Document ID: EPA-HQ-OPP-2002-0002-0004
Agency: epa
Document Type: Supporting & Related Material
Title: 
Posted Date: 2002-02-06T05:00Z

,
STUDYTITLE
DATA
REVPEW
IN
SUPPORT
OF
THE
REQUEST
FOR
TEE
E­?
UUWE"
PPON
OF
METHYL
ANTHRANILATE
FROM
THE
REQUWEMENTS
OF
.A
TOLERANCE
ON
ALL
RAW
AGRICULTUERAL
COMMODITIES
DATA
REQUIREMENTS
Food
Quality
Protection
Act.

AUTBOR
Leonard
R.
Askham,
PbD.

REVIEW
COMPLETED
ON
April
2,
1992
SUBMITTOR
Bird
Shield
Repellent
Corporation
P.
O.
Box
785
Pullman,
WA
99163
,
I'
STATEMENT
OF
DATA
CONFIDENTIALITY
CLAMS
No
claim
of
confidentiality
is
made
for
any
information
contained
in
this
study
on
the
basis
of
its
falling
within
the
scope
of
FIFRA
lO(
d)(
l)(
A),
@)
or
(C)

Submitter:
'
Bird
Shield
Repellent
Corporation
Submitting
Agent:&
eonard
R
Askham,
PhD.
Date
6
3
GOOD
LABORATORY
PRACTICE
STATEMENT
Bird
Shield
Repellent
­
EPA
Reg.
No.
66550­
1
L
This
submission
does
not
fall
within
the
requirements
of
40
CFR
Part
160
/

..
..
..
TITLE
Product
Identity
and
Composition
1.
Product
Identity
and
Composition
2.
MerckIndex
3.
Registry
of
Toxic
Effects
of
Chemical
Substances
Biological
Derivation
of
Methyl
Anthranilate
1.
2.

3.
4.
Methyl
Anthranilate
Content
of
Ohio
Concord
Grapes
Concord
Wine
Composition
as
Affected
by
hditurity
and
Processing
Technique
The
Odor
Quality
of
Labrusca
Grapes
An
Investigation
of
the
Volatile
Flavor
Composition
of
Vitus
Labrusca
Grape
Musts
and
Wines.
I.
Methyl
Anthranilate
­
Its
Role
in
the
Total
Aroma'Picture
ofLabrusca
Varieties
Isolation
and
Identification
of
Volatiles
from
Catawba
Wine
Methyl
Anthranilate
as
an
Aroma
constituent
of
American
Wine.
Inheritance
of
Methyl
Anthranilate
and
Total
Volatile
Esters
in
yi&
SPP­
Red
Wine
Aroma:
Identification
of
Headspace
Constituents
SECTION
Tab.
A'

Tab.
B
US/
FDA
Registration
of
Methyl
Anthranilate
asaGenerallyRecognized
as
Safe(
GRAS)
CompoundTab.
C
1.
21
CFR
182.60
­
Synthetic
Flavoring
Substances
and
Adjuvants;

2.
2
1
CFR
184.102
1
Benzoic
Acid
Methyl
Anthranilate
Methyl
Anthranilate
as
a
Flavor
and
Perfume
Ingredient
1
..
Fenaroli's
Handbook
Qf
Flavor
Ingredients,
Second
Edition,
Volume
2
2.
Perfume
Synthetics
and
Isolates
U.
S.
Department
of
Commerce
Scientific
Literature
Review
of
Anthanilates
in
Flavor
Usage.
Vol.
1.
Introduction
and
Summary
Tables
of
Data,
Bibliography
Tab.
D
Tab.
­E.

1.
Toxicity
to
Mouse,
Rate
and
Guinea
Pig
2.
Chemical
Identity
and
Physical
Properties
3.
Pharmacological
and
Toxicological
Effects
4.
NaturalOccurrence
5.
Flavor
and
Extract
Manufacturers'
Association
(FEMA)
and
5.
Addendum
to
Table
IV­
2
7.
Bibliography
8.
DataGuide
National
Academy
of
Science
WAS)
Use
Levels.

AcuteToxicityforMice,
Hamsters,
Birdsand
Fish
Tab.,
F
1
­
Food
Fiavoring
and
Compounds
of
Related
Structure.
I.

2.
Acute
Oral
Toxicity
and
Repellency
of
933
Chemicals
to
3.
Studies
of
Drug
Adsorption
from
Oral
Cavity:
Physio­
Acute
Oral
Toxicity
House
and
Deer
Mice.

chemical
Factors
AEecting
Absorption
from
Hamster
Cheek
Pouch.

998
Chemicals
to
One
or
More
Species
of
Wild
and
Domestic
Birds.
5
.
Comparison
of
Fish
Toxicity
Screening
Data
and
QSAR
Predictions
for
48
Aniline
Derivatives.
4.
The
Acute
Oral
Toxicity,
Repellency
and
Hazard
Potential
of
(@
T
TAB
A.
PRODUCT
IDENTITY
AND
COMPOSITION
TABLE
OF
CONTENTS
\.%/
'

1
.
Product
Identity,
and
Composition
2.
The
MerckIndex
3.
Material
SafetyData
Sheet
4,
Registry
of
Toxic
Effects
of
ChemicalSubstances,
Volume
1
(c)
Additional.
informtion
on
iqredient~
(i)
a~
chanical
name
is
methyl
arrthranilate
othersvnorrvms:
Anthranilic
acid,
methyl
ester
Mew1
2­
amhdIemoate
Methyl
o­
aminobenzoa~
2­
AminabenZoic
acid
m
y
1
ester
(ii)
CPS
Resistry
NLmJser:
.
134­
20­
3
(1985­
1986)
{iii)
StmAud.
Forrnrla
(i)
Methyl
an­
ate
is
mined
synthetically
by
.
.
present#
of
sulfuric
acid
(see
pspenaix
1).

(ii)
Fonnilatim
of
BIRD
SKIELD
(Refer
to
i0nfidmtia.
l
state­
esterifying
anthmdlic
add
w
i
t
h
methyl
alcdbol­
in
the
ment
of
fonuila,
fonn
8570­
4,
Confidmtial
Attadrment,
CnxS
F&
fer?
mce
1).
\
!

i.

i
I
THE
MERCK
INDEX
AN
ENCYCLOPEDIA
OF
CHEMICALS,
DRUGS,
AND
BIOLOGICALS
ELEVENTH
EDITION
Susan
Budavari,
Editor
Maryadele
J.
O'Neil,
Associate
Editor
Ann
Smith,
Assistant
Ediror
Patricia
E.
Heckelman,
Editorial
Assistant
Published
by
MERCK.
&
CO.,
INC.

R
A
H
W
A
Y
,
N
.J
..
U
.S
.A
.

1989
I
17.96%
H
12.08%
iN
18.65%.
0
21.30%.
CH,
NHCH,­
>H,
OH.
f
h
e
in
vivo
precursor
of
choline.
Has
been
isolated
'rom
a
mutant
of
Neurospora
c
m
s
n
which
has
lost
its
ability
o
synthesize
choline:
Horowitz.
J.
Bioi
Chem.
162,
413
'1946).
Prepd
by
mixing
ethylene
oxide
with
concd
methyl­
mine
soh
with
external
cooling:
Knorr.
Matthes.
Ber.
31,
1069
(1898);
Lowe
ct
02..
Brit.
pat.
763,434
(1956
to
Oxirane
Ltd.);
Nikolaev
et
al.,
Zh.
qbshch.
Khim.
33,
391
(1963).
Viscous
liquid.
Fishy
odor.
dm
0.937.
bp,
60
155­
156";
>p12
64­
659
n$
1.4385.
Miscible
with
water.
alcohol,
ether.
Zorrosive
to
skin,
cork,
metals.
Strong
base.
Forms
a
deli­
xuescent
salt
with
HCI.
LD,
orally
in
rats:
2.34
glkg,
Toxic
Substances
Lisr.
H.
E.
Christensen,
Ed.
(1974)
p
339.
Picrate,
yellow
crystals,
mp
148­
150".
Caurion:
Irritating
to
skin,
eyes,
mucous
membranes.

minophenol
sulfate;
p­
hydroxymethylaniline
sulfate;
Pho­
5940.
p­
Methylaminophenol
Sulfate.
Monomethyl
­p­

:ol;
Verol;
Rhodol;
Armol;
Elon:
Genol;
Graphol;
Photo­
Rex:
Pictol;
Planetol;
Metol.
C,,
H,
N,
O,
S:
mol
wt344.38.

:HOC6H,
NHCHJ2.
H2SO,.

Sol
in
20
parts
cold,
6
parts
boiling
water;
slightly
sol
in
alc;
Crystals.
Discolors
in
air.
mp
about
260'
with
decompn.

nsol
in
ether.
Keep
well
closed
and
protected
from
light.
C
48.82%,
H
5.85%.
N
8.14%.
0
27.87%,
S
9.31%.

USE:
Photographic
developer,
dyeing
Furs.

vlaniline.
C,
H,
N:
mol
wt107.15.
C
78.46%.
H
8.47%.
N
5941.
Methylaniline.
~­~
e~
hglbenze~
ramine:
rnonomerh­

13.07%.
C,
H,
NHCHJ.
Made
by
heating
aniline
chloride
and
methyl
alcohol
under
pressure.

exposure
to
air.
d
p
0.989.
mp
­5579
bp
194­
196'.
nZ1J
Colorless
or
slightly
yellow
liquid;
becomes
brown
on
1.5702.
Slightly
sol
in
water;
sol
in
alc,
ether.
LD
orally%
rabbits:
280
mglkg.

esrer;
methyl
2­
aminobenzoate;
neroli
oil,
artificial.
C,
H,­
5942.
Methyl
Anthranilate.
2­
Aminobenzoic
acid
mefhyl
NO,;
mol
wt
151.16.
C
63.56%.
H
6.00%
N
9.27%,
0
other
essential
oils
and
in
grape
juice:
also
obtained
syn­
21.17%.
Occurs
in
neroli.
ylang­
ylang.
bergamot,
jasmine,

thetically
by
esterifying
anthranilic
acid
with
CH,
OH
in
presence
of
HCI.

water:
freely
sol
in
alcohol
or
ether.
LD,
orally
in
rats.
Crystals.
d
1.168,
mp
24­
259
bpIs
135.59
Slightly
sol
in
mice:
2910.
3900
mglkg,
P.
M.
Jenner
et
al..
Fwd
Cosmer.
.
Toxicol.
2,
327
(1964).
CSEl
As
perfume
for
ointments:
manuf
synthetic
perfumes.

cenediono;
8­
methylanthraquinone.
C,,
H,,
O$
mol
wt
5943.
2­
h­
lethylanthraquinone.
2­.
MelhpI­
9,
f
0­
ahthro­

222.23.
C
81.06%.
H
4.54%
0
14.40%.
Occurs
in
teak­
wood.
Prepd
by
oxidation
of
2­
methylanthracene;
by
for­

Syn
4,
43
(1925);
from
6­
and
7­
methylanthraquinone­
l­
mation
from
phthalic
anhydride
and
toluene:
Fieser.
0%.

carboxylic
acids
with
powdered
copper
in
quinoline:
Fiescr.
Martin.
J.
Am.
Chem.
Soc.
58.
1443
(1436);
from
1.4­
naph­
thoquinone
and
isoprene:
Carothers.
Berchet,
ibid.
55.
2813
(1933).

Needles
from
alcohol,
mp
177".
Sublimes.
Very
sol
in
benzene,
toluene.
xylene;
sol
in
alcohol,
ether.
glacial
acetic
acid.
concd
H,
SO,;
insol
in
water.

noside;
methylarbutoside.
C,
H,
O,;
mol
wt286.28.
C
5944.
Methytarbutin.
4­
Melhox~
phengl­~­
D­~
lucopyra­

54.54%,
H
6.347'
0
39.12%.
Occurs
together
with
arbutin
I~
"
HANUFACNREB
WHEATEC
'
I
:
'
mD&
ss:
2
S.
076
0rchard"
Rd.
,,

Wheaton,
IL
60187
EMERGENCY
TELEPHONE:
708­
682­
3024
I
_.
Hethyl
Anthranilate
CBEHICdL
NAXE
AND
SYNONYMS:
Methyl
2
­
Aminobenzoate
*

TRADE
NAME
AND
SYNONYMS:
HETHYL
ANTHRANILATE,
HA
­
FCC
CHpfIcILL
FAKILY:
Aromatic
Acid
Ester
FOWLA:
C8H9N02'
6
4
2
2
.
C
H
NH
CO
H
CAS
REGISTRY
NUHBER:
134­
20­
3
DOT
SHIPPZHC
CLASSIFICATION:
Cherni­.
lc
,
NO1
PBODUCT
NUMBER:
..
*

..

S
E
C
T
I
O
N
11
.
HAZARDOUS
INGREDIENTS
M
T
E
R
U
L
:
.*

Methyl
99+
TLV
(Units)

SECTION
111
.
PHYSICAL
DATA
­
r"

ik."
d
­
,i
\?
"
..­

,:
BOILING
POINT,
76­
e
Xg:
206OC
6
FREEZING
POINT
:
24
*c
SPECIFIC
CBAVITY:
1.165
VAPOR
PRESSURE
AT
.2OoC.
:
<
lmm
Hg
VAPOR
DENSITY
(Air
­
1):
No
Data
Found
SOLUBILITY
IN
WATER:
Approximately.
O.
1%
X
VOLATI&
ES
BY
VOLUME:
0.5%
max.
a
s
w
a
t
e
r
]E;
VAPORILTION
RATE
:
No
Data
Found
*
*

APPEARANCE
AND
ODOR:
Colorless
t
o
pale
y
e
l
l
o
w
l
i
q
u
i
d
vith
a
b
l
u
i
s
h
fluorescence,
o
r
w
h
i
t
e
c
r
y
s
t
a
l
s
a
t
room
temp.

.­
I
.I"..
.­
.
.
...*.
­
­
­
.­
*.
Crape
like'
odor
'.
..
..
.
.
"

..

CTION
XV.
­­
FIRE
AND
EXPLOSION­
WARD
DATA
FLASH
POINT:
.
212'F
(CC)

FLAHHABLE
LIMITS
I
N
AIR:
No
Data
Found
EXTINGUISHING'
HEDIA:
Carbon
Dioxide
Dry
Chemical,
Foam.
SPECIAL
FIRE
FICXTING
PROCEDURES:,
F
u
l
l
p
r
o
t
e
c
t
i
v
e
e
q
u
i
p
m
e
n
t
i
n
c
l
u
d
i
n
g
s
e
l
f
­c
o
n
t
a
i
n
e
d
breaching
apparatus
should
be
used.
Uater
spray'may
be
i
n
e
f
f
e
c
t
i
v
e
.
If
water
1s
used,
fog
nozzels
are;,$.
referable.
Water
may
be
used
t
o
c
o
o
l
c
l
o
s
e
d
c
o
a
t
a
i
n
e
t
r
t
o
p
r
e
v
e
n
t
p
r
e
s
s
u
r
e
b
u
i
l
d
­u
p
.a
n
a
p
o
s
s
i
b
l
e
a
u
t
o
i
g
n
i
t
i
o
n
or
explosion
when
­exposed
to
­0
.

AUTOIGNITION
TEMPERATURE:
No.
Data
Found
..
extreme
heat.
E'
.:+
n.
uN"
suaL
FIRE
AND
\$

\
\­+
J
EXPLOSION
w
s
;
+Closed
containers
may
explode
(due
t
o
the
build­
up
of
'&
,4
.,
'\
I
Pressure)
wh?
n'@
XpOSed
t
o
extreme
heat.
>.
I
1
..
THRESHOLD
LIMIT
VALUE:
None
Assigned
EFFECTS
OF
OVEREwosu~~:
E
x
c
e
s
s
i
v
e
e
x
p
o
s
u
r
e
m
y
c
a
u
s
e
s
k
i
n
a
n
d
eye
i
r
r
a
t
a
t
i
o
n
.
Other
t
o
x
i
c
e
f
f
e
c
t
s
a
r
e
unknown.
Methyl
a
n
t
h
r
a
n
i
l
a
t
e
is
included
i
n
t
h
e
GKAS
(Generally
recognized
as
s
a
f
e
)
l
i
s
t
..

E
m
~G
m
c
y
AND
FIRST
AXD
PROCEDURES:
I
F
INHALED:
If
a
f
f
e
c
t
e
d
,
remove.
from
e
x
p
o
s
u
r
e
.
R
e
s
t
o
r
e
b
r
e
a
t
h
i
n
g
.
Keep
warm
and
q
u
i
e
t
.
I
F
ON
SKIN:
Wash
affected
a
r
e
a
t
h
o
r
o
u
g
h
l
y
w
i
t
h
s
o
a
p
a,
nd
water.
I
F
IN
EYES:
F
l
u
s
h
e
y
e
s
w
f
t
h
large
amounts
of
water
for
15
minutes.
Get
medical
a
t
t
e
n
t
i
o
n
.
IF
SWALLOWED:
Never
give
anything
by
mouth
t
o
a
n
u
n
c
o
n
s
c
i
o
u
s
p
e
r
s
o
n
.
G
i
v
e
s
e
v
e
r
a
l
g
l
a
s
s
e
s
o
f
water.
If
vomiting
is
not
spontaneous,
induce
vomiting
by
g
e
n
t
l
y
t
o
u
c
h
i
n
g
t
h
e
back
of
t
h
e
p
a
t
i
e
n
t
s
t
h
r
o
a
t
w
i
t
h
a
f
i
n
g
e
r
.
Keep
a
i
r
w
a
y
c
l
e
a
n
.
S
e
e
k
q
e
d
i
c
a
l
a
t
t
e
n
t
i
o
n
.

TOXICITY
Oral
LDSu
(.
Rat)
3,000
­
5,000
mg/
kg.

c
SECTION
.VI.
REACTIVITY
DATA.
.­
I
­

STABILITY:
Stable
WCOHPATIBILITY:
O
x
i
d
i
z
i
n
g
.,

HAZARDOUS
DECOHPOSITION
PRODUCTS:
BY
FIKE:
Carbon
Dioxide,
Carbon
Monoxide
,.
and
Oxides
of
Nitrogen.
HAZARDOUS
POLYMERIZATION:
Will
Not
Occur.
_.
.
..

~

.­
SECTION­
VI1
SPILL
OR
LEAK
PROCEDURES
..:
"

STEPS
TO
BETAKEN
IN
CASE
THEMATERIAL
IS
SPILLED
OR
RELEBEP:
'
S
p
i
l
l
e
d
material
can
be
c
o
l
l
e
c
t
e
d
a
n
d
h
e
l
d
for
reclaim
or
placed
in
a
cov'ered
waste
disposal
c
o
n
t
a
i
n
e
r
.

WASTE
DISPOSAL
.METHOD:
S
a
n
­i
t
a
t
y
l
a
n
d
f
i
l
l
or
i
n
c
i
n
e
r
a
t
e
Cr,
a
p
p
r
o
v
e
d
f
a
c
i
l
i
t
y
.
Do
not
i
n
c
i
n
e
r
a
t
e
c
l
o
s
e
d
c
o
n
t
a
i
n
e
r
.
Dispose
of
In
accordance
with
Federal,
State
,
.a
n
d
L
o
c
a
l
r
e
g
u
l
a
t
i
o
n
s
r
e
g
a
r
d
i
n
g
p
o
l
l
u
t
i
o
n
.
'

~­­­
I­"".
rrPT
SECTION
"".­
V
I
I
I
a
E
C
T
A
L
..
PR!
XE­
NJSS%
Z$
SN
RESPIRATORY
PROTECTION:
Use
a
r
e
s
p
i
r
a
t
o
r
a
p
p
r
o
v
e
d
for
organic
vapors,
fumes
and
mists
i
n
areas
laden
wtth
vapor.
VENTILATION:
L
o
c
a
l
e
x
h
a
u
s
t
p
r
e
f
e
r
a
b
l
e
,.
g
e
n
e
r
a
l
e
x
h
a
u
s
t
a
c
c
e
p
t
a
b
l
e
.
.
.

PROTECTIVE
GLOVES:
R
e
q
u
i
r
e
d
f
o
r
l
o
n
g
or
repeated
contact.
EYE
PROTECTION:
Wear
s
a
f
e
t
y
s
p
e
c
t
a
c
l
e
s
w
i
t
h
unperforated.
sideshield6.
OTHERPROTECTIVEEQUIPMENT:
Eye
wash,
Safety
shower,
\

..

