Chromatography
with
traces
glucose
was
tion
which
able
that
of the
of
ribose
found
to be
could
not
react
all
glyccgen
in
some
the
with
of the
fraction
revealed
instances.
only
By
the
presence
chromatographic
aldohexo3e
present
the
anthrcne
reagent.
glycogen
was
precipitated
in
of glucose
identification,
the
TCA
Therefore,
soluble
frac-
it seemed
upon
addition
prob-
of
alcohol
( 5:1
ratio).
Upon
increasing
the alcohol
concentration
to 10:1,
no additional
glycogen
could
be precipitated.
In view of the finding
that KOH
extraction
of
the glycogen
was approximately
twice
that of the TCA
method
( 30 per cent
KOH
would
destroy
free glucose
and its oligosaccharides
) , it appeared
that
the TCA
extraction
platelet
suspensions.
procedure
effected
The
seromucoid
fraction,
which
components
of different
electrophoretic
of the
total
total
hexose
of human
hexose
content.
Table
1.-Chemical
With
only
could
platelets,
respect
Analysis
No. of
exper.
and
Pentose
nlethylpentose
Hexose-total
for
extraction
23 per
cent
the
Present
bovine
cent
TCA
in Platelets
Bovine
of
of the
5 per
platelets
%
from
least
three
carbohydrate
accounted
for 46 per cent
platelets,
Carbohydrates
an
glyccgen
at
only
human
of the
Hum
Carbohydrate
fraction
contain
mobilities,21
but
to
partial
No.
platelets
%
of
exper.
dry
of
weight
dry
weight5
16
0.67
±
.11
17
0.8i
±
.11
16
4.41
±
.28
17
4.69
±
.31
.32
Glycogen-TCAf
extraction
6
1.16
±
.17
6
1.i6
±
Glycogen-KOHf
extraction
6
2.34
±
.17
6
2.26
±
.36
Glucosamine
10
0.10
±
.04
16
0.63
±
.07
Galactosamine
10
2.57
±
.i6
i6
1.99
±
.14
Sialic
14
0.72
±
.05
17
0.65
±
.04
U
acid
Including
fTrichloroacetic
fPotassium
the
standard
acid.
hydroxide.
error
of
the
mean.
From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
CARBOHYDRATES
OF
Table
HUMAN
AND
2.-Distribution
of the Total
Carbohydrate
Fractionated
Platelets#{176}
-
Human
Carbohydrate
fraction
No. of
exper.
Total
carbohydrate
5% TCAt-soluble
fraction
Glycogen-TCA
extraction
Seromucoid
are
expressed
tTrichloroacetic
residue
TCA
mucoid
the
.25
14
0.69
±
.09
3.52
±
15
0.34
± .08
1.97
± .20
13
0.36
± .07
2.40
± .29
16
0.08
± .02
0.77
± .12
0.30
±
1.66
±
of
dr’
determinations
platelet
TCA-insolubie
corresponding
above
values
( table
glycogen
could
homogenization
found
platelets
sonorated
that
the
could
this
13
0.03
± .007
0.49
±
9
0.15
±
0.81
± .H
standard
error
not
of
the
.01
.05
mean.
KOHt
medium
extraction
extraction
hydroxide
acid
by
the
human
cells and muscle
TCA.
Following
was
technic
technic
Glycogen,
platelet
)
in this
discrepancies
TCA
extraction
proand bovine
platelets.
to extract
conKOH
procedure.
that
all of the
by employing
proper
their
procedures,
it
from
values
extracted
sonorated
for
as Influenced
whole
or
with
KOH.
by
Human
TCA
TCA/KOH
extraction
300
.071
8.64
of Carroll
Fomig
et al.2a
et
al.2
platelets
glycogen
KOHt
extraction
548
15.1
as
resi-
Medium
Ratio:
of
experiments).
F-li,
extracted
glycogen
same
when
-
extraction
and
10 per cent
TCA
muscle
tilan
the
Roe#{176}maintained
.989
extraction
of
basis.