.
SECTION
IX
.
SPECIAL
PRECAUTIONS
1
PRECAUTIONS
TO
BETAKEN
IN
HANDLING
AND
STORAGE:
.Do
not
store
neer
open
flames
or
o
x
i
d
i
z
i
n
g
a
g
e
n
t
s
.
Avoid
stor€
ng
i
n
high
t
e
m
p
e
r
a
t
u
r
e
a
r
e
a
s
r
OTHERPRECAUTIONS:
Avoid
excessive
exposure:
Do
n
o
t
t
a
k
e
i
n
t
e
r
n
a
l
l
y
.

REVISED:
June
17,
1988
SUPEBCEDES:
July
20,
1984
..

,_

..
.,
..
~MEIHYLANIHRANIIATE"

safety
data
for
=thy1
anthranilab
summarized
in
the
follwjng
reference
bo&:
­in
mishy
of
Toxic
Eff­
O
f
chemical
substances
R
e
g
i
s
t
r
y
of
Toxic
Effects
of
chdm
substances
edited
by
R.
L.
Tatken
and
R.
J.
Lewis,
Sr.
1981­
82
edition.
U.
S.
Dept.
of
Health
and
Human
Services,
Public
Health
service
Centers
for
Disease
Control,
National
Institute
for
Occupational
Safety
and
fiealth,
Cincinnati,
OH
45226
me
above
ref­
lists
the
following
toxicity
data:

mth~
l
­late
CAS
RN:
134­
20­
3
Skin
Irritation
­
­it
500
mz/
24
hours:
Merate
Q
t
Edited
by
Rodger
L.
Tatken
and
Richard
J.
Lewis,
Sr.

US.
DEPARTMENT
OF
HEALTH
AND
HUMAN
SERVICES
Public
Health
Service
Centers
for
Disease
Control
National
Institute
for
Occupational
Safety
and
Health
Cincinnati,
Ohio
45226
June
1983
..
Cnnu
b34.71
"
"
RATAW
hU#
".

WECS
U6UOGRAPHIC
REFERENCES
1981­
82
M??
w
1.
2.

3.
4.

5.
6
.
7.

8.
Methyl
anthranilate
Content
of
Ohio
Concord
Grapes.
Concord
Wine
Composition
as
Affected
by
Maturity
and
Processing
The
Odm
Quality
of
Labrusca
Grapes.
An
Investigation
of
the
Volatile
Flavor
Composition
of
Vitus
Technique.

Labrusca
Grape
Musts
and
Wines.
I.
Methyl
Anthranilate
­
Its
Role
in
the
Total
Aroma
Picture
of
Labrusca.
Varieties.
Isolation
and
Identification
of
Volatiles
&om
Catawba
Wine
Methyl
anthranilate
as
an
Aroma
Constituent
of
American
Wine.
Inheritance
of
Methyl
anthranilate
and
Total
Volatile
Esters
in
Red
Wine
Aroma:
Identification
of
Headspace
Constituents.
­
vitis
spp.
a"
.
..

'
A
Research
Note
Methyl
Anthranilate
Content
of
Ohio
Concord
Grapes
JIM­
WEN
R.
LIU
and
JAMES
F.
GALLANDER
ABSTRACT
Concordgrapesgrown
at
five
locations
in
Ohiowerecollected
in
1982
and
1983
for
analysis
of
methyl
anthranilate
(MA)
in
frearun
and
heat­
extracted
juice
samples.
The
hiA
content
of
freerun
juice
ranged
from
0.33­
2.05
mg/
L
in
1982
and
0.14­
0.91
mg/
L
in
1983.
The
MA
content
was
higher
in
heat­
extracted
juice
than
in
frearun
juice.
Heat­
extracted
juice
contained
0.19­
2.60
mdL
of
MA
in
1982
and
0.22­
3.50
mg/
L
in
1983.
In
general,
the
MA
content
in­
creased
with
maturity
and
was
highest
in
grapes
grown
in
the
cool­
est
regions.

INTRODUCTION
METHYL
ANTHRANILATE
(MA),
the
methyl
ester
of
o­
aminobenzoic
acid,
is
one
of
the
several
compounds
which
contribute
to
the
characteristic
aroma
of
Concord
grapes,
This
compound
was
used
in
synthetic
grape­
flavored
products
before
it
was
found
in
grapes
(Acree,
1981).
The
measurement
of
MA.
provides
basic
information
for
the
con­
trol
of
flavor
quality
of
Concord
grapes
and
its
products.
There
are
several
methods
available
for
the
determination
of
MA.
These
include
a
colorimetric
procedure
(AOAC,
1980),
a
g
a
s
chromatographic
procedure
(Mattick
et
al.,
1963),
and
a
fluorometric
method
(Casimir
et
al.,
1976).
Robinson
et
aL
(1949)
reported
that
MA
of
New
York
Concords
developed
mainly
ih
the
last
stage
of
maturation
and.
declined
slightly
when
fruits
became
overmature.
Clore
et
al.
(1965)
and
Fuleki
(1972)
obtained
similar
results
with
Washington
and
Ontario
Concords,
respectively.
Clore
e
t
al.
(1965)
also
reported
that
season,
pruning,
vineyard
location,
and
type
of
soil
could
affect
MA
content
of
Corn­
cord
grapes.
In
Ohio,
about
72%
of
the
present
acreage
is
planted
t
o
the
Concord
cultivar
(Cahoon,
1984).
Approximately
97%
of
the
Concord
crop
in
Ohio
is
used
for
processing.
The
purpose
of
this
study
was
to
determine
the
MA
content
of
Ohio
Concord
grapes
as
influenced
by
season,
vineyard
location,
and
maturation.

MATERIALS
&
METHODS
Sample
collection
Fruit
samples
were
collected
in
the
1982
and
1983
seasons
from
five
different
vineyard
locations
in
Ohio.
Ai
each
location
10
vines
were
selected
for
harvest
a
t
three
different
levels
of
maturity.
me
fust
level
of
maturity
was
chosen
as
the
time
when
all
berries
had
completedcolorchange(
post­
veraison).
Thesecondandthird
maturity
wereabout
1­
2
and
2­
4
wk,
respectively,
afterthe
first
sampling.
The
grapes
were
harvested
a
t
increments
of
approximately
2'Brix
when
possible.
For
each
maturity.
two
berries
were
randomly
taken
from
each
of
the
middle
20
clusters,
located
in
the
upper
portion
of
the
fine.
The
berries
from
the
10
vines
were
pooled,
and
a
total
of
400
benies
Were
collected
for
each
replicate.
Duplicated
samples
were
kept
in
plastic
containers
and
stored
overnight
at
4°
C.

Authors
Liu
and
Gallander
are
affiliated
with
the
Dept
o
f
Horticul­
ture.
Ohio
Agricultural
Research
&
Development
Center,
The
Ohio
state
Univ.,
Wooster.
OH
446993.

280­
JOURNAL
OF
FOOD
SCIENCE­
Volume
50
(1985)
Sample
preparation
Berries
(100)
from­
eachreplicatewerepassedthrough,
press
(Squeezo
strainer)
to
obtain
free­
run
juice.
Approha
mL
free­
run
juice
was
centiifuged
on
an
IEC­
140
rotor
at
mately
900
x
g
for
10
min
to
remove
gross
particles.
natant
was
analyzed
for
soluble
solids
("
Brix)
with
a
refrac
Thejuice
was
then
stored
in
glass
containers
at
­17°
C
u
analysis
could
be
performed.
Heat
extracted
juice
was
prepared
by
macerating
appr
200g
of
berries
in
a
Waring
Blendor
at
low
speed
for
1%
macerate
was
placed
in
a
250
mLbeaker,
covered
with
glass,
andheated
in
a
water
bath
at
85°
Cfor
1
hr.
macerate
was
cooled
to
40°
C
and
drained
through
cheesecl
heat­
extracted
juice
was
then
centrifuged
as
described
for
run
juice
and
the
supernatant
was
decanted
and
stored
at
glass
containers
The
frozen
juice
samples
of
both
freerun
and
heat
e
x
n
a
thawed
and
brought
to
room
temperature
prior
to
analyk
fa
Analysis
of
MA
The
fluorometric
procedure
of
Casimir
et
d
(1976)
was
the
determination
of
MA.
A
10
mL
sample
of
grape
juice
wa
distilled
in
a
Cash
distillation
assembly
(Research
and
Devel
Products,
Berkeley,
CA)
rather
than
in
a
micro­
Kjeldahl
app
as
described
by
Casimir
et
aL
(1976).
Preliminary
experiments
indicated
that
in
order
to
use
the
assembly
for
steam
distillation,
a
slight
modification
on
the
p
dure
of
Casimir
et
aL
(1976)
wasnecessary
to
obtain
more
96.0%
recovery
of
MA.
Approximately
76
mL
of
distillate
lected
in
8
min
in
a
100
mL
volumetric
flask
containing
20
m
pH
7
buffer.
Thebufferwaspreparedaccording
to
Carimj
(1976).
The
collected
distillate
was
brought
to
vo1um.
e
wilh
distilled
water.
The
temperature
of
the
distillate
was
kept
at
1°
C.
Thefluorescence
of
the
distillate
was
then
measured
Model
J4­
7439
Fluoro­
colorimeter(
AmericanInstrumen
SilverSprings,
MD)
equippedwith
a
primaryfilter
(pea
mission
at
360
nm)
and
a.
secondary
filter
(at
415
nmand
The
fluorometer
was
calibrated
with
standard
solutio
anthranilate.
The
standard
eqor
of
the
mean
of
three
M
A
?O.
OlQ
mg/
L
for
a
grape
juice
samplecontaining
1
with
a
tom1
of
300
mLglass­
distilled
water.
The
Cash
assemblywasflushedbetweensamples
three
RESULTS
&
DISCUSSION
RESULTS
indicated
that
there
was
a
large
variation
in
content
between
two
replicated
juice
samples.
For
exa
the
MA
content
of
heat­
extracted
juice
collected
a
on
9/
9/
82
was
0.75
2
0.40
mg/
L,
and
the
MA
c
free­
run
from
Wooster
on
9/
14/
83
was
0.23
k
(Tables
1
and
2).
Fuieki
(1972)
reported
that
cons1
variability
was
observed
when
juice
t
o
be
analyzed
was
tained
by
pressing
the
grapes.
To
reduce
variability,
he
ommended
the
use
of
whole
grapes
instead
ofjuice
steam
distillation.

2.05
mg!
L
in
1982
and
O714­
O.
9i
mg/
L
in
1983.
For
extracted
juice,
MA
content
varied
from
0.19­
2.60
mgi
1982
and
0.22
to
3.50
mg/
L
in
1983.
In
general,
heat
traction
increased
MA
coritent
in
the
juice.
Rice
(1975)
ported
that
hot­
pressed
,,(=
oncord
juice
contained
mu
higher
MA
than
did
cold:$
ressed
juice.
Clore
et
d
(196
analyzed
the
MA
content
rjf
Washington
Concords.
fhe
M
The
MA
content
of
free­
run
juice
ranged
from
0.3
.

8
.
z*
2753
9t7
13.9
!
S­
IA
2948
911
5
15.7
0.77
2
0.09
1.16
t
0.06
OB5
t
0.02
0.72
t
0.03
23
18
a
m
2424
9/
7
2593
9/
15
12.4
0.67
2
0.05
1.00
i
0.12
14.5
1.13
f
0.06
15.2
1.34
i
0.16
0.93
t
0.08
1.02
i
0.20
0.33
t
0.04
0.19
3
0.02
0.69
t
0.02
0.75
3
0.4Q
.
15.6
1.76
t
0.04
2.22
2
0.60
0.91
2
0.05
0.00
f
0.12
1.87
t
0.12
0.54
t.­
o.
oa
1891
8130
11.7
201
2
919
21
92
9/
28
2183
911
14.2
2372
911
3
2479
9/
23
15.1
1.66
t
0.04
15.9
2.05
2
0.20
2087
9t9
14.0
0.70
+­
0.12
13.4
1.52
t
0.18
.~
11BC
0.42
3
O.
0Zd
0.55
*
0.01
2272
9123
15.0
"
­
1.55
i
0.33
2.60
t
0.05
2414
1015
17.4
1.34
2
0.01
1.63
3
0.03
.
of
degrees
above
50°
F
within
the
state
from
­

April
to
the
date
of
harvest;
1.
c..
X:
t(
dally
~~

maxlmum­
daily
mlnlmum)/
2­
50].

*~y
f
duplicate
F
#*
,
dup{
icates
t­
standard
error
of
mean
f
"
Degree
Date
Soluble
s
{
of
solids
MA
content
(rng/
L)
wm'vd
#:
ion
daysb
harvest
P
~r
i
x
)
Free­
run
Heat­
extract
2836
911
12.6'
.
0.17
f
0.
Old
3133
0.28
i
0.00
3299
9/
21
.16.4.
0.18
t
0.06
0.48
f
0.00
0.22
t­
0.01
2552
916
271
9
911
4
13.7
0.14
2
0.02
0.42
i
0.1
3
15.1
0.22
+­
0.04
912
1
0.62
i
0.06
16.3
0.30
:
0.02
0.70
3
0.02
2214
916
14.6
0.23.2
0.08
911
4
089
t­
0­
01
17.0
2448
912
1
18.1
0.41
t
0.13
1.69
t
0.05
0.63
k
0.1
81.90
t
0.1
0
2507
917
14.1
2668
0.28
t
0.02
*I
911
5
0.74
i
0.02
15.7
279
1
9/
22
OS1
i
0.06
1.97
t
0.03
034
t
0.09
0.44
f
0.04
056
t­
0.01
1.70
t
0.14
0.91
t­
0.06
350
i
0.1
0
,I
9/
12,
14.9
0.16
i
0.02
..
cpcs3
2840
antral)
2352
"­

1
16.1
0.70
i
0.05
232
+­
0.12
E
2377
9/
8
129
2555
9/
19
155
2624
9/
27
16.6
r
of
degrees
above
50'F
region
wltnin
the
state
from
April
to
the
date
of
harvest;
Le.,
Z[
(aally
cIU­
of
duplicate
&
O!
duplicate
f
standard
error
of
mean
c
5
m
e
a
t
for
the
whole
grape,
skin,
pulp.
juice
drained
from
central
Ohio
vineyards
(Morrow
and
Columbus)
tended
to
i.
w.
and
juice
drained
from
skins
was
6.0,
5.8,
1.8,
2.2
be
lower
than
those
from
northern
Ohio
(Wooster,
San­
@
Zd
ppm,
respectively.
They
concluded
that
most
of
the
dusky,
and
Madison)
at
comparable
levels
of
maturity.
:.
w
h
y
1
anthranilate
was,
located
in
the
skin
fraction.
Clore
et
a1
(1965)
reported
the
effect.
of
vineyard
location
fr.
gcneral,
the
MA
content
increased
with
maturity.
This
on
MA
content,
but
noticed
that
soil
type,
pruning
severity,
P
m
weement
with
the
findings
of
other
researchers
(Rob­
and
solar
radiation
could
also
influence
MA
content.
Based
t
t
at..
1949;
Clore
et
al.,
1965;
Flueki,
1972).
Robin­
an
the
total
number
of
degree
days,
the
MA
content
seemed
a
t
t
al.
(1949)
reported
that
MA
content
of
New
York
to
be
related
to
the
regional
climate.
Grapes
from
colder
PrRn
..
Concords
increased
during
ripening
and
decreased
regions
contained
higher
concentrations
of
MA.
The
MA
.
ftlt
when
the
fruit
became
overmature.
However,
Clore
content
in
1983
was
generally
lower
than
that
of
19S2.

4
@
d
(1965)
found
that
MA
content
did
not
necessarily
This
was
apparently
due
to
a
warmerseason
in
1983.
;
k­

bQnc
when
grapes
became
'
overmature.
In
this
study,
were
some
decreases
in
MA
contrni
in
1982
from
the
­c
m
a
t
u
r
i
t
f
t
o
t
h
e
t
h
i
r
d
m
a
t
u
r
i
t
y
f
o
r
grapes
from
the
REFERENCES
'*
4mk
and
Madison
vineyards.
Whether
this
Kas
due
to
.
A
~~~,
T.
E.
1981.
The
odor
quty
ofi&
rusca
=apes.
ACS
s
Y
m
p
~

m
i
n
a
t
i
o
n
of
MA
beyond
the
third
maturity.
No
de­
AOAC.
1980.
"Official
Methods
of
Andy&"
13th
e
a
Association
1
­
in
MA
content
was
observed
for
samples
collected
in
'
Cahoon.
G.
A.
1984.
Private
communication.
Horticulture
Dept.
i.:
m
3
*ason.
Ohio
Agriculturd
Research
6
Development
Center.
Ohio
State
'k
MA
content
of
Concord
grapes
from
southern
and
',
...
.
casimir.
D.
3..
Moyer,
J.
c­,
md
Mattick.
L.
R.
1976.
Fluorometric
%
Univ..
Wooster.
OH.

Volume
50
IISBSI­
JOURNAL
OF
.FOOD
SCIENCE­
281
Pa
&coming
overmature
is
not
clear,
since
there
was
no
sium
Series
170:
11.

of
Official
Analytical
Chemists.
U'ashington,
DC.

..
determination
of
methyl
anthranilate
In
Concord
grape
juict.
Daw
varietier
Ch.
4
In
"Chemistry
of
Winem"
JAOAC
69:
269.
Clore,
W.
J.,
Neubert.
A.
M.,
Carter:
G.
H..
Ingalsbe.
D.
W..
and
Bmm­
Robinson.
W.
B..
Shaulis.
N.
J..
and
Pederson.
C.
S.
19
mund.
V.
P.
1965.
Composition
of
U'ashington­
uro&
aced
Concord
studies
of
grapes
grown
for
juice
manufactum.
F
m
pap­
.nd
juices.
Wash.
Agr.
ESP.
Sta.
Tech.
Bull.
482
96.

papes
grown
io
Ontdo
during
ripening
in
the
1970
season.
Can.
J.
Plant
ScL
52:
863.
Matti4
L.
R..
Robinson.
W.
B..
Weirs,
L.
D.,
and
Barry.
D.
L.
1963.
Determination
of
methyl
anthranilate
in
grape
juice
by
electron
Rim.
A.
C.
1975.
Chemistry
of
a­
inemaking
from
native
American
dfinfty­
gas
Chromatography.
1.
Amic.
Food
Chem.
11:
334.
Webb.
P.
88.
Am.
C
h
c
a
SOC.
Adv.
Chem,
Sex.
197.
.
a
F
d
e
a
T.
1972.
Changes
in
the
chemical
composition
of
Concord
Ma
received
7/
13/
84;
accepted
8[
27/
84.
.­
:*

KINETICS
OF
WATER
UPTAKE
BY
FOOD
PRODUCTS..
.
From
page
279
REFERENCES
Baumann.
€I.
1966.
Apparatur
nach
Baumann
Zur
bestimmung
der
Anstrichm.
68(
9):
741.
flussigkeit
soufnahme
von
pulnigen
substanzen.
Fette,
Seifen,

Gerschenson.
L.
N..
Boquet.
R..
and
Bartholomai.
G.
B.
1983.
Ef­
fect
of
thermal
treatments
on
the
moisture
sorption
isotherms
tinurn
L.).
Lebensm.­
Wiss.
u.
Technol.
16:
43.
of
protein
isolate.
starch
and
flour
from
cheackpea
(Cicer
arie­

Hermansson,
A.
M.
1972.
Functional
properties
of
proteins
for
foods.
Swelling.
Lebensm.­
Wiss.
u.
Technol.
5(
1):
24.
Kuntz,
D.
A.,
Nelson,
A.
1
...
Steinberg,
M.
P..
and
Wei.
L.
S.
1970.
Control
of
chalkmess
in
soymilk.
J.
Food
Sci.
43:
1279.
Pilooof.
A.
M.
R.,
Bartholomai.
G.
B.
and
Chirife.
J.
1982.
Kinetics
of
nitrogen
solubility
loss
in
heated
flour
and
protein
isolates
Rdston.
M.
L.
and
Jennrich,
1­
1978.
Dud.
a
derivative
free
algo­
from
bean,
Phaseolus
vulgaris.
J.
Food
Sci.
47(
1):
4.

Romo.
C.
R.
1980.
The
extraction
characterization
together
with
rithm
for
nonlinear
least
squares.
Technometrics
20(
1).

nutritional
and
technological
properties
of
protein
contained
in
chilean
bean
and
rapeseed
meal.
Thesis.
National
CoUepc
ol,

Torgarsen,
H.
and
Toledo,
R.
T.
1977.
Physical
propenies
Technology.
X'eybridge.

preparations
related
to
their
functional
characteristics
g'
"4\
:
nuted
meat
systems.
J.
Food
Sei.
42(
6):
1615.
cpcc
:f
Urbanski.
G.
E..
Wei.
L.
S'.
Nelson.
A'I..
and
Steinberg.
M
p
.­
Flow
characteristics
of
soybean
constituents
contro&
ed'b,
'%

Ms
received
3/
14/
84;
reevked
7/
21/
84;
accepted
9/
24/
84.
of
total
to
imbibed
water.
J.
Food
Sei.
48:
691.
%.&
a*

assistance,
and
Dra.
0.
Kith
i
s
heartily
thanked
for
her
&
The
authors
ais0
acknowledge
Millipore
Co.
for
their
est
in
this
work
and
for
valuatile
comments.
I
i
I
.
l$
V*

Richard
R.
Nelson
and
Terry
E.
Acree
Respectively
Graduate
Research
Assistant
and
Associate
Professor
of
Biochemistry,
Department
of
FoodScience,
New
York
State
Agricultural
Experiment
Station,
Cornell
University,
Geneva,
New
York
14456.

The
authors
gratefully
acknowledge
the
gift
of
Concord
pigment
from
G.
Hrazdina,
assistancein
the
sensory
analysisfrom
R.
M.
Butts
and
1.
D.
Tyler,
andstatisticalas­
sistance
from
J.
Barnard.

This
manuscript
approved
by
the
Director
of
the
New
York
State
Agricultural
Experi­

Presented
at
the
Annual
Meeting
of
the
American
Society
of
Enologists,
June
23,

Received
June
23,1977.

Accepted
for
publication
January
17,1978.
ment
Station
for
publication
as
Journal
Paper
No.
3123.

1977,
Coronado,
California.

2
?
ABSTRACT
I
Concord
grapes
were
harvested
from
the
vine­
trifluoroethane).
Compounds
in
the
solvent
extract
vards
of
the
New
York
State
Agricultural
Experi­
mere
separated
by
gas
chromatography
and
their
f
kent
.Station,
Geneva,
during
the
19'76
vintage.
odors
evaluated
using
a
sniffing
device
attached
I
\vines
\vere
prepared
by
three
techniques
commonly
to
the
gas
chromatographic
effluent
port.
Nineteen
.­
for
producing
Concordmines
of
various
styles.
compounds
were
identified
by
combined
gas
chroma­

­­..
A
,vhite
\vine
was
prepared
from
12"
Brix
fruit
that
tographyjmass
­
spectrometry.
The­
wines
showed
pressed
immediately
after
crushing,
a
red
wine
vastly
different
varietal
characters
that
cannot
be
,
'..
as
prepared
from
16"
Brix
fruit
that
was
crushed
explained
by
variation
in
methyl
anthranilate
con­
fermented
on
the
skins,
and
another
red
wine
,
centration
alone.
Sensory
analyses
were
conducted
was
prepared
from
16"
Brix
fruit
processed
by
in
order
to
examine
the
correlation
of
odor
intensity
'
thelmal
vinification.
All
musts
were
ameliorated
in
with
methyl
anthranilate
concentration
and
total
to
produce
finished
wines
containing
12:;
volatile
concentration.
This
report.
demonstrates
that
v
,'
~
ethanol.
The
flavor
components
of
each
wine
the
volatile
composition
of
Concord
wine
differs
sig­
.
r
\\­
ere
extracted
with
Freon
113
(1,1,2­
trichloro­
1,2,2­
nificantly
with
maturity
and
processing
technique.
t
i
REVIEW
OF
LITERATURE
pounds
have
been
identified
by
Holley
et
al.
(6),
Literature
is
extensive
on
the
flavor
composition
Stevens
e
t
a].
(18)
Neudoerffer
et
a].
(lo),
and.
\
of
\f­
ine
and
other
a]
coho]
icbeverages.
The
corn­
Stern
et
d.
(16).
Particular
attention
has
been
given
pounds
identified
have
been
thoroughly
reviewed
bs­
to
the
role
of
methyl
anthranilate
in
the
varietal
i.
Kahn
(7)
and
by
Webb
and
Muller
(19).
There
has
Chal.
aCter
of
native
h
e
r
i
c
a
n
Varieties
in
work
b;
v
l
been
little
investigation,
however,
into
the
effects
of
Scott
(131,
Sale
and
U'ilson
(12),
Fufeki
(41,

.
'
the
\volatile
composition
of
table
wines.
Shaulis
and
This
report
examines
the
volatile
composition
of
;
Robinson
(
14)
noted
the
importance
of
seasonal
Concord
wine
by
instrumental
and
sensory
means
as
by
fruit
maturity
and
by
enological
tech­
­

'
'
season,
maturity,
and
fermentation
technique
on
l%
xIman
(3),
and
Nelson
et
a].
(9).

t
'hariation
on
methyl
anthranilate
concentration
in
affected
,'
Concord
and
Fredonia
grapes.
Stevens
et
al.
(17)
nique­
'.
compared
volatiles
isolated.
from
Grenache
juice
and
'
1
,
from
Grenache
Rose
wine.
Hardy
(5
)
examined
MATERIALS
AND
METHODS
:'flavor
development
in
Muscat
of
Alexandria
grapes
.
Wine
preparation:
The
experimental
wines
were
'
,,,
during
ripening,
and
Stern
et
al.
(15)
followed
cow­
.
prepared
from
40­
kg
lots
of
Concord
grapes
har­
k$?
sitional
changes
of
Zinfandel
\Tine
during
aging.
vested
during
the
19T6
\­
intage.
The
fruit
was
har­

\A,.;
The
volatile
composition
ofConcord
juice
has
vested
by
hand,
separated
from
the
steqs
and
leaves
1
studied
by
several
workers.
A
total
of
85
com­
in
a
modified
Healdsburg
stemmel­
(S
),
:and
crushed
83
.'
..

..

.
.
Am.
J.
Enol.
Vitic.,
VoL
23,
No.
2,1978
..

'!
I
­
~
~
~­~­­~
".
~
""

b
~u
I
Y
L
V
~~U
WINE
COMPOSITiON
:
4
in
a
fluted
roller­
type
crusher.
Sulfur
dioxide
was
added
as
potassium
metabisulfite
to
a
level
of
100
ppm.
All
musts
were
ameliorated
with
sucrose
to
21"
Brix
and
then
fermented
with
a
Montrachet
522
...
dry­
yeast
inoculum.
Enologicai
variables
were
as
follows:
a)
A
white
wine
was
prepared
from
Con­
cord
grapes
harvested
at
11.9"
Brix.
Immediately
Lw,
'
after
crushing,
the
juice
was
expressed
with
a
hy­
draulic
rack
and
cloth
press.
The
must
was
settled
for
12
hours,
and
the
clear
portion
was
then
racked
into
glass
fermentors
and
inoculated.
b)
A
red
wine
was
prepared
by
fermenting
crushed
16"
Brix
Con­
cord
grapes
in
contact
with
the
skin
for
five
days.
The
skins
were
then
removed,
ameliorant
was
added,
and
the
fermentation
was
completed
in
glass
fer­
mentors.
c)
The
third
sample
was
prepared
by
bringing
crushed
16"
Brix
grapes
to
60°
C
for
15
minutes
in
a
steam
kettle.
The
juice
was
then
drawn
off
and
settled
for
12
hours,
and
the
clear
portion
was
racked
into
glass
fermentors,
ameliorated,
and
inoculated.
\

Sofventextraction:
The
volatile
components
.of
the
,finished
wines
were
isolated
by
solvent
extrac­
tion
with
Freon
113
(1,1,2­
trichloro­
1,2,2­
trifluoro­
ethane).
Equal
parts
(3000
ml)
wine
and
solvent
were
stirred
for
30
minutes,
and
the
solvent
layer
was
removed,
dried
over
anhydrous
.MgSO,,
and
finally
concentrated
12,000­
fold
in
a
rotary
evapo­
rator
at
20°
C.
instrumentalanalysis:
Organoleptic
analysis
of
the
GC
effluent
was
done
on
a
Packard
800
gas
1::;
chromatograph
with
a
4­
m
x
2­
mm
glass
column
2
packed
with
10%
SP­
1000
on.
Chromosorb
W.
The
c,
sensory
character
as
each
compound
emerged
was
as­
sessed
.with
a
sniffing
device
attached
to
the
effluent
port
of
the
GC
(1).
Volatile
components
were
identi­
fied
with
a
Varian
1400
gas
chromatograph
equipped
with
a
similar
column
and
interfaced
to
a
Bendix
12
time­
of­
flight
mass
spectrometer.
The
eompo­
nents
were
quantified
with
a
Hewlett­
Packard
5830
A
gas
chromatograph
with
a
4­
m
x
2­
mm
stainless:
steel
column
and
­the
same
packing
material.
The
internal
standard
was
dodecanol.
.
.

Sensory
analysis:
Sensory
analyses
were
made
by
the
seven­
member
wine­
variety­
eyaluation
taste
panel
of
the
New
York
State
Agricultural
Experi­
ment
Station.
Two
questions
were
asked:
1)
are
there
detectable
differences
between
samples
;
and
2)
what
is
the
magnitude
of
any
differences?
Com­
pletely
randomized
triangle
tests
were
used
for
the
first
analysis,
and
scoring
on
an
unstructured
scale
was
used
for
the
second.
Several
other
samples
were
included
for
reference
:
White
Riesling,
Delaware,
Catawba,
and.
,two
synthetic
samples
prepared
by
adding
the
19
compounds
identified
by
GC/
MS
in
amounts
found
in
the
thermally
vinified
sample
to
a
solution
of
12%
v/
v
ethanol
and
0.75oJo
w/
w
,._".
tartaric
acid
in
distilled
water.
Methyl
anthranilate
i
was
omitted
from
one
of
the
synthetic
samples.
To
(\<
exclude
bias
due
to
color
differences,
pigment
iso­
.
.I
iated
from
Concord
grape
juice
was
added
to
an
samples
until
the
color
matched
the
thermally
pipi.
f
ied
sample.

RESULTS
AND
DISCUSSION
Fig.
1
shows
typical
chromatograms
of
the
&re,
Concord
extracts.
A
total
of
64
compounds
were
de.
tected
in
the
cold
pressed
white
wine
(coded
Cpw),
64
compounds
in
the
red
wine
fermented
on
the
skins
21
Fig.
1.
Chromatograms
of
12,000­
fold
essenceextracted
from
cold­
pressed
white,
fermented­
on­
skins
red,
and
thermally
vinified
red
Concord
wine.

L
y
,

Am.
J.
Enol.
Vitic.,
Voi.
29,
No.
2,
'1978
.,

..
.~
,
"..

Table
1.
Volatile
composition
ofConcord
wine.
R
"

i
Cornpound
Cold­
Fermented
Thermally
Retention
pressed
on
skins
vinified
time
(W
)
(w
l
l
)
(W
l
)
(mid
"

trt;,
4
acetate
a
Is.?
butyl
acetate
f:
hyf
butyrate
1
tsjamyl
acetate
.
b~
amyt
alcohol
Ethyl
hexanoate
i
Hexyl
acetate
Ethyl
lactate
'
i
:
.:`:
'::?.
Hexanot
..

Diethylsuccinate
Phenethyl
acetate
[
Hexanoic
acid
'.
Phenethyl
alcohol
'
Octanoic
acid
250
29
140
1400
61
00
730
ndb
280
20
370
50
1
20
1800
9000
41
0
36
340
140
5
49
1300
470
460
120
280
340
130
'
96
1900
1200
5200
4300
6300
2400
51
0
200
170
6200
8900
630
57
230
170
41
1100
640
240
860
1600
71
00
5600
4.3
7.4
8.0
10.9
13.8
14.7
16.0
18.4
18.6
19.7
21.8
28.7
29.8
34.6
34.8
37.6
43.1
­
Methyl
anthranilate
660
380
560
553
­
Decanoic
acid
2200
640
2300
57.7
*Concentration
of
compounds
calculated
ona
single­
strength
wine
basis
as
determined
with
dodecanol
used
as
the
internal
?
standard.

*
nd
=
not
detected.

Table
2.
Classification
of
.volatile
components
of
Concord
wine.

..
..
,­

i
Processing
techniques
Class
CPW
OSR
TVR
7.8
Acids
10.4
4.2
'
9.5
Alcohols
11.3
13.5
?..
:
.
..
16.2
..
.
Esters
3.9
2.2
3.6
&:.
d
Total
essence
27.4
22.3
37.1
i
Acetates
1.8
(ppml
2.4
i
.'
.

CpW,
`cold­
pressed
white;
OSR,
on­
the­
skinsred;
N
R
,
ther­
mally
vinified
red.
..
.,

..
Am.
3.
Enol.
Vitic.,
CONCORD
WINE
COMPOSlTlON­
85
triangle
tests.
The
results
are
given
in
Table
3.
Six
'

of
the
seven
panelists
could
successfully
distinguish
CPW
from
OSR,
all
seven
panelists
could
distinguish
OSR
from
TVR,
and
five
panelists
could
distin­
guish
CPW
from
TVR.
The
respective
results
are
significant
at
p
=
0.01,
p
=
0.001,
and
p
=
0.05
(2).
The
intensity
of
typical
"American"
or
"la­
brusca"
character
was
investigated
by
scoring
on
an
unstructured
scale
(Fig.
2)
from
0,
no
detectable
"labrusca"
character,
to
20,
extremely
intense
"la­
brusca"
character.
The
compiled
data
were
evalu­
ated
by
a
two­
way
analysis
of
variance
followed
by
Duncan's
multiple­
range
test.
The
results
are
shown
in
Table
4.
It
is
apparent
that
the
white
wine
made
from
low
Brix
grapes
is
significantly
lower
in
"la­
brusca"
character
than
the
other
Concord
samples.
Although
it
is
not
surprising
that
the
Catawba
wine
was
scored
relatively
high
in
"labrusca"
char­
acter,
it
is
interesting
that
the
methyl
anthranilate
concentration
of
that
sample
was
less
than
0.1
ppm.
It
can
also
be
seen
that
the
synthetic
wines
were
poor
imitations
of
"1abrusca"­
flavored
wines,
re­
gardless
of
the
presence
or
absence
of
methyl
anthranilate..
Statistically,
the
CPW
wine
has
no
more
"labrusca".
character
than
the
Delaware
or
the
White
Riesling
wines.
That
appears
to
confirm
the
thought
that
the
compounds
responsible
for
Concord
varietal
character
are
formed
during
the
latter
stages
of
maturation.
Although
Robinson
and
Shaulis
(11)
showed
that
to
be
true
for
methyl
anthranilate
formation,
this
work
indicates
that
it
is
not
responsible
for
varietal
character.
Although
both
red
wines
have
distinctive.
``
la­
brusca"
character,
they
are
quite
different
from
each
other.
The
compounds
responsible
for
the
difference
have
not
yet
been
positively
identified.
It
seems,
however,
that
the
extremely
high
level
of
acetates
in
the
thermally
vinified
wine,
particularly
isobutyl,
isoamxl,
and
phenethyl,
may
play
a
role.
Table
5
lists
the
acetates
detected
and
the
relative
amounts
found
in
the
two
red
Concord
wines.
Each
com­
pound
has
a
strong
fruity
or
spicy
aroma
that
could
contribute
to
the
character
of
the
thermally
vinified
wine.

Table
3.
Varietal
character:
difference
test.

(Randomizedtriangletests)
Comparison
%
correctresponse
Level
of
significance
CPWa
vs.
OSRb
86
.01
CPW
vs.
TVRc
71
.05
OSR
vs.
TVR
loo
.
.001
a
Cold
pressed
white
wine.

Red
wine
fermented
in
contact
with
skins.

e
Thermally
vinified
red
wine.

I
Vol.
29,
No.
2,1978
..
~~
~~

..
..

..
86­
CONCORDWINECOMPOSITION
­

Table
4.
Analysis
of
variance
and
Duncan's
multiple­
range
test.

Sample
Treatment
Mean
Concord
Therm.
vinif.
14.43
Concord
Ferm.
on
skins
12.36
Catawba
Cold
press
11.36
Concord
Cold
press
7.07
Delaware
Cold
press
6.21
Riesling
Cold
press
5.64
Synthetic
No
met.
anth.
5.07
Synthetic
With
met.
anth.
1.93
­____

Error
df
=
42,
Error
ms
=
30.1477,
F
=
4.2000,
LSD
=
6.0397.

In
conclusion,
the
thermally
vinified
wine
had
a
higher
odor
intensity
than
the
other
wines.
It
also
had
a
higher
level
of
total
extractable
volatiles.
Even
so,
the
white
wine
was
significantly
lower
in
odor
intensity
yet
had
a
higher
level
of
extractable
volatiles
than
the
wine
fermented
on
the
skins.
The
lower
level
of
total
volatiles
in
the
sample
fermented
on
the
skins
did
not
appear
to
affect
odor
intensity
significantly.
Thermal
vinification,
through
the
production
of
high
levels
of
acetates
or
other
compounds,
yields
a
wine
with
maximunl
intensity
of
Concord
aroma.
It
can
be
concluded,
however,
that
the
Concord
varietal
character
is
composed
of
­one
01­
more
trace
compo­
nents
that
have
yet
to
be
identified
and
that
these
compounds
are
formed
during
the
latter
stages
of
maturation.
.
The
nineteen
compounds
identified,
including
methyl
anthranilate,
constituted
over
90%
of
the
total
extractable
essence.
These
compounds
certainly
contribute
to
the
"\*
inous"
character
of
Concord
wine
but
make
little
01­
no
contribution
to
Concord
varietal
character.
Identification
of
the
compounds
responsible
for
the
varietal
character
in
Concord
and
other
wine
varieties
will
rest
yith
the
identifica­
tion
of
trace
components
present
in
pg/
1
quantities.

ODOR
EVALUATION
OF
''AMERICAN
CHARACTER
Taster
Date
Directions
­
Examine
the
odor
of
each
wipe:
Rate
the
intensity
of
typical
"American"
character
according
to
your
own
defini­
tion
and
experience
by
placing
a
vertical
line
at
the
appropriate
position
on
the
scale
provided..

Sample
"American"
Low
High
71
23
"

44
60
Fig.
2.
Unstructured
scaleused
in
scoring
intensity
of
"labrusca"
odor
in
experimental
wines.
Responses
were
con­
verted
for
statistical
analysis
using
a20­
pt
grid.
1
1
Table
5.
Relative
acetate
concentration
in
red
Concord
wine.

CompoundFerrn.
on
skins
Therm.
vinification
X
in,­;
~

Ethyl
acetate
­"­.
38
Isobutyl
acetate
50
200
300
Isoamyl
acetate
1
E?
6200
244
Hexyl
acetate
36
57
96
58
Phenethyl
acetate
860
796
"

370
510
\

LITERATURE
CITED
1.
Acree,
T.
E.,
R.
M.
Butts,
R.
R.
Nelson,
and
C.
y.
L
~~,
,Sniffer
to
determine
the
odor
of
gas
chromatographic
efil,,.
ents.
Anal.
Chem.
48:
12,1821
(1976).
2.
Amerine,
M.
A.,
and
E.
B.
Roessler.
Wines:
Their
SensorV
Evaluation.
W.
H.
Freeman
and
Co.,
San
Francisco.
Appendix
D,
p.
181
(1976).
3.
Friedman,
t.
E.
Concord
grape
quality.
N.
Y.
S.
Nortic.
sot.
Proc.
121:
132­
6
(1976).
4.
Futeki,
T.
Methyl
anthranilate
and
total
volatile
estero
content
of
grape
cultivars
grown
in
Ontario.
hpubl.
Paper
presented
at
the,
Annual.
Meeting
of
the
American
Society
of
Enologists,
EasternSection,
Erie,
Pennsytvania,
August,
1976.
5.
Hardy,
P.
J.
Changes
in
volatiles
of
Muscat
grapes
dur­
ing
ripening.
Phytochemistry
9:
709­
75
(1970).
6.
Holley,
R.
W.,
B.
Stoyla,
and
A.
D.
Holley.
The
identifica.
tion
of
some
volatile
constituents
of
Concord
grape
juice.
Food
Res.
20:
326­
30
(1955).
7.
Kahn,
J.
H.
Compounds
identified
in
whiskey,
wine,
and
.
beer:
a
tabuiation.
J.
Assoc.
Off.
Anal.
Chem.
52:
1766­
78(
1969).
8.
Moyer,
J­
C.
An
experimental
grape
stemmer.
Farm
Res.
23:
1,15
(1957).
9.
Nelson,
R.
R.,
T.
E.
Acree,
C.
Y.
Lee,
and
R.
M.
Butts.,
Methyl
anthranilate
as
an
aroma
constituent
of
American
wine.
J.
Food
Sci.
42:
57­
9
(1977).
IO.
Neudoerifer,
T.
S.,
S.
Sandler,
E.
Zubeckis,
and
M.
D.
Smith.
Detection
of
an
undesirable
anomaly
in
Concord
grape
i
by
gaschromatography.
J.
Agric.