Fractionation
procedure
revealed
pg.
-
of
the human
( methylpentose
( 100 pg. )
98.9
-
F-I
or
glycogen
100
sero-
co-precipitation
series
hexose,
fucose
extracted
TCA
#{176}Trichloroacetic
liver
with
Suspending
Suspending
mllediulll
5 or
skeletal
and
in Extractable
7.05
to
considerable
the KOH
in sonorated
more
standard
cent
precipitated
residues
yielded
manand
bovine
platelets
in this
determinations,
cent
the
98.9
due
namely
platelets.
However,
essentially
the
3.-Variations
gelatin
be
included
fractions,
be extracted
from
and
re-extraction
g.
to the
fraction
on a dry weight
by the Glegg
and Eidinger’
by
chemical
proteins.
amount
of
twice
as much
have
shown
from liver and
Carroll,
Longley
Glycogen
since
similar
monosaccharides
approximately
9 per
attributed
could
not
the
either
observed
whole
remained
fraction
of tile TCA-insoluble
fucose
in both
human
glycogen
45 to 55 per
be
platelet
were
obtained
1 ) were
seromucoid
difference
the
twice
contained
mentioned
than
from
platelets
Table
Potassium
the
.43
.38
of the sugars
revealed
to bovine
platelets,
polysaccharide
Some
investigatorst242a
siderably
less glycogen
On the other
hand,
Saline
2%
Tullis
including
essence,
seromucoid
residue
bovine
between
cedures
.11
weight
and chromatography
galactose
and
to containing
fraction
also
was
±
with
( hexosamine
also.
In the
Hexoses
3.63
however,
In addition
seromucoid
due
and
methylpento.e
.11
residue
fraction
Hydrolysis
nose,
glucose,
the
exper.
±
in
insoluble
fraction;
of the
platelet.
Pentose
0.60
%
was,
seromucoid
tile
Hexosea
of
17
and chromatography
fractions.
\Vith
respect
tile
Bovine
No.
acid.
illSoluble
analysis
in both
of
as
in Chemically
plateleta
Pentose
and
methylpentose
fraction
#{149}AU
values
1177
PLATELETS
BOVINE
per
ml.
Ratio:
TCA/KOH
.548
553
.027
542
.016
From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
1178
WOODSIDE
In addition,
the
obtained
for
KOH
Probably
the
tion of glycogen
present
seen
in the
that
values
sonorated
TCA-soluble
by the
and
insoluble
per
cent
in 0.85
TCA
as well
recovery
( tile
pension
where
of the
30
per
found
to be
related
to the
( table
4 ) . Sonoration
metabolism
of the
suspensions
from
were
the
the
blood
higher
within
2 hours
of platelets
implying
that
could
exert
was
either
time
intervals
of time
revealed
total
hexose
obtained
more
plasma
biochemical
and
when
for
Thus,
it is evident
of the
Table
4.-Variations
that
other
both
one
to
human
Sound-treated
in
1 (lay
of
greater
platelet
found
that
and
Glycogen
blood
than
of
Contentsof
the
Human
saline,
or
catabolism
bovine
platelets
described
governing
in
blood,
both,
of the
in
loss
listed
table
in
4.
of total
Platelets#{176}
Ratio:
prolonged
incubation:
0 hr.
control
Total
hexose
-
Glycogent
Hexose
Glycogen
wilole
lfl
blood
plus
0
2.32
0.98
at
4 C.
24
2.36
0.89
saline
02
0 91
platelets
1 day
in
i day
Whole
3
collection
platelets
1 day
Whole
point
and of treatment
major
sources
it was
increased
previously
factors
and
the
allowed
for
since
whole
elements
in an
those
Hours
incubation
4 C.
treatment
sus-
the
at 4 C. in saline
did not result
were
left suspended
in whole
appear
that
the carbohydrate
-
Preliminary
gelatin
to
between
in whole
important
in TotalHexose
was
in
glycogen
values.