FoodChem,
13(
6):
584­
8
(1965).
11.
Robinson,
W.
B.,
and
N.
J.
Shaulis.
Ripening
studies
of
grapes
grown
in
1948
for
juice
manufacture.
Fruit
Prod.
J.
Am.
.
Food
Manuf.
29:
2.36­
7,54,62
(1949).
12.
Sale,
J.
W.,
and
J.
8.
Wilson.
Distribution
of
volatile
flavor
in
grapes
and
grapejuices.
J.
Agric.
Res.
33:
307­
10
13.
Scott,
R.
D.
Methyl
anthranilate
in
grape
beverages
and
flavors.
Ind.
Eng.
Chem.
15:
732­
3
(1923).
14.
Shaulis,
N.
J.,
and
W.
B.
Robinson.
The
effect
of
season,
i
pruning
severity,
and
trellising
on
some
chemical
character­
­:
istics
of
ConcordandFredonia
grape
juice.
Proc.
Am.
Hortic.
.

15.
Stern,
0.
J.,
G.
Guadagni,
and
K.
L.
Stevens.
Agingof
wine:
changes
in
Zinfandel
volatiles.
Am.
J.
Enol.
Vitic.
26:

16.
Stern,
0.
J.,
A.
Lee,
W.
H.
McFadden,
and
K.
L.
Stevens.
Volatiles
from
grapes:
identification
of
volatiles
from
Concord
essence.
J.
Agric.
Food
Chem.
15:
llOO­
3
(1967).
17.
Stevens,
K.
L.,
R.
A.
Flath,
L.
Alson,
and
D.
J.
Stern.
Volatiles
from
grapes:
comparison
of
Grenache
juice
and
Grenache
Rosewine.
J.
Agric.
Food
Chem.
17:
1102­
6(
1969).
18.
Stevens,
K.
L.
A.
Lee,
W.
H.
McFadden,
and
R.
Teranishi.
Volatiles
.from
grapes:
some
volatiles
from
Concord
essence.
J
Food
Sei.
30:
1106­
7
(1965).
19.
Webb,
A
D.,
and
C.
J.
Muller.
Volatile
aromacompo­
nents
of
wines
and'
other
fermented
beverages.
Adv.
&PI.
Microbiol.
15:
75­
146,(
1972).
(1926):

SOC.
62:
214­
20
(1953).

208­
13
(1975).
1
Am.
J.
Enol.
Vitic.,
Vol.
29,
No.
2,1978
..

..
:..
.
_...
­.
.
..
.
x
..
..
.
.
..
.
.
.
­
.
.
..,_
2
The
Odor
Quality
o
f
Labrusca
Grapes
TERRY
E.
ACREE
New
York
StateAgricultural
Experiment
Station,
Cornell
University,
Geneva.
NY
14456
C
u
l
t
i
v
a
r
s
o
f
t
h
e
s
p
e
c
i
e
s
vinifera
are
the
most
p
r
e
v
a
l
e
n
t
g
r
a
p
e
s
p
l
a
n
t
e
d
i
n
­t
h
e
w
o
r
l
d
.
O
r
i
g
i
n
a
l
l
y
grown
i
n
Europe
and
Asia
.
v
i
n
i
f
e
r
a
g
r
a
p
e
s
.
o
f
w
h
i
c
h
,t
h
e
r
e
a
r
e
s
e
v
e
r
a
l
thousand
named
c
u
l
t
i
v
a
r
s
(i
).
a
l
s
o
d
o
m
i
n
a
t
e
t
h
e
v
i
t
i
c
u
l
t
u
r
e
of
the
New
Vorld.
In
North
&erica.
however,
two
o
t
h
e
r
s
p
e
c
i
e
s
of
(h
b
r
u
s
u
,
Bailey
and
L
p
t
u
n
d
i
f
o
u
.
Hicha'ux)
are
growo
i
n
s
i
g
n
i
f
i
c
a
n
t
q
u
a
n
t
i
t
i
e
s
(?).
The
s
e
v
e
r
e
c
l
i
r
c
a
t
e
a
n
d
v
i
r
u
l
e
n
t
d
i
s
e
a
s
e
i
n
c
e
n
t
r
a
l
and
e
a
s
t
e
r
n
N
o
r
t
h
America
made
early
c
u
l
t
i
v
a
t
i
o
n
o
f
v
i
n
i
f
e
r
a
g
r
a
p
e
s
l
a
r
g
e
l
y
m
s
u
c
c
e
s
s
f
u
l
,
a
t
l
e
a
s
t
i
n
t
h
e
E
n
g
l
i
s
h
C
o
l
o
n
i
e
s
.
T
h
i
s
s
t
i
m
l
a
t
e
d
t
h
e
h
y
b
r
i
d
i
z
a
t
i
o
n
and
c
u
l
t
i
v
a
t
i
o
n
o
f
t
h
e
c
o
r
e
t
o
l
e
r
a
n
t
n
a
t
i
v
e
g
r
a
p
e
s
p
e
c
i
e
s
f
o
r
many
years.
With
t
h
e
d
e
v
e
l
o
p
e
c
t
of
n
e
t
h
o
d
s
f
o
r
d
i
s
e
a
s
e
c
o
n
t
r
o
l
acd
the
expansion
of
v
i
t
i
c
c
l
t
u
r
e
i
n
t
o
c
l
i
m
a
t
e
s
more
amenable
t
o
.v
i
n
i
f
e
r
a
g
r
a
p
e
s
,
t
h
e
p
e
r
c
e
n
t
o
f
n
a
t
i
v
e
s
p
e
c
i
e
s
h
a
s
d
e
c
r
e
a
s
e
d
­
t
o
l
e
s
s
t
h
a
n
f
i
v
e
p
e
r
c
e
n
t
of
the
total
Korth
American
.grape
production
(2
).
Alth6uL.
h
dwarfed
by
t
h
e
s
i
z
e
of
t
h
e
v
i
n
i
f
e
r
a
g
r
a
p
e
c
r
o
p
,
i
n
excess
of
4.000.00@
t
o
n
s
,
t
h
e
p
r
o
d
u
c
t
i
o
n
af
labrusca
grapes
has
increased
i
n
the
last
30
y
e
a
r
s
.
T
h
i
s
c
o
n
t
i
n
u
e
d
d
e
n
a
c
d
f
o
r
l
a
b
r
u
s
c
a
g
r
a
p
e
s
is
due
t
o
t
h
e
i
r
s
u
p
e
r
i
o
r
q
u
a
l
i
t
y
for
the
production
of
grape
juice
and
j
e
l
l
y
.
I
t
is
t
h
e
u
n
i
q
u
e
f
l
a
v
o
r
o
f
labrusca
grapes.
and
i
n
p
a
r
t
i
c
u
l
a
r
t
h
e
c
u
l
t
i
v
a
r
Concord;
that
i
s
r
e
s
p
o
n
s
i
b
l
e
f
o
r
t
h
e
i
r
s
u
p
e
r
i
o
r
i
t
y
.
I
n
f
a
c
t
.
t
h
e
.
f
l
a
v
o
r
o
f
labrusca
grapes
has
.
become
t
h
e
s
t
a
n
d
a
r
d
of
i
d
e
n
t
i
t
y
€o
r
g
r
a
p
e
juice
and
j
e
l
l
y
i
n
n
o
r
t
h
America.
TtiE
m
O
R
OF
LaBBllscBsBhEEs
The
major
flavor
differences
among
grape
prcducts
nade
with
d
i
f
f
e
r
e
n
t
s
p
e
c
i
e
s
o
f
g
r
a
p
e
s
a
r
e
f
o
u
n
d
i
n
t
h
e
o
l
o
r
.
It
is
e
a
s
y
t
o
d
i
s
t
i
n
g
u
i
s
h
t
h
e
o
d
o
r
s
of
g
r
a
p
e
j
u
i
c
e
s
made
from
vinifera.
l
a
b
r
u
s
c
a
.
a
n
$
m
u
s
c
a
d
i
n
e
s
p
e
c
i
e
s
.
I
n
c
o
n
t
r
a
s
t
,
t
h
e
t
a
s
t
e
of
g
r
a
p
e
s
,
t
h
a
t
is
t
h
e
i
r
s
w
e
e
t
n
e
s
s
.
.
sourness,
and
b
i
t
t
e
r
n
e
s
s
.
is
frequently
manipulated
by
p
r
o
c
e
s
s
o
r
s
t
o
t
h
e
e
x
t
e
n
t
t
h
a
t
a:;
y
t
z
s
t
e
difference
among
grapes
of
l
i
f
f
e
r
e
n
t
s
p
e
c
i
e
s
a
r
e
o
b
l
i
t
e
r
a
t
e
d
.
The
one
word
which
has
been
used
€o
r
c
e
n
t
u
r
i
e
s
t
o
'
d
e
s
c
r
i
b
e
t
h
e
o
d
o
r
c
h
a
r
a
c
t
e
r
o
f
n
a
t
i
v
e
g
r
a
p
e
s
is
foxy".
In
f
a
c
t
e
a
r
l
y
..

009~­
6~
5~/~
1/
0170­~
01
lSOS.
OO/
O
0
1981
American
Chemical
Society
12
QUALITY
OF
SELECTED
FRUITS
AND
VEGETABLES
2.
ACREt
European
v
i
s
i
t
o
r
s
t
o
t
h
e
New
World
named
t
h
e
n
a
t
i
v
e
l
a
b
r
u
s
c
a
g
r
a
p
e
s
g
r
o
w
i
n
g
i
n
New
England
the
Northern
Fox
G
r
a
p
e
,
a
n
d
t
h
e
grapes
growing
i
n
t
h
e
S
o
u
t
h
t
h
e
S
o
u
t
h
e
r
n
Fox
Grape.
The
o
r
i
g
i
n
a
l
meaning
of
t
h
e
word
"foxy"
was
e
x
p
l
i
c
i
t
l
y
s
t
a
t
e
d
i
n
1722
by
Robert
Beverly
when
h
e
d
e
s
c
r
i
b
e
d
the
smell
o
f
.t
h
e
s
e
g
r
a
p
e
s
a
s
resembling
t
h
a
t
o
f
a
fox(&).
This
musky
a
n
i
m
a
l
­l
i
k
e
a
r
o
n
a
is
s
t
i
l
l
c
o
n
s
i
d
e
r
e
d
a
n
u
n
d
e
s
i
r
a
b
l
e
a
t
t
r
i
b
u
t
e
i
n
&r
a
p
e
s
.
b
u
t
"f
o
x
y
"
m
u
s
t
not
b
e
c
o
n
f
u
s
e
d
w
i
t
h
t
h
e
p
o
w
e
r
f
u
l
f
r
u
i
t
y
I
f
l
o
r
a
l
,
o
r
candy
o
d
o
r
s
e
s
s
e
n
t
i
a
l
t
o
t
h
e
q
u
a
l
i
t
y
o
f
g
r
a
p
e
j
u
i
c
e
a
n
d
j
e
l
l
y
.
Even
t
h
o
u
g
h
t
h
e
s
e
o
d
o
r
s
may
n
o
t
b
e
a
p
p
r
e
c
i
a
t
e
d
i
n
most
wine
types,
they
are
n
o
t
t
h
e
same
as
f
o
x
i
n
e
s
s
.
T
h
e
c
u
l
t
i
v
a
r
s
of
w
r
u
s
c
m
(Concord,
Catawba.
Delaware,
etc.)
were
appreciated
by
e
a
r
l
y
A
m
e
r
i
c
a
n
g
r
a
p
e
b
r
e
e
d
e
r
s
b
e
c
a
u
s
e
o
f
t
h
e
i
r
g
e
n
e
r
a
l
l
a
c
k
of
foxiness.
However,
Niagara
i
s
t
h
e
o
n
e
r
e
m
a
i
n
i
n
g
.
counercial
c
u
l
t
i
v
a
r
w
h
i
c
h
h
a
s
a
l
v
a
y
s
b
e
e
n
c
o
n
s
i
d
e
r
e
d
f
o
x
y
.
T
h
e
r
e
f
o
r
e
,
f
o
x
i
n
e
s
s
s
h
o
u
l
d
n
o
t
b
e
c
o
n
s
i
d
e
r
e
d
t
h
e
d
o
m
i
n
a
n
t
o
d
o
r
q
u
a
l
i
t
y
o
f
L
n
b
r
u
s
c
a
g
r
a
p
e
s
.
I
n
t
h
i
s
p
a
p
e
r
,
t
h
a
t
o
d
o
r
q
u
e
l
i
t
y
c
o
m
o
n
t
o
611
f
a
b
r
u
s
c
a
g
r
a
p
e
s
will
be
r
e
f
e
r
r
e
d
t
o
a
s
"l
a
b
r
u
s
c
a
c
h
a
r
a
c
t
e
r
.
D
u
r
i
n
g
t
h
e
s
e
n
s
o
r
y
s
t
u
d
i
e
s
o
f
h
u
n
d
r
e
d
s
of
d
i
f
f
e
r
e
n
t
g
r
a
p
e
c
u
l
t
i
v
a
r
s
c
o
n
d
u
c
t
e
d
a
t
t
h
e
E
x
p
e
r
i
m
e
n
t
S
t
a
t
i
o
n
i
n
Geneva,
New
`iurk.
p
a
n
e
l
i
s
t
s
h
a
v
e
,
w
i
t
h
g
r
e
a
t
r
e
g
u
l
a
r
i
t
y
,
i
d
e
n
t
i
f
i
e
d
t
h
e
presence
of
l
a
b
r
u
s
c
a
c
h
a
r
a
c
t
e
r
i
n
t
h
e
o
d
o
r
of
l
a
b
r
u
s
c
a
g
r
a
p
e
s
and
e
[
t
e
n
d
o
t
h
e
s
a
n
e
w
i
t
h
h
y
b
r
i
d
c
r
o
s
s
e
s
b
e
t
w
e
e
c
y.
b
b
r
u
s
u
and
cther
g
r
a
p
e
s
p
e
c
i
e
s
.
F
u
r
t
h
e
m
o
r
e
,
t
h
e
y
c
a
n
d
e
t
e
c
t
t
h
e
p
r
e
s
e
c
c
e
o
f
s
p
e
c
i
f
i
c
o
d
o
r
­
c
o
n
p
o
n
e
n
t
s
,
w
h
i
c
h
i
n
v
a
r
y
i
n
g
d
e
g
r
e
e
s
,
seem
t
o
make
L
I
~
'
t
h
e
t
o
t
a
l
s
e
n
s
o
r
y
e
f
f
e
c
t
.
F
o
u
r
d
e
s
c
r
i
p
t
o
r
s
w
h
i
c
h
are
irequently
used
by
t
h
e
s
e
p
a
p
e
l
i
s
t
s
a
r
e
f
o
x
y
­,"f
l
o
r
a
l
".
n
e
t
h
y
l
~n
t
h
r
a
n
i
l
a
t
e
­l
i
k
e
'
*,
and
"cotton
candy".
It
is
c
e
r
t
a
i
n
l
y
u
n
w
i
s
e
cia
a
s
s
u
m
e
t
h
a
t
t
h
e
s
e
o
d
o
r
c
o
m
p
o
n
e
n
t
s
a
r
e
.
i
n
every
c
a
s
e
.
r
e
l
a
t
e
d
t
o
s
i
n
g
l
e
c
h
e
m
i
c
a
l
s
p
e
c
i
e
s
.
b
u
t
t
h
e
y
a
r
e
p
r
o
b
a
b
l
y
less
c
o
m
p
l
i
c
a
t
e
d
i
n
t
h
e
i
r
c
t
m
i
s
t
r
y
t
h
a
n
t
h
e
conpound
mix
t
h
a
t
p
r
o
d
u
c
e
s
t
h
e
t
o
t
a
l
p
e
r
c
e
p
t
i
o
n
o
f
l
a
b
r
u
s
c
a
c
h
a
r
a
c
t
e
r
.
Such
d
e
s
c
r
i
p
t
o
r
s
are
u
s
e
f
u
l
i
n
o
u
r
a
t
t
e
m
p
t
s
t
o
s
o
r
t
o
u
t
t
h
e
s,,
alI
number
of
odor­
active
compounds
present
i
n
n
a
t
u
r
o
l
products.
However,
they
have
l
i
t
t
l
e
m
e
a
n
i
n
g
f
r
o
n
o
n
e
l
a
b
o
r
a
t
o
r
y
t
o
t
h
e
n
e
x
t
.
F
o
r
e
x
a
m
p
l
e
.
t
h
e
a
r
o
m
a
d
e
s
c
r
i
b
e
d
i
n
o
u
r
v
o
r
k
as
cotton
candy"
appears,
for
r
e
a
s
o
n
s
w
h
i
c
h
w
i
l
l
b
e
e
x
p
l
a
i
n
e
d
l
a
t
e
r
,
t
o
b
e
d
u
e
t
o
t
h
e
same
compoufid
responsible
f
o
r
t
h
e
s
t
r
a
w
b
e
r
r
y
o
d
o
r
d
e
t
e
c
t
e
d
i
n
t
h
e
l
a
b
o
r
a
t
o
r
y
of
Rapp(
j)
in
Gemany.
Tne
c
o
n
f
u
s
i
o
n
t
h
e
s
e
n
o
n
­c
h
e
m
i
c
a
l
d
e
s
c
r
i
p
t
o
r
s
c
r
e
a
t
e
,
w
i
l
l
be
ninimized
once
WE
know
t
h
e
c
a
u
s
a
t
i
v
e
a
g
e
n
t
s
for
o
d
o
r
p
e
r
c
e
p
t
i
o
n
.
Then
we
can
use
a
chemical
name
t
o
d
e
s
c
r
i
b
e
t
h
a
t
p
e
r
c
e
p
t
i
o
n
.
In
t
i
t
&
meantime,
we
must
t
o
l
e
r
a
t
e
t
o
some
e
x
t
e
n
t
t
h
e
u
s
e
o
f
t
h
e
s
e
v
o
r
d
s
i
n
o
u
r
d
a
y
­t
o
­d
a
y
r
e
s
e
a
r
c
h
.
,IETHYL
AHTHRAFI~
Studies
Gf
t
h
e
v
o
l
a
t
i
l
e
c
o
m
p
o
s
i
t
i
o
n
of
grapes
have,
through
the
y
e
a
r
s
,
r
e
v
e
a
l
e
d
t
h
e
p
r
e
s
e
c
c
e
of
so
many
odor­
active
coapocnds
t
h
a
t
i
t
is
v
e
r
y
u
n
l
i
k
e
l
y
t
h
a
t
a
s
i
n
g
l
e
compound
i
s
responsible
for
more
than
a
f
e
w
p
e
r
c
e
n
t
o
f
t
h
e
t
o
t
a
l
o
d
o
r
c
h
a
r
a
c
t
e
r
o
f
g
r
a
p
e
s
,*
f
any
kind(
h).
Furthermore,
among
t
h
e
h
u
n
d
r
e
d
s
of
v
o
l
a
t
i
l
e
s
..

..
..

..
present
o
d
o
r
a
c
t
present
together
c
u
l
t
i
v
a
r
a
specif
One
e
v
e
r
a
s
s
)
(1).
h
i
produce
i
n
g
r
a
p
e
anthrani
3
abrus
ca
i
n
o
n
l
y
nethyl
a
t
h
i
s
co
t
h
e
l
a
b
r
labrusca
Ha
n
t
o
have
o
d
o
r
t
b
r
to
v
i
n
i
labrusca
There
a
propyl,
i
n
l
a
b
r
u
contribu
placed
For
exarn
i
n
d
i
c
a
t
e
t
h
e
q
u
a
l
t
h
e
p
e
r
t
h
e
anou
e
s
t
e
r
c
breeders
odor
cha
One
preseccc
products
f
l
a
v
o
r
s
of
flavc
unintent
FOXIKES$

f
r
o
n
1
2
been
ur
"2
­
has
an
grapes.
I
n
so
abrusca
s
n
d
t
h
e
e.
The
in
1722
ipeS
a
s
ma
i
s
candy
.
Even
types
.
ruscana
e
a
r
l
y
ack
of
­r
e
r
c
i
a
l
r
e
f
ore,
i
t
y
o
f
,
t
o
a
l
l
grape
Jar
New
zd
t
h
e
pes
and
and
x
c
e
o
f
3
make
c11
are
..
foxy­

0.

t
h
a
t
u
t
.
t
h
e
:la
t
urz
1
s
r
a
t
o
r
y
.roGk
as
plained
i
o
r
t
h
e
.en;;
any.
.
i.
11
be
epdion.
n.
'
I
n
f
t
h
e
s
e
2.
ACREE
Odor
Quality
of
Lahrrtsca
Grapes
13
p
r
e
s
e
n
t
i
n
g
r
a
p
e
s
(F
i
g
u
r
e
I.)*
only
a
very
feK
probably
have
any
o
d
o
r
a
c
t
i
v
i
t
y
.
T
h
e
r
e
m
u
s
t
e
x
i
s
t
a
number
of
c
h
m
i
c
a
l
compounds
p
r
e
s
e
n
t
i
n
g
r
a
p
e
s
a
t
p
a
r
t
i
c
u
l
a
r
c
o
n
c
e
n
t
r
a
t
i
o
n
s
t
h
a
t
when
taken
together
produce
t
h
e
c
h
a
r
a
c
t
e
r
i
s
t
i
c
o
d
o
r
q
c
a
l
i
t
y
o
f
a
p
a
r
t
i
c
u
l
a
r
c
u
l
t
i
v
a
r
.
A
s
s
o
c
i
a
t
e
d
w
i
t
h
e
a
c
h
of
t
h
e
s
e
o
d
o
r
­a
c
t
i
v
e
c
h
e
m
i
c
a
l
s
is
a
s
p
e
c
i
f
i
c
.
though
n
o
t
n
e
c
e
s
s
a
r
i
l
y
u
n
i
q
u
e
,
o&
or
q
u
a
l
i
t
y
.
One
such
compound
i
s
m
e
t
h
y
l
a
n
t
h
r
a
n
i
l
a
t
e
,
t
h
e
f
i
r
s
t
conpound
ever
associated
w
i
t
h
o
d
o
r
c
h
a
r
a
c
t
e
r
of
a
p
a
r
t
i
c
c
l
a
r
g
r
a
p
e
s
p
e
c
i
e
s
(2
).
O
r
i
g
i
n
a
l
l
y
i
d
e
n
t
i
f
i
e
d
i
n
n
e
r
o
l
i
o
i
l
(8)
it
vss
u
s
e
d
t
o
produce
synthetic
grape­
flavored
products
before
i
t
vas
observed
i
n
g
r
a
p
e
s
.
In
1923
S
a
l
e
a
n
d
W
i
l
s
o
n
(2
)
a
n
a
l
y
z
e
d
t
h
e
m
e
t
h
y
l
a
n
t
h
r
a
n
i
l
a
t
e
c
o
n
t
e
n
t
i
n
55
c
u
l
t
i
v
a
r
s
of
g
r
a
p
e
s
.
i
n
c
l
u
d
i
n
g
b
o
t
h
labrusca
and
v
i
n
i
f
e
r
a
c
u
l
t
i
v
a
r
s
.
They
f
o
u
n
d
m
e
t
h
y
l
a
n
t
h
r
a
n
i
l
a
t
e
i
n
,
o
n
l
y
14
of
t
h
e
s
e
.
I
n
1976
using
gas
chromatography.
we
found
m
e
t
h
y
l
a
n
t
h
r
a
n
i
l
a
t
e
i
n
only
8
of
4
5
c
u
l
t
i
v
a
r
s
(m).
Certainly
t
h
i
s
compound
p
l
a
y
s
a
n
i
m
p
o
r
t
a
n
t
r
o
l
e
i
n
t
h
e
f
l
a
v
o
r
of
sone
of
t
h
e
l
a
b
r
u
s
c
a
g
r
a
p
e
s
b
u
t
i
t
i
s
n
o
t
­
s
o
l
e
l
y
r
e
s
p
o
n
s
i
b
l
e
f
o
r
t
h
.e
l
a
b
r
u
s
c
a
c
h
a
r
a
c
t
e
r
.
Nany
v
i
n
e
s
w
i
t
h
a
s
t
r
o
n
g
l
a
b
r
u
s
c
a
c
h
a
r
a
c
t
e
r
h
a
v
e
b
e
e
n
f
o
u
n
d
t
o
have
a
methyl
a
n
t
h
r
a
n
i
l
a
t
e
c
o
n
t
e
n
t
vel1
below
t
h
e
a
p
p
a
r
e
n
t
odor
threshold
(m).
F
u
r
t
h
e
r
n
o
r
e
,
t
h
e
a
d
d
i
t
i
o
n
o
f
t
h
i
s
compound
t
o
v
i
n
i
f
e
r
a
v
i
n
e
s
a
t
v
e
r
y
h
i
g
h
c
o
n
c
e
n
t
r
a
t
i
o
n
s
d
o
e
s
n
o
t
p
r
o
d
u
c
e
a
labrusca
aroma
n
o
r
d
o
e
s
i
t
produce
a
v
i
n
e
w
i
t
h
any
foxiness.
There
are
small
amounts
of
o
t
h
e
r
a
n
t
h
r
a
n
i
l
a
t
e
e
s
t
e
r
s
.
e
t
h
y
l
.
propyl,
=.­
butyl
and
the
analogous
compound
p­
aminoacetophenone
i
n
l
a
b
r
u
s
c
a
g
r
a
p
e
s
b
u
t
t
h
e
r
e
are
n
o
d
a
t
a
e
x
p
l
a
i
n
i
n
g
t
h
e
i
r
precise
contribution,
i
f
any,
t
o
t
h
e
=e
t
h
y
l
a
n
t
h
r
a
n
i
l
a
t
e
­l
i
k
e
aroma.
I
n
t
h
e
l
a
s
t
60
y
e
a
r
s
,
p
e
r
h
a
p
s
t
o
o
much
e
q
h
a
s
i
s
has
beer.
placed
on
t
h
e
m
e
t
h
y
l
a
n
t
h
r
a
n
i
l
a
t
e
c
o
n
t
e
n
t
of
l
a
b
r
u
s
c
a
g
r
a
p
e
s
.
For
example.
ueasurements
of
t
h
i
s
cornpour.
6
heve
beer.
used
t
o
i
n
d
i
c
a
t
e
g
r
a
p
e
n
a
t
u
r
i
t
y
(u
);
it
has
been
used
as
a
n
i
n
d
i
c
z
t
o
r
of
t
h
e
q
u
a
l
i
t
y
o
f
g
r
a
p
e
p
r
o
d
u
c
t
s
(u
),
and
as
z
means
of
nonitorinL
t
h
e
p
e
r
f
o
r
m
a
n
c
e
o
f
e
s
s
e
n
c
e
r
e
c
o
v
e
r
y
e
q
u
i
p
z
e
n
t
(u
).
F
u
r
t
h
e
r
m
o
r
e
,
t
h
e
amount
of
methyl
.
a
n
t
h
r
a
n
i
l
a
t
e
c
o
m
b
i
n
e
d
.
w
i
t
h
t
h
e
v
o
l
a
t
i
l
e
e
s
t
e
r
c
o
n
t
e
c
t
of
grapes
is
p
r
e
s
e
n
t
l
y
b
e
i
n
g
used
t
o
a
i
d
t
h
e
g
r
a
p
e
b
r
e
e
d
e
r
s
i
n
t
h
e
i
r
a
t
t
e
m
p
t
t
o
p
r
o
d
u
c
e
new
c
u
l
t
i
v
a
r
s
w
i
t
h
c
e
r
t
a
i
n
o
d
o
r
c
h
a
r
a
c
t
e
r
i
s
t
i
c
s
(&).
One
of
t
h
e
d
a
n
g
e
r
s
o
f
p
l
a
c
i
n
g
t
o
o
much
inportance
on
the
p
r
e
s
e
n
c
e
o
f
m
e
t
h
y
l
a
n
t
h
r
a
n
i
l
a
t
e
i
n
g
r
a
p
e
p
r
o
t
u
c
t
s
i
s
t
h
a
t
s
u
c
h
products
may
tend
t
o
snell
more
l
i
k
e
t
h
e
s
i
m
p
l
e
i
m
i
t
z
t
i
o
n
g
r
a
p
e
f
l
a
v
o
r
s
r
a
t
h
e
r
t
h
a
n
t
h
e
n
a
t
u
r
a
l
p
r
o
d
u
c
t
.
I
t
i
s
one
of
t
h
e
g
o
a
l
s
of
f
l
a
v
o
r
r
e
s
e
a
r
c
h
t
o
p
r
e
v
e
n
t
s
u
c
h
i
r
o
n
i
e
s
f
r
o
e
o
c
c
b
r
r
i
n
g
unintentionally.
FOXINESS
So
f
a
r
o
u
r
a
t
t
e
m
p
t
s
to
i
s
o
l
a
t
e
L?
pure
chemical
component
f
r
o
m
l
a
b
r
u
s
c
a
o
r
a
u
s
c
a
d
i
n
e
g
r
a
p
e
s
.
w
i
t
h
a
c
l
e
a
r
f
o
x
y
odor
have
b
e
e
n
u
n
s
u
c
e
s
s
f
u
l
.
T
h
e
r
e
is,
h
o
w
e
v
e
r
r
o
n
e
c
c
n
p
o
n
e
n
t
,
grans­
2­
texen­
1­
01.
apparently
present
in
a
l
l
g
r
a
p
e
s
p
e
c
i
e
s
.
t
h
a
t
h
a
s
a
n
o
d
o
r
.
v
e
r
y
siniilar
t
o
t
h
e
f
o
x
y
oZor
cooponect
of
Kiagara
g
r
a
p
e
s
.
P
o
s
s
i
b
l
y
.
t
h
i
s
c
o
m
p
o
u
n
d
.
i
s
p
r
e
s
e
n
t
i
n
f
o
x
y
­s
n
e
l
l
i
n
g
14
<
..
"
."
7
I
31VLllN33NO3
A
000'tl
NO113VUJ
*%
E
2.
ACRE1
grapes
B
t
h
e
i
r
odo
concentra
10
times
breeders
commercia
grapes
t
c
o
m
e
r
c
i
a
foxy
f
l
a
v
x€
uLu&
w
I
n
a
present
d
e
a
c
t
i
v
a
t
p
o
l
a
r
i
t
y
and
100%
p
o
l
a
r
i
t
y
e
x
t
r
a
c
t
s
was
t
h
e
c
h
a
r
a
c
t
e
r
presence
reported
n
a
t
u
r
e
1
p
The
labrusca
anouhts
o
v
i
n
i
f
e
r
a
products
Concord
t
h
a
t
foun
chardonna
Bece
products
t
h
e
o
d
o
r
c
o
n
t
r
i
b
u
t
s
u
b
s
t
a
n
t
i
threshold
t
h
e
n
e
t
h
grapes
is
Besi
o
t
h
e
r
goo
such
con
r
e
n
i
n
i
s
c
c
COTTOA
Q!

Experimer
q
u
a
l
i
t
y
c
f
o
r
t
h
e
has
been
p
o
l
l
u
t
i
o
r
t
h
e
o
d
o
r
1.3­
cycl0
For
16
QUALITY
01;
SELECTED
FRUITS
AND
VEGETABLES
2:
ACRI
1:

40
20
100%

40
20
100%

80
60
40
20
100%

80
60
40
20
20
40
60
80
100
120
140
M
/e
i
I
20
40
80
100
120
140
M
/e
QUALITY
OF
SELECTED
FRUITS
AND
VEGETABLES
COMPOUND
ODOR
QUALITY
0
DAMASCENONC
FOXY
ANIMAL
.
FlORAl
FRUITY
COTTON
CANDY
STRAWBERRY
BURNT
SUGAR
".
"

2.

90
_­
Anhvestigation
of
the
Volatile
Flavor
Composition
of
V'tus
Labrusca
Grape
Musts
and
Wines.
1.
Methyl
Anthranilate
­
Its
Role
in
the
Total
Aroma
Picture
of
Labrusca
Varieties.*

Richard
H.
Tomlinson
and
Joe
boison*
I'

Contribution
from
Ckemistq
Dqartment,
McNaster
LTniversitzj,.
Canada
U
S
&Ill
and
the
Chemistry
Department,
University
of
Saskatchewan;
Saskatoon,
Canada
S7N
OW0
.
L'anthranilate
de
mifhyle
u
4tZ
considhi
comme
6tant
le.
compos4
volatile
le
pfas
&terminant
dans
l'ar6me
des
vins
et
des
moicts
de
misinq
uConcozrl"
et
f
u
t
longtemps
considirC
conrme
Ctant
l'agent
r&
Cla­
tmr
du
go5t
dissimzd&
associ4
ti
l'ardmedes
uins
'Concord"
et
des
autres
variltts
Vitis
labrusca.
Des
risultats
exphimentaux
notweaux
uiennent
demon­
trerqzte
des
compos&
colatiles
aromatiques
azttres
qtre
l'anthmnilate
de
d
t
h
y
l
e
peurent
dtre
des
agents
ri­
vdlateurs
del'ar6me
camcteristique
de
ces
variit4s
nard­
amh'caines.

Abstract
Methyi
anthranilate
has
been
proposed
as
the
single
most
important
volatileflavor
component
in
Concord
grape
musts
and
wines
and
has
long
been
held
respon­
sible
for
imparting
the
typical
"ffoxiness"
associated
with
the
flavor
of
Concord
and
other
Vitis
labrusca
Varieties.
New
experimental
evidence
is
presented
to
demonstrate
that
volatileflavor
compounds
other
than
methyl
anthranilate
m
y
be
respomible
for
imparting
the
typical
flavor
associated
with
these
native
North
American
varieties.

$
Presented
as
part
of
a
session
on
uAnalysis
of
Trace
Components
in
Alcoholic
Beverages",
Cas&
86
Xeeting,
Toronto,
Ontario,
Canada,
October
64,1986.
*
Author
to
whom
correspondence
m
y
be
directed.
e
n
t
address:
Agricuiture
Canada.
Food
ProductionandInspectionBranch,
Animal
Pathology
Laboratory,
116
Veterinary
Road,
University
of
Saskatchewan
Campus,
Saskatoon,
Saskatchewan,
­
Canada
C717
9Dr)
Methyl
anthranilate
has
been
considered
the
single,
­
"­

.most
important
volatile
flavor
component
in
Concord
grape
­musts
and
wines
since
its
presence
was
fwt
detected
in
Concord
grapes
by
Power
and
Chestnut
in
1921
(1).
Subsequently,
its
presence
in
several
Vitis
.

labmica
varieties
has
been
confirmed
by
Scott
(21,
Sale
and
Wson
(31,
Robinson
et
al.
'(
4­
7),
and
by.
HoIIey'
et
d.
(8).
This
compound
has
been
held
responsible
for
imparting
the
"foxiness"
associated
with
the
flavor
of
the
native
North
American
varieties
Witis
Zabntsca)
and
their
hybrids.
This
viewpoint
has
been
so
preva­
lent
in
the
minds
of
vintners
and
viticulturalists
in
the
eastern
United
States
and
Canada,
where
the
grapes
of
the
pative
vines
form
the
backbone
of
the
wine
industry,
that
viticultural
and
vinification
practices
in
this
region
have
hitherto
been
Focussed
on
the
reduc­
tion
or
total
removal
of
this
compound
from
the
prod­
ucts
they
make
for
consumers.
These
practices
have
unfortunately
not
yielded
the
anticipated
superior
­wines.
Could
it
be
that
vintners
and
viticulturalists
in
the
eastern,
United
States
and
Canada
have
geared
their
efforts
to
the
removal
of
a
cornpound
which
is
not
sole13
responsible
for
imparting
the
"distasteful"
characteristics
associated
with
these
varieties?
More
recent
investigations
into
the
volatiie
aroma
composition
of
the
native
North
American
grapes
and
wines
have
,paid
particular
attention
t
o
the
role
of
methyl
anbhfanilate
in
the
overall
aroma
of
these
vari­
.
'

­
eties
(9­
15).
iri?
lile
it
has
been
established
that
the
vola­
tile
aroma
eomposition
of
Concord
depends
signifi­
cantly
on
p
e
processing
technique
(141,
it
is
becoming
increasingly
$ear
that
the
flavor
character
is
ti^^
of
these
native
&iieties
cannot
be
explained
by
the
presence
Investigation
of
Volatile