Even
more
the platelets
were
removed
formed
the
similar
cent
Furthermore,
resulting
platelet
carbohydrates.
The
total
hexose
values
table
5 varied
in a manner
re-
glycogen
in the glycogen
and
4 ) . These
variations
presumably
of platelets
accelerated
influences
be
glycogen
resulted
gelatin
of collection
some
of the
collection.
or the
3 it can
when
per
the
variations
( table
prolonged
incubation
of isolated
whole
platelets
in as low values
as obtained
when
tile platelets
blood
for extended
periods
of time.
It would
metabolism
those
completely
of the
procedure
glycogen-2
of platelets
carbohydrates
after
table
be
However,
degrades
large
encountered
varying
higher
values
maintained
striking
than
is negligible).
and isolation
of platelets.
The influence
prior
to carbohydrate
determination
variation
endogenous
From
could
7 per cent
the KOH
the
bonding
higher
factor
preventing
total
precipitanature
and amount
of proteins
procedure.
likely
KOCHOLATY
procedure.
solutions
from
most
At the beginning
of the investigations,
hexose
values
of human
platelets
were
were
cent
TCA
fractions.6’27
gelatin
only
In contrast,
KOH
polysaccharide-protein
the
the
KOH
glycogen
cent
50 per
by
saline
as the
was suspended
in 2 per cent
recovered
by the TCA
procedure.
complete
approximately
extracted
most
important
contributing
from
TCA
solutions
was
glycogen
covered
were
platelets
AND
whole
in
blood
plus
0
3.08
1.19
at
4 C.
16
3.10
1.05
0
5.88
2.44
16
5.09
1.51
saline
88
platelets
whole
hours
in
plus
6 hours
#{176}Values
fGlycogen
blood
in
expressed
extracted
saline
as
per
by
cent
the
of
procedure
dry
weight.
of
Fong
et
al.26
.8
.63
From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
CARBOHYDRATES
Table
OF
HUMAN
of Storage
5.-Effect
Human
2
p
s-os
#{149}
at 4 C.
I.E
on
the
2
n
ca
o
a)
E
2.58 ± .16
0.68 ± .16
6
7
8.95±.29
0.38±.14
hours
6
hours
3
2
5.42±.47
0.51±.21
rep orted
as well
blood),
the
5)
3.66
total
If this
.
cent
to 0.68
0.38
was
precipitated
-
-
6
their
exposure
5
correspon
ding
5.40±.06
case,
0.73±07
an
variation
platelets
cent
to whole
on
be
upper
limit
by
losses
for
or double
human
of
blood.
platelets
These
Wagner,’5
still
to
glycogen
highest
and
mean.
on human
in whole
attributable
the
the
the
leukocyte
glycogen
by
from
leukocytes
isolated
as reported
in ribose
content14’25
upon prolonged
storage.14
for
error
glycogen
determinations
after
collection
( 3 hours
possibly
blood
eta ndard
of platelets
blood,
may
procedure
varying
by
of Glegg
the
63
per
glycogen.
alcohol
cent
in
from
would
value
table
ob-
5 can
leakage
variations
0.64
not
platelets
content
glycogen
and/or
These
of
to 0.73
be
ribose
ranged
per
cent
Fraction
also
fucose,
human
and
contained
Cytochemical
polysaccharides29’3#{176}
two
)
,
distinct
Fraction
contained
( F-Il
II
to eliminate
6) These
acid,
human
ribose
bovine
the
which
fractions
( F-I )
I
acid
,
which
mucopolysac-
contained
and
and
has
for
the
been
sialic
one
( table
presence
presence
confirm
acid
galactose,
platelets.
were
In addition,
presented
and
fractions
the possible
results
would
galactosamine,
and
bovine
platelets.
galactosamine
evidence
Identification
of galactosamine
sence
of glucosamine
in F-I
.
Eidinger,1
or
more
composed
of aldohexoses,
methylpentoses,
hexosamine
and a
amount
of protein,
was precipitated
by 84 per cent ethanol.
Apthis fractionation
procedure
to human
and
bovine
platelets
re-
mannose,
of both
and
concentration.
ethanol,
the presence
of glucuronic
in F-I fractions
of both
galactose,
tend
(table
wi th
of whole
the
per
by
and
polysaccharides
considerable
plication
of
platelets
-
with
the studies
loss of the glycogen
of whole
fractionation
obtained
tions
l.a
0.64 ± .41
platelets.
were
vealed
ribose
E___
2.47 ± .11
of the total
glycogen
in whole
blood).
in pentose
and methylpentose
illustrated
from
charides
3
weight
is the
cent)
were
to normal
from the
In the
-
-
at 4 C. Since
initial
were
done
9 hours
ml.
(66 per
variations
bovine
dry
compared
a rapid
per
attributed
derivatives
for
of
glycogen
(32
mg./100
tamed
The
cent
as glycogen,
for
(table
per
and stored
platelets
accounted
be
as
may
be
who reported
blood
bovine
and
___
10
a
.
hours
observations
Wagner,15
from
.c
a
days
#{149}A11
values
hexose
a-
)<
E
Z
1-7
3
days
24
a
‘a
E
5.4
a
a
4
of Platelets#{176}
platelet.
a
‘oc
0a
_
Z
E#{149}
1-2
Content
e
.
88
.ca
#{149}‘
4,
Carbohydrate
Bovine
.
‘c
051
d
.
1179
PLATELETS
a
u#{176}
24
BOVINE
platelet.
r:
#{176}
mi
AND
glucose
and
Glucosamine,
present
F-Il
in
fractions
F-Il
frac-
of human
6).
for
of these
the
absence
constituents3133
glucuronic
acid
with
the
of both human
and bovine
of
acid
muco-
in platelets.
concurrent
platelets
abwould
of either
hyaluronic
acid
or heparin
the preliminary
findings
of Kerby
and
From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
1 180
WOODSIDE
Table
____________
Source
6.-Monosaccharides
Hydrolysis
of
Present
in isolated
Fractions
and Paper
Chromatography
Fraction
fractions
GA
Ga-N
+
+
+
+
0
0
Humanpiatelets
Bovineplatelets
HumnanRBC
Fraction
Ga
Gl
R
Ga-N
Gl-N
Ga
+
+
+
+
+
+
+
±
±
+
+
0
0
0
+
0
+
-
0
-
Prothrombin
0
monomer
always
0, monomers
+.
present;
GA:
glucuronic
mannoee;
fractions).
F:
fucose;
Langley34
platelets.
Ill
platelets
the
R. :
by
monomer
ribose;
Odell,
always
galactosamine;
SA:
to
the
Jr.,
absent;
Gl.N:
sialic
acid
(determined
and
Andersonm2
F
H
SA
-
+
+
-
+
+
±
±
-
+
+
-
+
+
+
-
+
-
-
0
on
is i- agreement
sulfate
( table
F-I
fractions
in
have
as adenosine-phosphate
identified
II
M
traces
but
galactose;
chemically
tile
)
6
aliquots
of
the
human
and
its
glucose;
M:
unhydrolyzed
B in
sulfate
fractions
of both
occasionally
Gi:
with
F-I
in
After
GI
absent
Ga:
ii
the existence
of chondroitin
sullate
tentative
identification
of chondroitin
of chondroitin
been
monomer
±,
glucosamine;
bovine
platelets.
ribose
derivatives
The
lets
Ga-N:
with
respect
addition,
monomers
and
present;
-.
not
determined.
acid;
KOCHOLATY
i and
I
Gl-N
AND
human
in rat
findings
of
of both
human
bovine
plate-
and
derivatives.14’28
Most
of the glucose
present
in F-I is probably
derived
from
glycogen.