Flmor
Composition
in
Wines
­7
or
absence
of
methyl
anthranilate
alone.
Some
native
I.
D.)
glass
column
packed
with
Superpak
20
M
(Anal­

.
.
varieties
such
as
Baco
Noir,
Elvira,
Delaware
(161,
abs,
Connecticut,
U.
S.
A.)
held
in
a
Varian
1800
Gas
Aurore
(16,18),
and
Catawba
(16,17,21)
show
no
.Chromatograph.
The
injection
port
temperature
was
detectable
concentrations
of
methyl
anthranilate
and
set
at
250%
and
the
oven
temperature
was
programmed
yet
possess
the
distinctive
Labrusca
flavor.
Various
­
"­.
from
20­
200°
C
at
G"
C/
min
and
held
at
the
final
temper­
authors
have
also
concluded
that
methyl
anthranilate
ature
for
10
min.
A
glass­
lined
microsample
splitter
done
cannot
be
the
most
significant
odorous
compo­
fitted
at
the
end
of
the
colUmn
provided
an
approxi­
nent
in
Vitk
labrusca
varieties
(12,14,1921):
Although
mately
1:
lO
split
ratio
of
the
chromatographic
effluent
previous
investigators
have
speculated
that
the
possi­
to
oneof
two
flame
ionization
detectors
(FID
set
at
ble
presence
of
one
or
more
compounds,
other
than
250°
C)
and
an
exit
port.
Th'e
exit
port
effluent
was
methyl
anthranilate,
may
rather
be
responsible
for
either
directed
to
a
fraction
collector
or
a
heated
recep
imparting
the
typical
hbrusca
flavor
to
these
vari­
tacle
(SNIFF
PORT)
where
the
nose
of
the
experi­
eties
(14),
there
is
no
evidence
in
the
literature
that
menter
could
evaluate
its
significance.
such
compourids
have
been
detected
and
identified.
This
paper
presents
new
experimental
evidence
to
clarify
the
role
of
methyl
anthranilate
in
the
total
aroma
picture
Packed
Column
GClMS
Analysis
of
the
native
North
American
varieties
in
a
continuing
research
effort
to
identify
the
compound(
s)
that
To
provide
qualititiie.
'identification
of
the
compo­
.
'

adequately
portray
the
flavor
characteristics
of
the
nents
separated
by
gas
chromatography.,
GC'separa­
.
Yitis
labrusca
varieties.
tion
was
conducted
on
an
identicalcolumnused
for
the
packed
column
GC
analysis
held
in
a
Varian
3700
Gas
p,:?
Experimental
focussing,
magnetic­
sector
mass
spedrometer
(>
IX70­
.
­.

.
~
..
..
Chromatograph
coupled
to
a
high­
resolution,
double­

Ld
70F,
VG
Analytical
Ltd.
Altrincham,
England)
Aiamtus
and
Materials
.
a
jet
separator.

r
.
A
newdesign
of
solvent
extractor
was
developed
for
this
analysis.
It
was
used
in
conjunction
with
a
new
design
of
concentrator
apparatus
to
successfully
isolate
and
enrich
the
volatile
flavor
fraction
from
95
mL
of
Cabernet
Sauvignon
wine
using
250
mL
of
purified
Freon­
11
(trickdomfluoromethane)
as
extracting
solvent
(E).,
Wine
samples
for
this
analysis
were
obtained
from
Chateau­
Gai
Wineries,
St.
Catharines,
Canada,
An&
Wineries,
Canada,
and
the
private
collection
of
one
of
the
authors
(RHT).

Isolation
and
Enrichment
of
the
Volatile
Flavor
Extract
Volatile
flavor
components
in
Pichon
Lalande
(Vini­
fm),
Vidal
(Labncscu),
Concord
(Lubrusca),
MouIin
Rouge
(Labrusca),
Delaware
(Labrusca),
Elvira
(L
a
b
c
a
),
and
Similkameen
Red
(Labmsca)
were
.
isolated
and
concentrated
2500­
fold,
using
the
previ­
ously
descriied
method
(22).
The
concentrated
extract
i
"'
was
stored
in
a
screw­
capped
vial
for
further
experi­

I'
mentation,
GC
andtor
GCNS
analysis.
GC
Experimekts
to
Evaluate
Regions
of
Organoleptic
Significance
(SNIFF
PORT)

Aliquots
(5­
10
pL)
of
the
concentrated
volatile
flavor
extract
were
injected
into
the
packed
column
GC
and
a
sensory
description
of
the
separated
and
eluting
components
was
obtained
(Figure
1).

Experiments
to
Evaluate
the
Amma
Contribution
of
the
Total
Extract
to
the
Wine
In
order
to
determine
if
the
nature
of
the
solutions
produced
by
mixing
the
flavor
extract
with
the
wine
base
depended
on
the
method
of
mixing,
a
total
of
20
pL
of
the
concentrated
volatile
flavor
extract
was
injected
into
the
GC
and
trapped
into
150
mL
of
cooled
wine
base
via
thelfraction
collector.
Samples
were
saved
for
sensory
evaluation
(Table
I).
Another
20
JLL
aliquot
of
the
same
flavor
extract
was
added
diiectly
to
a
second
150
mL
portion
of
cooled
wine
base.
The
resulting
mixtures
were
saved
for
sensofy
evaluation
(Table
11).
,.
I
'
I
GC
~qeriments
to
Reconstitute
Vitis
,.
,
.
Gas
chromatographic
separation
of
the
concentrated
The
..
failure
of
our
attempts
to
produce
homogeneous
..
I
'
,
.
PackedColumn
GC
Analysis
Labrusca
Wines
.
,
1,;
.
iz
'1
1
)I
;i
,
..

­
.
,
,­,
­
­
"

,,,:
r
;
­
­
4
­
:
.
­..
c­­
­A­
­­
a
+I...
..:"
"
.
­.

P
c
c
Richard
H.
Tomlinson
&
3w
Boisrm
2
.I
t­
i
I!
..
'

P
E
N
t
I
C
.
TIlocri=>

Figure
1.
Sensory
description
of
the
eluting
components
in
the
chromatographic
profile
of
a
concentrated
volatile
flavor
extract
of
Concord
wine
on
a
6
h.
X
0.4­
in.
O.
D.
(2
mm
LD.)
Superpak
21,
Jf
glass
column.
(Peaks
represetit
FID
recorder
responses
while
the
descriptors
represent
the
analyst's
perception
at
the
sniff
port
of
the
components
eluting
from
the
CC.)

bases
for
the
Labrusca
varieties
using
the
previously
described
isolation­
concentration
procedures
used
successfilIy
for
Pichon
Lalande
(22)
led
us
to
conduct
these
procedures
with
a
modified
version
of
the
solvent
extractor
and
concentrator
operated
under
carefully
controlled
inert­
atmospheric
conditions
(22).
The
isolated
and
enriched
volatile
flavor
extracts
obtained
under
these
new
operating
conditions
were
used
to
repeat
the
previously
described
experiments
.to
eval­
uate
the
aroma
contribution
of
the
total
extract
to
the
wine.
The
results
of
the
sensory
analysis
performed
on
the
resulting
homogeneous
solutions
are
show.
in
Table
111.

GC
Experiments
to
Evaluate
the
Contribution
of
Sections
of
the
Chromatographic
Profile
to
the
Wine
In
order
to
assess
the
aroma
contribution
of
various
fractions
(especially
methyl
anthranilate)
of
the
elution
profile
to
the
wine,
a
total
of
20
pL
of
the
concentrated
volatile
flavor
extract
obtained
under
the
modsed
conditions
was
injected
into
the
column
and
the
k
c
­
tions
eluting
from
the
Superpak
20
M
column
were
trapped
jnto
150
mL
of
cooled
\vine
bases
as
pictorially
"""
­
"_
L
­
1
:.­
T"
..
Figure
2.
Partial
Reconstructed
Ion
Chromatogram
of
a
Standard
Mixture
of
volatile
flavor
compounds
depicting
the
experiments
conducted
to
evaluate
the
flavor
contribu­
tion
of
sections
of
the
chromatographic
profile
on
the
wine.
X.
Ethyl
hexanoate
2.
Amyl
butyrate
3.
Ethyl
Iactate
4.
cis­
3­
hexen­
1st
5.
.Ethyl
octanoate
6.
Zmethylbutyi
hexanoate
7.
Benzaidehyde.
8.
I­
Octanol9.
y­
Butyrolac­
tone
10.
Ethyl
benzoate
11.
Ethyl
decanoate
12.
Diethyl
succinate
13.2­
Phenethyl
acetate
1.1.
Hexanoic
acid
15.
2­
Phenethyl
alcohol
16.
t­
cinnamaidehyde
17.
Octanoic
acid
IS.
Methyl
anthranilate
19.
Decanoic
acid
20.
Dh+
Llli,
l­
O?
0
7%"
­­
I..­*
1
Investigation
of
Volalile
Flavor
Composition
in
Wina
Table
1.
Experiments
t
o
Evaluate
the
Aroma
Contribution
of
the
Total
.Extract
to
the
Wine.

20
pL
of
Flavor
Extract
(a)
150
mL,
Wme
base*
(b)­

Concord
Xoulin
Rouge
Vidal
Elvira
Delaware
S
i
e
e
n
RH
Pichon
Lalande
Observation
aftera
+
b
Concord
­

f
u
r
f
a
c
e
Film
Forme
Concord
Moulin
Rouge
Vidal
Elvira
Delaware
Moulin
Rouge
..
.
..
S
i
e
e
n
Rei
Pichon
Lalande
I
~
~~

Concord
Xoulin
Rouge
Vidal
Elvim
Delaware
Pichon
L'aIapd.
Sirnilkameen
Rec
Von­
Homogeneous
m
.
"med
.
Table
111.
GC
E­
xperiments
to
Reconstitute
\'itis
Labnuar
Wines
using
the
lfodiied
Isolation­
Concentration
Method.
"
Vidai
Strong
Burned
Odor
leveloped
Concord
Jfoufin
Rouge
Vidd
Elvira
Delaware
S
i
e
e
n
Red
Pichon
Lalande
Elvira
Delaware
Concord
Xoulin
Rouge
Vidal
Elvira
Delaware
S
i
e
e
n
Red
Pichon
Lalande
Table
IV.
GC
Experiments
to
Evaluate
the
Flavor
Contribution
of
Sections
of
the
Chromatographic
Profile
to
the
Wine.

Concord
Moulin
Rouge
Vi&
Elvira
Similkameen
Red
f­
Delaware
Sirnilkameen
Red
Pichon
Lalande
Flavor
of
PI
Regenerated
Homogeneous
Soin.
Formed
No
Detectable
hbrusca
Flavor
Perceived
?oncord
Houlin
Rouge
Vidal
Zlvira
Pichon
Lalande
leiaware
S
i
e
e
n
Red
?ichon
Lalande
Wine
base
is
the
bland
product
,remaining
after
the
volatile
flavor
La:"
L­
­
*
L
94
Richard
H.
Tonrlinson
&
Joe
Boison
(a)
all
components
eluting
up
to
and
including
2­
sitions
of
Vitis
l
a
h
s
c
a
varieties
have
been
investi­
phenethyl
alcohol
were
trapped
into
wine
base
A;
gated
using
techniques
essentially
developed
for
Vi?&
(b)
all
components
eluting
up
to
but
excluding
methyl
fm
varieties
without
any
major
modifications.
After
anthranilate
were
trapped
into
wine
base
B;
,the
apparatus
developed
for
this
analysis
was
modified
(c)
all
components
eluting
from
2­
phenethyl
alcohol
"
to
enable
the
isolation­
concentration
procedures
to
be
(inclusive)
up
to
but
excluding
methyl
anthranilate
were
trapped
into
wine
base
C;
(d)
all
components
eluting
from
methyl
anthranilate
(inclusive)
and
thereafter
were
trapped
into
wine
base
D;
and
(e)
all
components
eluting
except
Freon
solvent
were
trapped
into
wine
6ase
E
to
serve
as
control.
The
results
of
the
sensory
evaluation
of
the
solutions
obtained
in
this
experiment
are
presented
in
Table
IV.
Sensory
evaluation
of
samples
was
done
by
a
group
of
faculty,
staff
and
graduate
students
of
the
Chemistry
Department,
3lcMaster
University,
who
had
been
trained
to
recognize
the
Labnlsca
flavor.

Results
and
Discussion
The
results
of
the
GC
experiments
presented
in
Table
I
show
that
in
all
cases
where
the
voIatile
flavor
extract
was
of
the
Labnwca
origin,
the
introduction
of
the
flavor
extract
into
the
wine
base
through
the
GC
frac­
tion
collectorproducednon­
homogeneous
solutions.
Note
that
when
the
volatile
flavor
extract
was
of
the
Vinifera
origin,
homogeneous
solutions
were
obtained
regardless
of
the
origin
of
the
wine
bases.
The
results
shown
in
Table
I1
were
obtained
by
introducing
the
flavor
extracts
directly
into
the
wine
bases
thus
elim­
inating
the
GC
step
for
sample
introduction
used
for
obtaining
the
results
presented
in
Table
1.
Since
both
methods
of
sample
introduction
info
the
wine
bases
produced
non­
homogeneous
solutions,
it
was
concluded
that
the
results
obtained
in
Tables
I
and
I1
were
not
caused
'by
the
method
of
sample
introduction,
but
may
have
been
caused
by
processes
occurring
prior
to
the
sample.
introduction
step,
i.
e.,
the
isolation­
concentration
procedures.
Any
changes
caused
by
the
isolation­
concentration
procedures
wouid
affect
either
the
flavor
extract
orr
the
wine
base
or
both.
In
an
attempt
to
identify
the
source
zpd
cause
of
the
change
in
the
Labmsca
varieties,
it
was
observed
that
the
flavor
extract
of
Pichon
Lalange
and
White
Riesling
wines,
'both
made
from
Vingma
grapes,
dissolved
readily
into
their
o
m
wine
bases
and
all
the'
wine
bases
of
Labmsca
origin
to
regenerate
the
characteristic
Cabernet
Sauvignon
or
Riesling
aroma.
This
obser­
vation
led
us
to
conclude
that
it
is
the
flavor
eitract
of
,the
4bbrusca
rather
than
the
wine
basg
that
may
have
'hnilergobe
change
during
the
sample
prepaiation
stages.
Ext,
ensive
review
of
the'liteqture
dnd
our
own
"*
l"
A­
l"
L­
"_..
­1
*
.I
L
3
­
.
.­
­
conducted
under
carefully
controlled
inert
atmospheric
conditions
(22),
it
became
possible
to
produce
homo­
geneous
solutions
and
therefore
reconstitute
all
the
Vitis
lafrrusca
wines
as
shown
in
Table
111.
Any
further
sample
extraction
and
concentration
procedures
relat­
ing
to
the
LQbncsca
varieties
were
therefore
performed
under
the
modified
operating
conditions.
Having
developed
a
method
that
retains
the
original
nature
of
the
volatile
flayor
extract
to
permit
the
reconstitution
of
Lahcsea
\vines,
it
was
observed.
from
the
results
of
the
experiments.
conducted
to
assess
the
aroma
contribution
of
the
eluting
components
to
the
wine
that,
besides'the
control
sample
E,
only
fractions
B
and
C
trapped
into
the
wine
bases
produced
solutions
that
typsed
the
Lubncsca
character;
fraction
D
(methyl
anthranilate)
produced
a
solution
that
could
hardly
be
described
as
typically
Lubncsca.
It
is
noteworthy
to
observe
that
these
results
and
conclusions
were
the
same
for
all
thetabntscu
varietiesinvestigated
includ­
ing,
E
l
v
h
andDelaware,
which
even
showedno
detectableconcentrations
of
methyl
anthranilate.
Since
,

fraction
€3
also
encompasses
fraction
C,
which
by
itself
imparts
.the
Labrusca
flavor
to
the
wine,
it
may
be
concluded
that
fraction
C
must
be
the
most
organo­
leptically
significant
portion
of
the
profile
of
the
vola­
tile
flavor
composition
of
Vitis
labnlsca
varieties
on
the
Superpak
20
M
column.
This
conclusion
is
rein­
forced
by
the
results
of
the
preliminary
experiments
to
evaluate
regions
of
organoleptic
significance
in
the
chromatographicprofile
(Figure
1).
The
results
in
Figure
1
deiine
even
more
precisely
the
region
of
Organoleptic
interest
in
the
chromatographic
profile
by
confining
it
to
the
labelled
section
which
is
comprised
of
compounds
with
similar
retention
times
to
octanoic
acid.
Any
experiments
designed
to
detect
and
identify
the
compound(
s)
responsible
for
imparting
the
typical
Labrusca
flavor
to
the
native
North
American
vari­
eties
should
therefore
be
confined
to
the
section
of
the
chromatogram
labelled
'kegion
of
organoleptic
inter­
est"
(Figure
1).

Conclusions
The
results
of
these
experiments
confii
the
suspi­
cions
of
previous
investigators
that
the
presence
or
absence
of
methyl
anthranilate
alone
cannot
explain:
the
typicalflavorassociated
with
the
native
North
.
'
­
L
Investigation
of
Volatile
Flavor
Composition
in
Wines
tant
odorous
compound
in
these
varieties.
Unlike
Vitis
vinifera
varieties,
isolation
and
concentration
proce­
dures
for
the
examination
of
the
volatile
flavor
compo­
sition
of
Vitis
labnrsca
grape­
musts
and
wines
must
be
conducted
under.
carefully
controlled
inert
atmos­
pheric
conditions
t
o
prevent
any
possiblechemical/
structural
modifications
to
the
nature
of
the
volatile
flavor
extract.
The
experimental
evidence­
presented
in
this
paper
demonstrates
that
compounds
other
than
methyl
anthranilate
are
responsible
for
imparting
the
typical
Labmca
flavor.
Such
compound(
s)
are
those
with
similar
retention
times
to
that
of
octanoic
acid
on
the
Superpak
20
M
column.
This
conclusion
should
be
of
considerable
significance
to
vinters
and
viticultuy­
alists
in
the
eastern
United
States
and
Canada
in
help
ing
them
to
review
not
only
their
vinification
and
viti­
cultural
practices
but
also
their
perception
of
the
role
­
played
by
methyl
anthranilate
in
the
total
aroma
picture
of
the
Labntsca
varieties.
"

...
".
­.
::

ps;
References
L
,l
I
.

1.
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B.
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r
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d
\

'"
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H.
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B.
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w
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e
r
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D.
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'
E.
Kepner,
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8
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R.
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E.
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i.
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0.
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Canada,
1984.

­
A
Cornparison
of
the
Effect
of
Toxic
and
Non­
Toxic
Metals
on
the
Aromatic
System
of
Salicylic
Acid
Wiodzimierz
Lewandowski
Contribution
from
Department
of
Physico­
Chemical
Analyses,
Agricultuml
University
.of
Warsaw,
03­
528
Warsaw,
Rakoutiecka
26/
30?
Poland
Received:
November
10,
1986
Accepted
(inrevised
form):
May
29,
1987
R6sumC
Les
spectres
d'abso?
ption
IR
et
Raman
des
salicy­
lates
de
mercure(
II),
cadmium
et
aluminium
ont
dtd
6tudi6s
et
interpritks.
On
acompark
l%=
fluence
de
mereure(
1)
et
"(
II),
pLomb(
II),
cadmium,
mgne­
sium,
zinc,
aluminium,
fer(
III),
lanthane(
II0,
neodyme(
III),
disprosium(
III),
ytterbiumllll),
.sodium
et
potassium
sur
le
noyau
aromatiQue
de
l'aeide
salicilique.
Pour
les
investigations
on
a
applipud
de
la
spectroscopie
d'absorption
Llectronique
(UV,
VIS)
et
la
spectroscopie
d'oscillation
(celle
$absorption
a
l'infrarouge
et
celle
de
Raman).
On
a
constat6
que
tes
,
.­
.
..
.
.
­
,­
;.
.
­
.
.
.
.
­
contraire
de
aluminium,
lunthadIII),
n&
iynte(
III),
disprosium(
III),
ytterbium(
III),
fer(
III),
zinc
et
magwsiz~
ml
perturbent
le
systi?
meammatique
de
l'acide
salicilique.

Abstract
'
Assignments
areproposed
for
the
IR
absorptionand
'
Raman
spectra
of
mercury(
II),
cadmium
and
aluminium
salicylates.
Th
effect
of
mercury(
I)
and
4
1
1
,
l
e
d
l
l
),
cadmiuwi
as
weld
as
of
magnesium,
zinc.
aluminium,
iron(
III),
lanthanum(
III),
.,
f
.
..
..

&.
r
1188
J.
A&.
Food
Chem.,
Vd.
26,
No.
5,.
1978
Jeon,
I.
J.,
Reineccius,
G.
A,
Thomas,
E
L.,
J
.
Agric.
Food
Chem,
24.433
(1976).
Kinsella,
J.
E.,
Chem.
fnd.,
36
(1969).
.
Kirk,
J.
R.,
Hedrick,
T.
I.,
She,
C.
M.,
J.
Dairy
Sci.
51,492
(1968);
Langler,
J.
E.,
Day,
E.
A.,
J.
Dairy
Sci.
47,1291
(1964).
;
Ldlard,
D.
A.,
Day,
E.
A.,
J.
Dairy
Sci.
44,
623
(1961).
Muck,
G.
A,,
Tobias,
J.,
U'hitney,
R.
M.,
J.
Dairy
Sci.
46,
774:
(1963).

Whitney,
R.
M.
J
.
Dairy
Sci.
46,
671
(1963).

6,232
(1964).

(1963).
Nawar,
U'.
W.,
Lombard,
S
.
H.,
Dall,
H.
E.
T.,
Ganguly,
A.
S.,

Parks,
0.
W.,
Keeney,
M.,
Katz,
I.,
Schwartz,
D.
P.,
J.
Lipid
Res.

Parks,
0.
W.,
Keeney,
M.,
Schwartz,
D.
P.,
J.
Dairy
Sci.
46,295
Parks,
0.
W.,
Patton,
S.,
J.
Dairy
Sci.
44,
1
(1961).
Parks,
0.
W.,
Schwartz,
D.
P.,
Keeney,
M.,
Nature
(London)
202,
185
(1964).
patel,
T.
D.,
Calbert,
H.
E.,
Morgan,
D.
G.,
Strong,
E'.
M.,
J.
DLrY
Sci.
45,601
(1962).
**"*
rrw­
*­

Patton,
S.,
J.
Dairy
Sci.
36,
1053
(1952).
Patton,
S.,
Josephson,
V.,
Food
Res.
22,316
(1957).
scan
la^,
R
A,
U
y
,
R
C.,
Libbey.
L.
M.,
%,
E
A,
J.
Dairy
schwartz,
D.
P.,
Parks,
0.
W.,
Y
o
n
d
e
,
R
A,
J.
Am.
Oil
Cbm.
,
i
Stark,
W.,
Forss.
D.
A,
J.
Dairy
Res.
33,31
(1966).
.1
Thomas.
E.
L.,
Burton,
H.,
Ford,
J.
E.,
Perkin,
A.
C.,
J.
Bairn
;
Si.
51,
1001
(1968).

Soc.
43,128
(1966).
1
Res.
42,285
(1975).
­1
Toothill,
J.,
Thompson,
S.
Y.,
Edwards­
Webb,
J..
J.
Dairy
R
~~.
1
4
37,29
(1970).
7
.
withers,
M.
K.,
J.
Chromatogr.
66,249
(1972).
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J.
G.,
BimRistle,
R.,
J.
Dairy
Res.
40,169
(1973).

Isolation
and
Identification
of
Volatiles
from
Catawba
Wine
Richard
R.
Nelson,
Terry
E.
Acree,*
and
Robert
M.
Butts
.
­

The
volatile
composition
of
three
Catawba
wines
prepared
from
grapes
grown
in
the
vineyards
of
the
New
York
State
Agricultural
Experiment
Station
during
the
1976
vintage
were
analyzed
by
instrumental
and
sensory
means.
The
three
wines
differed
according
to
the
enological
technique
employed
for
their
production.
Volatiles
were
isolated
by
solvent
extraction,
separated
and
quantified
by
gas
chroma­
tography,
arid
identified
by
combined
gas
chromatography­
mass
spectrometry.
Although
some
variation
in
volatile
composition
due
to
processing
technique
was
observed,
sensory
analyses
comparing
the
win?
with
corresponding
model