In addition,
part of the glucose
present
could
exist as a polysaccharide
in combination
with
the galactose
and/or
galactosamine
which
were
also present
in this fraction.
The
biologic
significance
of
been
elucidated.
Recently
platelets
a polysaccharide
similar
to the
mers
isolated
( table
6
)
F-Il
fractions
from
F-lI
one
may
fractions
samine
suggested
on
and bovine
tions
( table
Table
per
cent
ments,
platelets
6).
and
yields
are not present
carbohydrate
that
in both
8.1
per
up
to two
cent
small
total
of the
residue
After
residue
was
platelet
of
3 times
found
this
monocells
specific
and
studies
fibrinogen.
in
the
total
and
bovine
)
gluco-
and
of
monomers
than
yield
has been
from 75 to 92 per
drying
rat
be
blood
platelets
glycoproteins
respectively
greater
preparations.
the supernatant
not
antigenicity.’16
of
similar
in
red
aldohexoses
Subsequent
presence
of human
times
has
from
could
of the
human
human
active
hexose,
one-half
F-lI
immunologically
two
amounts
fractions
variation
show that
of F-Il.
the
least
contain
in either
F-I or F-Il
moiety
was found
in
extraction
remaining
for
at
mechanism.
revealed
the
of the
and
of
of hydrolyzed
of physiologically
F-Il
in
obtained
and
which
similarity
and
presence
responsible
very
and
A similar
above
data
precipitation
at 40 C. and
ethanol,
the
only
F-I
present
the
platelets
the
amld prothrombin
7 reveals
for
6. From
identified
the blood
clotting
bovine9
platelets
7 were
obtained.
Eidinger.1
The
ethanol
have
)
s
Odell,
Jr.,32 have
of uronic
acids
of human
paper
chromatograms
the possible
existence
may
influence
human38
and
counted
Crostitm7
and
free
in table
on
complexes
and
polysaccharide(
listed
speculate
carbohydrate-protein
Bestetti
the
Anderson
which
was
washed
Human
F-Il
hexoses
fracare
other
listed
ac-
( 7.3
platelets
In
.
those
have
which
experiI1
table
reported
by Glegg
and
cent of the total hexoses
In addition,
solution
supernatant
a highly
soluble
after
84 per cent
solution
with
12 to 15 ml.
to contain
additional
in vacuo
of 95 per
hexoses.
cent
The
From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
(;ARBOHYDRATES
OF’
Table
HUMAN
AN!)
7.-Percentage
1181
PLATELETS
BOVINE
of Carbohydrate
isolated
in
Fractions
i and
Yield
Source
Human
platelets
Bovine
platelets
#{176}Thetraces
( 40 .tg. equivalent
large
mg.
I
II
I
II
4.1
33.9
10.1
51.3
of sialic acid in Fraction
to 0.4 tg. sialic acid).
variety
probable
of
and
I
hexose-containing
complexity
human
%
Fraction
of
bovine
weight
Pentose
and
methyl.
pentose
Hexose
0.79
6.52
0.04
0.04
0.08
0.13
0.006
0.022
1.32
0.02
0.09
0.008
6.79
0.06
0.09
0.015
dry
weight
may
Sialic
fractions
carbohydrate
further
protein
moieties
acid
glucuronic
acid
exemplified
existing
in
the
both
hednetointerferenceof
protein
the
ii
as % dry
Expressed
platelets.
Sunit
1. Average
platelets
weight
larger
values
for
cose,
galactose,
acid
and sialic
acid were
Glucose
and ribose
2.
the
total
carbohydrate
content
were
8.47 and 8.77 per cent of the dry
basis,
human
platelets
were
calculated
than
bovine
platelets.