solutions
indicate
that
the
major
identifiable
components
are
of
little
im­
portance
in
determining
the
aroma
of
Catawba
wine
as
influenced
by
processing
technique.
4
Catawba
vines
have
been
cultivated
in
the
northeastern
United
States
for
over
150
years.
Currently,
in
New
York,
over
10
000tons
are
produced
annually
and
over
90%
of
that
is
used
for
wine
production
(New
York
State
Crop
Reporting
Service,
1976).
Catawba
grapes
can
be
used
in
the
production
of
either
white
or
ros6
wines
depending
upon
enological
technique,
and
much
of
the
white
wine
produced
is
used
in
sparkling
wine
cuv6es.
The
literature
dealing
with
the
volatile
composition
of
wines
and
winegrapesisextensive.
Kahn
(1969)
and
Webb
and
Muller
(1972)
have
tabulated
hundreds
of
compounds
that
have
been
identified
in
wines
and
other
'alcoholic
beverages.
Many
native
American
grape
varieties
including
Concord
and
Catawba
have
characteristic
aroma
components
that
appear
to
be
unique
to
some
varieties
with
iabrusca
parentage.
Although
many
workers
(Holley
et
al.,
1955;
Neudoerffer
et
al.,
1965;
Stevens
et
J.,
1965;
and
Stern
et
al.,
1967)
have
studied
the
volatile
composition
of
the
Concord
variety,
no
such
investigations
have
been
con­
ducted
with
Catawba.
Methyl
anthranilate,
a
compound
long
thought
to
be
of
major
importance
in
the
aroma
of
labrum
varieties
(Sale
and
Wilson,
19261,
now
appears
to
be
far
less
important
than
previously
thought
(Nelson
et
d.,
1977a).
Friedman
(1976)
believes
that
methyl
anthranilate
is
of
little
im­

Department
of
Food
Science
and
Technologyv
New
Yoyk
state
A&
ultural
Experiment
Station,
Geneva,
New
Yo&
14456.
­
_­­
"­l
.".1
."
.."".
n*,
fi
portance
in
the
aroma
of
Concord
grapes,
even
though
ita
concentration
in
that
variety
&­
relatively
high.
h
e
h
e
et
aL
(1959)
noted
that
the
distinctive
Catawba
aroma
was
apparently
not
due
to
methyl
antbrdte
and
that
other
more
important
compounds
must
be
present.
This
report
examines
the
volatile
composition
of
Ca­
tawbawines
prepared
by
three
different
enological
techniques.
In
addition,
it
attempts
to
assess
the
effect
of
processing
technique
on
the
occurrence
of
these
volatils
in
Catawba­
wine.
MATERIALS
AND
METHODS
Wine
Preparation.
Catawba
grape6
were
harvested
at
16.6"
Brix
in
October
of
1976.
The
fruit
was
divided
into
three
20­
kg
lots
for
fermentation.
From
one
lot
a
white
Catawba
wine
was
prepaied
by
immediately
pressing
the
7
crushed
grapes
while
rose
wines
were
prepared
from
the
other
two.
One
of
the
rose
wines
was
prepared.
by
fer.
meriting
the
juice
in
contact
with
the
ekins
for
5
days
[d
(FS)]
whil'e
the
other
was
thermally
vin%
ed
[rose
(m71.
"hemal
Mlcation
comista
of
hating
the
crushed
grapes
in
a
steam
kettle
to
60
O
C
for
16
min,
followed
by
im­
mediate
pressing.
Fermentations
were
conducted
at
20
*C
and
other
standard
enological
procedures
as
described
b'
Nelson
et
al.
(1977b)
were
followed
in
each
case.
Volatile
Isolation.
The
Catawba
wine
volatile6
were
isolated
using
organic
solvent
extraction
with
Freon
113
(1~
1P­
tricbloro­
1,2,2uor~~~
e,
"Precision
Cleaning
Agent,
Du
Pont").
33qd
volumes
of
wine
and
fieon
(2700
&I
were
stirred
fdr
1
h.
The
Freon
phase
was
then
draws
bff,
dried
over
anhydrous
magne+
m
sulfate,
and
Con­
*
retention
concentration,
ppm
time,
rose
rosd
compound
min
white
(TV)
(FS)
"
ethyl
acetate
p
)
jsobutyl
acetate
(c)
ethyl
butyrate
(d)
isoamyl
acetate
(e)
isoamyl
alcohol
(f)
ethyl
hexanoate
(g)
hexyl
acetate
(h)
ethyl
lactate
ii
j
n­
hexanol
(j)
cjs­
3­
hexene­
1­
01
(k)
ethyl
actanoate
(1)
~inaloiil
(m)
butyric
acid
(n)
ybutyrolactone
'
(0)
ethyl
Gecanoate
(p)
diethyl
succinate
(9)
2­
phenylethyl
acetate
(r)
hexanoic
acid
(6)
2­
phenylethyl
alcohol
(t)
octanoic
acid
(u)
methyl
anthranilate
3.25
6.22
6.74
9.14
11.37
12.84
14.07
16.18
16.43
17.41
19.61
23.18
24.13
25.02
26.07
27.39
31.70
32.76
33.17
38.87
46.41
0.15
0.16
0.02
0.03
0.04
0.03
i
0.17
0.12
0.09
2.57
3.19
1.68
6.27
3.19
1.68
0.57
0.49
0.47
0.04
0.13
0.20
0.17
0.24
0.45
0.34
0.12
0.76
0.06
0.03
0.13
1.16
0.74
0.55
0.02
0.05
0.10
0.04
'I"
'
0.12
ndb
Tr
0.06
0.53
0.33
0.26
0.30
0.56
0.95
1.09
3.79
2.90
3.23
­1.35
5.51
5.00
7.45
7.50
9.20
7.35
1.80
ndb
0.07
Tf
'a
only
trace
quantity
detected.
Not
detected.

centrated
in
a'rotary
evaporator
with
water
bathat
20
"C.
The
final
extract
concentration
was
13500­
fold
(0.2
mL),
md
each
extract
had
a
characteristic
Catawba­
like
aroma
while
the
aqueous
phase
was
nearly
odorless.
Instrumental
Analysis.
The
Catawba
extracts
were
analyzed
by
combined
gas
chromatography­
mass
spec­
trometry
using
3­
pL
injections.
The
system
consisted
of
a
Sarian
Series
1400
gas
chromatograph
with
a
4
m
X
2
mm
i.
d.
glass
column
packed
with
10%
SP­
lo00
on
100­
120
mesh
Chromosorb
W.
A
temperature
program
from
60
to
200
"C
at
4
"C/
min
was
employed.
The
gas
chromatograph
was
interfaced
through
a
LleweUyn
type
methyl
silicone
membrane
separator
to
a
Bendix
Model
12
Time­
of­
Flight
mass
spectrometer
equipped
with
CVC
Mark
IV
solid
state
electronics
and
with
a
computerized
data
collection
system.
Spectra
were
taken
at
70
eV
and
identification
was
done
by
comparison
of
experimental
spectra
with
published
spectra
and
with
those
of
authentic
standards.
Comparison
of
component
retention
times
with
those
of
the
authentic
standards
was
considered
to
be
confirmatory.
Quantification
of
volatile
components
was
done
using
a
Hewlett­
Packard
5830
A
gas
chromatograph
Chromatographic
conditions
were
the
same
as
above
exckpt
that
a
stainless
steel
column
of
the
same
dimensiow
was
used.
Quantitative
estimation
was
done
by
comparing
;the
peak
area
of
2­
phenylethyl
alcohol,
a
major
component,
with
that
of
an
internd
standard
(n­
decanol)
added
to
the
Freon
extract
at
a
level
of
2
ppm
relative
to
the
original
wine.
The
concentrations
of
the
other
components
were
then
calculated
directly
using
relative
peak
area
ratios.
Sensory
Evaluation.
The
odors
of
the
individpal
components
in
the
three
extracts
were
characterized
by
the
authors
using
a
sniffing
device
(Acree'etm
a].,
1976)
attached
to
the
effluent
port
of
a
Packard
Model
800
gas
chromatograph.
Chromatographic
conditions
were
the
same
as
in
the
GC­
MS
system.
In
order
tb
examine
the
role,
if
any,
of
the
identified
compounds
in
determining
Catawba
odor,
model
wine
solutions
were
prepared
using
12%
v/
v
ethanol­
1%
w/
w
tartaric
acid,
pu;
ified
Concord
grape
anthodyanin
gigmeht,
and
the
identified
volatiles
in
distilled
water.
The
v&
tiles
Were
added
to
the
three
model
solutions
at
the
level
de­
termined
for
each
compound
iq
the
corresponding
au­
t
i
..

I
30
40
50
RETENTION
nME
(MIN)

Figure
1.
Chromatogram
of
the
13500­
fold
Freon
extract
of
Catawba
wine
fermented
in
contact
with
the
skins.

tbentic
wine.
The
concentration
of
each
compound
added
to
model
solutions
we
listed
in
Table
I.
All
compounds
identified
were
added
to
the
corresponding
model
solution,
except
when
only
a
trace
was
detected
none
was
added.
To
eliminate
panel
bias
due
to
color
differences,
pigment
was
added
to
the
wins
and
to
the
model
solutions
at
levels
such
that
each
sample
had
a
t­
ypical
ros&
color.
An
ex­
perienced
1Zmember
panel
completed
three
sets
of
randomized
triangle.
difference
tests.
Only
odor
was
Considered
by
the
panel
90
the
samples
were
not
tasted.
The
Catawba
wines
were
first
tested
against
each
other,
and
then
the
model
solutions
were
examined
for
aroma
,
differe?
ces.
Finally
each
winewas
tested
against
its
corresponding
model
solution.
Samples
were
presented
at
room
temperature
in
standard
wine
glasses
in
individual
tasting
booths.
Sample
size
was
50
mL.
RESULTS
A
N
D
DISCUSSION
In
the
white
Catawba
wine,
36
compounds
were
present
in
concentrations
sufficient
for
quantitative
estimation.
In
the
rose
wine
fermented
in
contact
with
the
skins
and
in
the
thermally
vinified
ros6
wine
42
and
50
compounds
were
detected,
respectively.
Of
these,
19
compounds
found
­

in
the
white
wine
were
identified
and
21
compounds
in
the
other
two
extracts
were
identified.
Methyl
anthranilate
and
y­
butyrolactpne
were
not
detected
in
the
white
Ca­
tawba
wine.
The
compounds
that
were
identified
were
generally
those
present
in
the
largest
quantities.
The
ide'ntified'compounds
had
a
total
concentration
that
was
ne+
y
constant
at
30.8
ppm
in
all
three
wines
and
rep­
res,
ented'
from
95.7
to
97.5%
of
the
total
extractable
volatiles;
F
i
1
shows
a
chromatogram
of
the
13
5oO­
fold
extract
of
the
wine
fermented
in
contact
with
the
skins.
The
letters
"a"
through
"v"
correspond
to
the
identified
cornpourids
listed
in
Table
I
with
their
concentrations
in
the
differmt
wines.
The
concentrations
of
the
maj0rit.
y
of
$he'compunds
appear
to
be
little
affected
by
enologid.
technique.
However,
the
acetate
esters,
particularly
isohmy1
acetate
tpd
2­
phenylethyl
acetate.
are
distinctly
more
+bundant
in
'the
thermally
vinified
wine.
Decanoic
acid
(v)
was
identified
in
each
of
the
wine
extiacts
but
its
concentrktion
could
not
be
reliably
estimated
due
to
excessivk
chromatographic
tailing.
I!@
ii$
J~
skrongly
oro
ow
compounds
were
detected
by
gas
chrpdaxikaphip
,effluent
sniffing.
Some,
with
extremely
lo?
&&
Fent
thresholds,
were
present
in
concentrations,
,,
nble
E.
Results
of
Randomized
Triangle
Teb
Comparing
the
Aroma
of
Catawba
Winea
and
of
Model
Solutions
correct
comparison
response%
significance
wines
vs.
wines
­white
vs.
rosh
(TVIb
10
0.001
white
vs.
r
o
d
(E'S)=
9
0.01
res;
(TV)
vs.
rosd
(FS)
9
0.01
white
vs.
r&
(TV)
9
0.01
white
vs.
ros6
(FS)
7
NSd
ro&
(TV)
vs.
rose
(FS)
9
0.01
white
11
0.001
rosi
(TV)
1
2
0.001
ros6
(FS)
10
0.001
Maximum
nurpber
of
correct
responses
is
12.
Ther­
mally
vinified
rose
wine.
Rosd
.wine
fermented
in
con­
tact
with
the
skins.
Not
significant.

too
low
to
give
any
detector
response.
No
single
compound
was
thought
to
have
a
distinctly
Catawba­
like
aroma..
The
results
of
the
triangle
difference
tests
are
shown
in
Table
11.
The
panel
found
that
the
aroma
of
each
Ca­
tawba
wine
was
significantly
different
from
that
of
the
other
twowines
(99%
level'of
confidenceor
better).
Clearly,
if
processing
technique
had
no
significant
effect
on
Catawba
wine
aroma,
further
sensory
investigation
would
be
unproductive
but
this
is
apparently
not
the
case.
The
panel
was
then
asked
to
distinguish
among
the
three
model
solutions.
The
aroma
of
the
solution
imitating
the
thermally
vinified
wine
was
significantly
different
from
that
of
the
other
two
solutions
(99%
level
of
confidence).
­.

Apparently
some
compositional
difference
in
the
identified
compounds
is
organoleptically
significant
and
peculiar
to
the
thermally
vinified
wine.
Although
proof
is
not
yet
available,
the
mostobviouscompounds
to
which
this
difference
can
be
attributed
are
isoamyl
acetate
and
2­
phenyktiiyl
acetate
because
of
their
high
concentration
in
that
sample.
The
solutions
corresponding
to
the
wine
fermented
in
contact
with
the
skins
and
the
white
wine
were
indistinguishable
by
the
panel.
This
indicates
that
the
identified
compounds,
even
though
they
represent
over
95%
of
the
total
extractable
volatiles,
do
not
account
for
the
significant
aroma
differences
that
occur
in
Catawba
wine
due
b
processing
technique.
In
the
final
triangle
test
each
wine
was
judged
against
its
corresponding,
model,
solution.
A
highly
significant
difference
(99.9%)
was
found
in
each
case.
The
three
model
solutions
were,
in
fact,
very
poor
imitations
of
the
authentic
wine.
The
high
concentratiofi
of
acetate
esters
in
the
thermally
vinified
wine
may
contribute
to
its
dis­.
tinctive
aroma
but
it
certainly
does
not
define
that
aroma.
models
vs.
models
wines
vs.
models
.
.""
.,
"
q
*­

It
appears
that
the
identified
components,
although
they
do
contribute
aroma,
contribute
little­
or
nothii
to
Ca.
tawba
varietal
character
as
influenced
by
enologicd
technique.
The
nature
of
wine
and
wine­
grape
aroma
is
not
we0
understood.
Brander
(1974)
has
suggested
that
e
n
m
y
the
same
volatile
Components
are
present
in
all
wine
varieties
and
that
the
aroma
differences
­0ng.
Virietia
are
due
to
these
components
beiig
present
in
differing
ratios.
On
the
other
hand,
Stern
(1975)
stresses
the
h­
portance
to
wine
aroma
of
compounds
present
in
trace
quantities.
The
concentrations
of
these
compoun&
is
often
too
low
to
give
any
detector
response
whataoever
cment
methodology,
but
they
may
be
of
great
0rganoIeptii
importance
if
they
have
sufficiently
low
thresholds.
The
major
volatile
components
detected
and
identified
in
the
three
Catawba
wines
do
contribute
aroma
to
those
wines.
However,
the
model
solutions
containing
these
compounds
at
their
appropriate
concentrations
are
easily
disthguished
from
the
authentic
wine.
It
can
be
condud&
that
the
unidentified
trace
components,
although
they
comprise
less
than
5%
of
the
total
Freon­
extractable
volatiles,
are
of
critical
importance
to
the
aroma
of
Ca­
tawba
wine
as
influenced
by
processing
technique.
LlTERATURE
CITED
Acree,
T.
E,
Butts,
R.
M.,
Nelson,
R.
R.,
Lee,
C.
Y.,
Am!.
Chem.
48(
12),
1821
(1976).
Amerine,
M.
A,,
Roessler,
E.
B.,
Filipello,
F.,.
Hifgardia
28(
1&),
i
500­
501
(1959).
Brander,
C.
F.,
Am.
J
.
Enol.
Vitic.
W(
l),
13­
18
(1974).
I
Friedman,
I.
E.,
N.
Y.
Hortic.
SOC.
Roc.
121,
132­
136(
1976).
1
Holley.
R.
W.,
Stoyla,
B.,
Holley,
A.
D.,
Food
Res.
20.326330
I
(1955).
"
"_

Kahn,
J.
H.,
J.
Assoc.
Off.
Anal.
Chem
52(
6),
1166­
1123
(1%
9).
Nelson,
R.
R.,
A~
ree,
T­
E.,
Lee,
C.
Y.,
Butts,
R.
M.,
J.
Food
Sci.

Nelson,
R
R,
A
m
,
T.
E.,
Robinson,
W.
B.,
Pool,
R
M.,
Bertino,

Neudoerffer,
T.
F.,
Sbdler,
S.,
Zubeckis,
E.,
Smith,
M.
D.,
J
.

New
York
Crop
Reporting
Service,
Survey
of
Wineries
and
Grape
.
Sale,
J.
W.,
Wilson,
J.
B.,
J.
Agric.
Res.
33(
4),
301­
310
(19261.
Stern,
D.
J.,
Guadagni,
D.,
Stevens,
K.
L,
Am.
J.%
nol.
Vitic.
26(
4).

Stern,
D.
J.,
Lee,
A.,
McFadden,
R
.
H.,
Stevens,
K.
L.,
J.
Agric.

Stevens,
K.
L,
McFadden,
W.
H.,
Teranishi,
R.,
J.
Food
Sci.
30,

Webb,
A.
D.,
Muller,
C.,
Ado.
Appl.
Micro&
ioZ.
15,75­
146
(1972).
42,57­
59
(1977a).

J.
J.,.
N.
Y.
Food
Life
Sci.
Bull.
No.
66
(July
1977b).

Agric.
Food
Chem.
13(
6),
584­
588
(1965).

Processing
Plants,
Albany,
N.
Y.,
1976.

208­
213
(1975).

Food
Chem.
15(
6),
1100­
11,03
(1967).

1006­
1007
(19651.

Received
for
review
October
25,1977.
Accepted
May
4,1978.
Presented
at
the
174th.
National
Meetingof
the
American
ChemicalSociety,
Chicago,
Ill.,
Aug1977.
Approved
by
the
Director
of
the
New
York
State
Agricultural
Experiment
Station
as
Journal
Paper
No.
3107.
..
f
­
.~

R.
R.
NELSON.
1.
E.
ACREE,
C
Y.
LEE
and
R.
M.
BUTTS
Dept.
of
Foodscience
&
Technology
New
York
Sfate
Agricultural
Experiment
Station,
Geneva,
NY
14456
METHYL;
ANTHRANILATE
AS
AN
AROMA
CONSTITUENT
OF
AMERICAN
WINE
i
INTRODUCTION
lf[
f`
WINE
INDUSTRY
of
the
Eastern
United
States
and
Can­
s
based
on
the
native
American
grapes
and
on
hybrids
and
rlr,­;
ions
thereof.
Certain
native
grapes
have
a
characteristic
r!,,,
nF
flavor
which
is
disagreeable
t
o
many
wine
consumers.
1
rwrilc
over
100
yr
of
intensive
grape
breeding,
this
"foxy"
,.&
c??
s,
t,
r
still
prohibits
production
of
European
style
wines
from
\\
h.
7
The
origin
of
the
term
"foxy"
and
its
association
with
the
grapes
I:
.
._­
­
~

­
Amric­
n
varieties
is
unclear,
but
the
most
reasonable
theory
,,
b
K
d
on
the
similarity
of
the
wild
musky
odor
of
the
grapes
r:
th
the
characteristic
animal­
like
odor
of
a
fox
or
of
a
fox's
dtr.
The
gapes,
which
were
originally
called
Fox
Grapes
were
fuflhCr
subdivided
into
the
Xorthern
Fox
Grapes
(Viris
iabrus­
r;)
and
the
Southern
Fox
Grapes
(Vilis
rotundifolio)
(Bailey,
i
PO8
).
Methyl
anthranilate
has
been
suggested
as
an
important
rr.
mpofient
of
the
characteristic
Concord
(the
most
important
ra::
vc'
American
variety)
aroma
for
over
50
yr
(Power,
1921).
has
since
been
implicated
as
a
major
contributor
to
"foxi­

e
a
*'
in
American
wines
(Sale
and
Wilson,
1926;
Winkler,
f
Q
7
2
).
Amerine
et
al.
(1959),
using
Catawba
as
an
example,
rqgcsted
that
"foxy"
varieties
need
not
contain
methyl
mthranilate.
This
paper
is
intended
to
evaluate
the
importance
of
methyl
3n:
hranilate
as
an
odor
constituent
of
wines
produced
from
*dely
differing
grape
varieties.

MATERIALS
&
METHODS
*Hca
i(
different
varieties
of
wine
were
obtained
from
the
experimental
*2
z
k
:I
~o
n
of
the
New
York
State
Agricultural
Experiment
Station
for
*­
w.
this
experiment.
AU
wines
were
prepaied
according
to
the
stan­
d
*.
`
rmt­
dure
of
Pool
et
al.
(1976).
The
wines
represent
four
snecies
(
'1
T
~c
wineswere
extracted
with
Freon
11
3,
concentrated,
andana­
'
­4
k
r
r
methyl
anthranilate
by
the
gas
chromatographic
method
of
~c
!s
.~t
r
e1
at.
(1976)­
Sensory
evaluation
Two
separate
sensory
analyses
were
performed.
First,
the
threshold
of
methylanthraniiate
in
wine
(V.
viuiferu
cv.
WhiteRiesling)
was
estimated
by
comparing
the
odor
of
wines
with
varying
concentrations
of
added
methyl
anthranilate.
Rankings
were
done
by
the
taste
panel
to
determine
the
point
at
which
methyl
anthranilate
could
no
longerbe
detected.
Second,
an
evaluation
of
relative
foxiness
in
selected
varieties
wasperformed
by
the
same
panel.
The
wines
were
selected
to
vary
in
nature
andintensity
of
odor
and
to
containmethylanthranilate
at
various
levels.
This
permits
a
direct
comparison
of
foxiness
and
methyl
anthranilate
concentration.
The
sevenpanelists
employed
inthiswork
are
experienced
in
the
critical
sensory
evaluation
of
wines.

RESULTS
&
DISCUSSION
THE
THRESHOLD
of
methyl
anthranilate
in
wine
was
esti­
mated
by
sensory
evaluation.
The
panel
was
given
clean,
young
White
Riesling
wine
samples
with
methyl
anthranilate
added
at
various
levels.
The
ranking
was
conducted
in
two
groups
of
four
samples
each
(0.00,
0.01,
0.03,
0.1
and
0.1,0.3,
1.0,
3.0
ppm).
The
results,
shown
in
Table
1
,
indicate
that
the
panel
could
successfullf
distinguish
higher
levels
of
methyl
anthranil­
ate.
They
were,
howeker,
unable
to
correctly
rank
the
less
'
concentrated
set.
The
threshold
can
be
estimated
from
these
results,
Each
set
of
four
wines
contained
a
sample
with
0.1
ppm
methyl
anthranilate.
The
panelists
could
correctly
rank
this
sample
within
the
set
of
higher
concentrations
while
they
could
not
do
so
within
the
less
Concentrated
set.
This
indicates
that
0.1
ppm
could
not
actually
be
detected
but
was
ranked
.
correctly
by
the
process
of
elimination
only
when
samples
of
higher
concentrptipn
were
available
for
reference.
The
sample
containing
0.3
pp,
m
was
consistently
ranked
correctly
so
it
can
be
assumed
that
this
concentration
of
methyl
anthranilate
is
at
or
near
the
threshold
level
in
White
Riesling
wine.
With
this
information,
a
mdre
meaningful
estimation
of
the
importance
of
methyl
anthranilate
in
wine
aroma
can
be
made.

Table
1­
The
threshold
of
methyl
anthranilate
in
white
riesling
wine
Methyl
anthranilate
(ppm)
Correct
rank
AVQ
ran@

0.00
1
3.1
0.01
2
2.3
0.03
3
1.9
0.1
0
`4
2.7.
­
0.10
4
4.3
0.30
5
5.3
1
.oo
6
5.9
3­
00
7
6.6
,a
Average
rank
of
seven
experienced
panelists
Volume
42
(1977)­
JOURNAL
OF
FOOD
SCIENCE­
57
.
...
~~
~
~~~­~
~

f
"JCIlJRNAL
OF
FOOD
SCIENCE­
Volume
42
(?
Y//
J
The
majority
of
the
45
varieties
analyzed
were
found
to
be
;
free
of
methyl
anthranilate.
Table
2
fists
these
varieties,
their
parentages,
and
the
vintage
of
each
sample.
Only
six
varieties
3.
Additional
samples
of
Niagara,
Concord
and
Delaware
were
(?
'
analyzed
and
considerable
variation
in
the
level
of
methyl
anthranilate
due
.to
vintage
and
enological
technique
was
found.
In
1949,
Robinson
et
al.
reported
that
methyl
anthra­
nilate
was
formed
in
grapes
during
the
final
stages
of
ripening.

sample
was
harvested
at
16.0°
B
and
showed
3.1
ppm
methyl
t
i
9
%
were
found
to
contain
methyl
anthranilate
as
shown
in
Table
L
%,/

i
This
certainly
seems
to
b
e
t
h
e
case
for
Niagara.
The
1970
i
!4
1
'

Table
2­
Wine
varieties
without
methyl
anthranilate
Variety
Vintage
­
French
hybrid9
Aurore
(5.788
X
5.29)
Baco
22A
(Folle
blanche
X
Noah)
Chancellor
(S.
5163
X
S.
880)
Chelois
(S.
5163
X
s.
5593)
OeChaunacb
6.51
63
x
S.
793)
Fochb
(Mgt.
101­
14
X
Gold
Riesling)
b
n
d
o
t
451
1
(L244
X
S.
V.
12­
375)
Ravat
51
(5.6095
X
Pinot
blanc)
S
e
i
b
e
l
10868
6.5163
X
5.5593)