By chemical
and
TCA-soluble
residue
glucuronic
acid,
fractions
contained
fraction
and
tosamine
bovine
fucose,
glucosamine,
polysaccharide
and
rihose,
glucosamine,
galactosamine,
glucuronic
detected
in both
human
and bovine
platelets.
were
found
to be the only
monomers
present
in the
mannose,
5 per cent
TCA-insoluble
of human
weight,
respectively.
On a dry
to be approximately
one-third
chromatographic
analysis,
gIn-
of both
glucose,
human
galactose,
galactosamine
both
of
glucuronic
acid
human
and bovine
mannose,
and
and
sialic
bovine
monomers
as
human
and
platelets.
fucose,
acid.
The
platelets
well
as
The
ribose,
acid
mucb-
contained
galactose,
galac-
glucose.
and
ribose.
3. Glycogen
was
contrast,
resulted
30
in
per
human
length
bovine
from
total
Platelets.
of storage
separation
KOH
of the
whole
extraction
the glycogen.
and glycogen
These
variations
of
platelets
in whole
1. Valores
medie
e bovin
pro
esseva
Calculate
super
approximativemente
le
analyse
chimic
e
human
e bovin
de
galactosamina,
acido
tile
ethanol
contents
were
or in 0.85
p1atelets
vas
by
partially
glycogen
bovine
giycogen
were
blood
Total
and
incomplete.
precipitation
found
in both
dependent
per
on
NaCI
cent
the,
after
blood.
SuMMiuo
human
platelets.
human
followed
hexose
recoveries
ill both
and
bovine
cent
cent
complete
4. Variations
in both
TCA
from
sonorated
ethanol
precipitation
of
per
subsequent
however,
In
found
\VIS
by 5
extracted
IN
INTERLINGUA
le contento
total
de
8,47
pro
cento
base
un
e 8,77
hydrato
del
de pesos
sic, plachettas
tertio
plus
grande
que
chromatographic
glucosa,
glucuronic.
demonstrava
galactosa,
e acido
mannosa,
sialic.
de
carbon
peso
sic,
in
plachettas
respectivemente.
human
le
se revelava
como
plachettas
bovin.
Le
le presentia
fucosa,
ribosa,
in plachettas
glucosamina,
From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
1182
WOODSIDE
2. Glucosa
e ribosa
plachettas
de
human
acido
tose,
esseva
como
trichioroacetic.
mannosa,
e acido
Le
fucosa,
sialic.
e bovin
le sol
etiam
Le
non
de
man
per
e bovin.
esseva
Tamen,
Per
de
kalium
le caso
in
parte
0,85
del
con
per
plachettas
intervallo
pro
cento
1. Glegg,
R.
isto,
human
e etiam
Glycogeno
sonorate
de
con
ethanolic
esseva
plachettas
ethanolic
extraction
precipitation
total
human
de
de
plachettas
glucuronic
plachetta.
ab
galac-
30
hu-
glycogeno
pro
cento
resultava
de
in le corn-
glycogeno.
in le contentos
del
de
del
cento
glucosa,
del
precipitation
contrasto
del
typos
le caso
galactosamina,
e acido
trichioroacetic
subsequente
sequite
recuperation
4. Variationes
acido
in
contineva
acide
KOCHOLATY
in 5 pro
glucosamina,
galactosamina
in ambe
de
le
incomplete.
hydroxydo
plete
cento
residuo
glucuronic,
de
tanto
solubile
mucopolysaccharido
monomeros
5 pro
trovate,
le fractiones
solubile
acido
galactosa,
glucosa,
e ribosa.
3. Glycogeno
esseva
trovate
extrahite
in
assi
ribosa,
fraction
contineva
monomeros
bovin,
AND
de
e etiam
immagasinage
NaCl
post
br
hexosa
e de
glycogeno
esseva
bovin.
Iste
variationes
dependeva
in
total
o in
del
plachettas
separation
ab
in
sanguine
le sanguine
trovate
total.
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1960 16: 1173-1183
Carbohydrates of Human and Bovine Platelets
E. E. WOODSIDE and W. KOCHOLATY
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