Geneva
hybrids
Canada
Muscat
Cayuga
White
GW­
1
(Catawba.
X
Seneca)

i/.;...,
?,
GW­
2
(Seyval
X
Schuylerl
I
GW­
4
(SeyvaI
X
Sene=
')
L
T
GW­
5
(Pinot
Blanc
X
Ontario)

~

GW­
6
(S
e
y
~l
X
Seneca)

GW­
7
fSeywI
X
Schuyler)
GW­
8
(Pinot
Blanc
X
Aurore)
GW­
10
(SeyvaI
X
Chardonnay)
GR­
1
(Buffalo
X
Baco
Noir)
GR­
3
(Buffalo
X
Baoo
Noir)
GR­
6
(Buffalo
.x
Baco
Noir)
GR­
7
(Buffalo,
X
Baco
Noir)
N.
Y.
Muscat
Vitis
aestivalis
Wild
Summer
Vir;$
labrusca
(and
crosses)
Diamond
1970
1972
1969
1970
1970
1973
1970
1973
1973
1969
1974
1970
~

1967
1967
1971
1967
1968
anthranilate
while
the
1975
sample
matured
to
only
12­
50
and
showed
0.6
ppm.
,
Of
the
six
varieties
containing
methyl
anthranilate,
Catau
ba,
Delaware
and
Elvira
may
have'insufficient
methyl
anthr
nilate
to
influence
the
aroma
of
these
wines
since
the
COncel
trations
determined
are
near
the
threshold
level.
These
wine
however,
are
decidedly
American
in
character.
A
wide
range
of
native
American
wines
are
called
"fox)
but
methyl
anthranilate
appears
t
o
be
important
in
only
a
fe
of
them.
In
order
to
examine
the
correlation
of
"foxiness
with
methyl
anthranilate
concentration,
a
series
of
wines
w;
selected
for
further
taste
panel
work.
The:
wines
were
selected
to
have
a
wide.
range
of
apparer
foxiness
and
to
subject
the
panelists
to
other
varietal
aromi
that
may
or
may
not
be
considered
foxy
by
the
individu
judges.
The
six
varieties
selected
were.
Canada
Muscat,
Carlo
Concord,
Moored,
Niagara
and
White
Riesling.
A
seventh
sari
ple
was
prepared
by
adding
0.7
ppm
methyl
anthranilare
t
the
same
White
Riesling
wine.
In
the
experiment
panelists
we1
asked
to
smell
the
wines
and
rank
them
according
to
decrea
ing
foxiness.
The
results
are
shown
in
Table
4.
There
appeal
to
be
no
correlation
between
methyl
anthranilate
concentr;
tion
and
the
relative
foxiness
of
these
wines.
The
fact
that
the
White
Riesling
sample
with
0.7
p
p
~
added
methyl
anthranilate
was
ranked
higher
in
foxiness
tha
the
pure
Riesling
may
indicate
that
the
ester
makes
some
cor
tribution
to
the
foxy
character.
However,
other
varieties
wit
less
methyl
anthranilate
were
consistently
judged
higher
in
thl
character.
In
fact,
three
of
the
four
wines
judged
highest
i
Table
3­
Wine
varieties
with
methyl
anthranilate
Methyl
anthranilate
V
a
r
i
e
t
y
V
i
n
t
a
g
e
Parentage
bpm)

1972
Niagara
1970
V.
labrusca
1972
Concord.
1970
V.
labrusca
1
967
Concord
1975
V.
Iabrusca
1972
Concord
1972
V.
labrusca
1972
Niagara
1975
V.
labrusca
.
1973
lves
1972
.
V.
labrusca
1969
Catawba
1972
V.
Iabrusca
X
?
3.102
1.752
1
.110
0.699
0.61
7
0.381
0.1
78
Delewaren
1975
,
V.
labrusca
X
?
0.102
Elvira
.
1972
V.
labrusca
X
V.
riparia
trace
Oelaware
1973
V.
iebrusca
X
?
0.
M)
O
"

a
Hot
pressedsamples
.
1970
1966
Dutchess
1967
Eumelan
1967
lsabella
.
­
1966
Moored
1975
Table
4­
Evaluation
of
"'foxiness"
in
selected
varietal
wines
Vincent
.
1970
Methyl
Vitis
rotundifolia
nnthranilate
Carlos
1974
Variety
foxiness'
.
.
,
fppml
Fry
1974
Hunt
1974
Carlos
5.1
'
0.0
I
Concord
49
1.7
­
Canada
Muscat
4
­4
Pinot
Blanc
1971
0.0
Moored
Pinot
Blanc
4.1
1972
0.0
A:
1
..­\

Sylvaner
Niagara
1973
3.7
0.6
Rieslingb
3.3
White
Riesling
1972
Riesling
2.4
0.0
0.7
Vitis
vinifera
:r
­
i
",
._

a
Seedling
abbreviations;
S
=
Seibel;
Mgt.
=
Millardet
et
de
Grasset;
a
Rank
averages,
seventasfers
on
a
seven
point
scale
bHot
pressed
samples
L
=
Lendot;
S.
V.
=
Seyve­
Villard,
.
.
.
.
Methyl
anthranilate
added
(0.7
ppm)
using
standard
solution
in
EtOH
­
..

­I,,
..

..
.­
,
.
­
..
ataw­
*
inthra­
,
oncen.
win?­
.
4
rhb
&Some
"foxy"
wine
varieties
frm
of
methyl
snttiranilstr
­­­"
Variety
Parantap
d
WiW
Omda
Muscat
V.
vinifera
X
V.
labrusce
Carlos
V.
rotundifofia
Diamond
labrusca
X
?
outchess
V.
labrusca
X
7
FrY
V.
rotundifolia
M,
wouri
Riesling
V.
labrusca
X
V.
riparia
Eumetan
V.
aestivalis
tabella
V.
labrusca
Moored
V.
labrusca
(hybrid)
N.
y.
Muscat
V.
vinifera
X
V.
labrusca
vtncent
V.
labrusca
(hybrid)
R
d
foxiness
were
entirely
free
of
methyl
anthranilate
including
Carlos
which
was
rated
the
most
foxy
of
all
wines
tested.
The
case
of
Carlos
(a
bronze
variety
of
the
muscadine
grape,
I?
rc,
rundifoliu
according
to
Ferree,
1975)
is
not
unique.
Table
5
lists
several
more
varieties,
both
red
and
white,
of
various
par­
entages
that
are
generally
considered
t
o
produce
foxy
wines
and
art'
free
of
methyl
anthranilate
a
t
levels
determined
in
this
work.

CONCLUSIONS
METHYL
ANTHRANILATE
occurs
with
relative
infrequency
in
wine
varieties.
It
is
limited
to
some
varieties
of
V.
hbrusca
and
a
few
crosses
thereof.
In
addition,
the
concenlration
methyl
anthranilate
present
in
many
cases
appears
lo
be
bel
the
threshdld
level
which
furlher
minimizes
its
importance
win
e.
aroma.
If
present
in
large
quantities,
as
in
some
samples
of
Concc
and
Niagara
wines,
methyl
anthranilate
prohahly
contribu
to
the
characteristic
"grapey"
or
"fruity"
aroma­
of
thc
wines.
However,
any
correlation
of
methyl
anthranilate
wi
foxiness
seems
to
be
absent.
It
cannot
he
said
that
mett
anthranilate
does
not
contribute
t
o
foxiness
but
it
is
clear
t
h
it
is
not
the
primary
source
of
the
odor.
Unlike
the
occuren
of
methyl
anthranilate,
foxiness
is
not
limited
to
varieties
K
.
lahrusca
but
is
found
in
a
wide
range
of
native
Americ.
wines
of
dissimilar
parentage.
Further
analytical
­and
sensory
work
yifl
be
required
determine
the
cause
of
the
native
American
wine
aron
known
as
foxiness.

REFERENCES
Amerine,
h1.
A..
Roesrler,
E.
B.
and
Filipello.
F.
1959.
Modern
sensol
methods
of
evaluating
wine.
Hilgardia
28:
477.
Bailey,
L.
H.
1898.
"The
Evolution
of
our
Native
Fruits."
The
Ma
millan
Co..
New
York.
Ferree.
M.
E;
1975,
Muscadine
grape
culture.
Bull.
739.
Univ.
of
G,

Nelson.
R.
R..
Aeree,
T.
E..
Lee,
C.
Y.
and
Butts,
R.
M.
1976.
Gas­
liqui
CoL
of
A&,
Athens.
Ga
chomatographic
determination
of
methyl
anthranilate
in
win1
JAOAC.
In
pres
Pool.
R.
M..
Robinson.
W.
B..
Einset.
J.,
Kimball.
K.
H..
Watson
J.
P.
an
Bertino,
J.
J.
1976.
Vineyard
and
cellar
notes
1958­
197a.
Specis
Power,
F.
B.
1921.
The
Detection
of
Methyl
Anthranilate
in
Fmi
Report
No.
22..
N.
Y.
St.
Ag.
Exp.
Sa..
Geneva.
N.
Y.

Juices
J.
Amer.
Chem.
SOC.
43:
377.
Robinson.
W.
B..
Shaulis.
N.
J.
and
Pederson,
C.
S.
1949.
Ripenin
Studies
of
Grapes
Grown
in
1948
for
Juice
Manufacture.
.Fruit
Prod
J.
and
A
m
Food
Mfg.
29(
2):
36.
'Sa'e.
J.
WI
and
Wilson.
J.
B.
1926.
Distribution
of
voIatile
flavors
L
Winkler.
A.
J.
1972.
''General
Viticulture."
University
of
Caliiorni;
grapes
and
grape
juices
J.
Agr.
R
e
s
33:
301.

hfs
received
6/
2/
76;
revised
7/
9/
76;
accepted
1/
16/
16.
bess.
Berkeley.
Calif.
iNHERITANCE
OF
METHYL
ANTHRANILATE
AND
TOTAL
A.
G.
Reynolds,
T.
Fuleki,
and
W.
D.
Evans
Respectivelyformergraduatestudent.
Department
of
HorticulturalScience.
University
of
Guelph.
Guelph,
Ontario.
Canada
N1G
2W1;
Research
Scientist,
Horticultural
Products
Laborato­
ry.
Horticultural
Research
Institute
of
Ontario,
Vineland
Station,
Ontario
LOR
2EO;
and
Associate
Professor,
Department
of
Horticultural
Science,
University
of
Guelph.
Mr.
Reynolds
is
currently
pursuing
further
graduate
study
in
the
Department
of
Pomology
and
Viticulture,
New
York
State
Agricultural
Experiment
Station,
Cornell
University,
Geneva,
New
York
14456.
Taken
in
part
from
the
MSc
Thesis
of
the
senisr
author.

Thesenior
authorwishes
to
thank
Dr.
R.
E.
Subden,
Department
of
Botany
and
Genetics,
University
of
Guelph,
forhisencouragementandguidancethroughoutthecourse
of
this
work.
Presented
at
the
Fifth
Annual
Meeting
of
the
Eastern
Section,
American
Society
of
Enologists,
August
8.
1980.
Erie,
Pennsylvania.

Financial
Assistance
from
the
Natural
Sciences
and
Engineering
Research
Council
of
Canada
is
gratefully
acknowledged.

Manuscript
25
May
1981
,

Revised
manuscript
received
2
November
1981.

Accepted
for
publication
7
November
1981.

ABSTRACT
Methyl
anthranilate
and
total
volatile
esters
concen­
trations
were
determined
for
two
families
of
grape
'
seedlings
which
resulted
from
crosses
made
at
Vineland,
Ontario
in
1972.
Chi­
square
analysis
of
the
segregation
patterns
suggested
that
three
dominant
complementary
genes
were
involved
in
the
inheritance
of
methyl
anth­

,.,
c­
~

ranilate
and
two
for
total
volatile
esters.
High
heterosis
r\.
j
and
broadsense
heritability
values
for
both
characters
In
Ontario,
,grapes
are
grown
on
approximately
10000
ha,
about
half
of
which
are
still
planted
to
the
traditional
Vitis
labruscana
cultivars.
Improvement
of
the
Ontario
grape
and
wine
industry
to
allow
it
a
greater
competitiveness
domestically
has
involved
both
the
planting
of
French
hybrid
and
V.
uinifera
cultivars,
and
the
introduction
of
new
wine­
grape
cultivars
from
grape
breeding
programs
such
as
that
of
the
Horticultural
'
Research
Institute
of
Ontario
a
t
Vineland
Station,
On­
tario.
This
latter
breeding
program
has,
as
one
of
its,
objectives,
the
elimination
of
the
undesirable
"foxy"
or
labrusca
flavor
character
from
its
select,
ions;
however,
due
to
the
climatic
constraints
of
the
area,
the
objectives
of
winter
hardiness,
disease
resistance,
and
vigor
often
'
take
precedence
over
this
objective,
such
that
the
utili­

'
zation
of
labrusca­
flavored
but
hardy
cultivars
as
par­
en'ts
is
often
necessary.
Because
of
this
problem,
this
study
was
initiated,
to
elucidate
a
mode
of
inheritance
for
the
labrusca
flavor
charact.
er,
and
to
make
recom­
mendations
for
its
avoidance
in
wine­
grape
breeding
programs.
dyid'

~

.
..
An
early
reference
to
"foxiness"
in
grapes
was
made
;:
by
Hedrick
et
al.
(9),
who
described
the
aroma
of
certain
/­,;
'
V­
labruscana
cultivars
as
being
reminiscent
of
a
fox's
den
or­
burrow.
Early
attempts
to
characterize
this
flavor
Liq;/
j
chemically'
were
reported
by
Power
and
Chesnut
(Is),
,II
I
l
l
suggested
dominance.
Statistical
differences
between
the
families
and
examination
of
the
ancestries
of
the
parental
cultivars
allowed
the
postulation
of
genotypic
formulae
for
the
parents.
No
correlation
wasfound
between
methyi
anthranilate
and
volatile
esters,
or
among
either
of
these
characters
and
soluble
solids,
winter
hardiness,
or
vigor.

.
"
.­
?'
.;
,

who
found
a
relationship
between
labrusca
flavor
char­
acter,
V.
labrusca
ancestry,
and
the
ester
methyl
anth­
ranilate
(MA).
Sale
and
Wilson
(18)
reported
similar
results,
but
indicated
that
other
volatile
esters
such
as
ethyl
acetate
also
made
an
important
contribution
to
the
flavor.
Ho'lley
et
al.
(11)
detected
eight
components
in
the
essence
of
Concord
juice,
of
..
which
MA,
methyl
acetate,
and
ethyl
acetate
were
predominant.
They
concluded,
however,
that
many
other
important
aroma­
tic
substances
were
probably
overlooked
due
to
the
iimitation
of
their
methods.
The
development
of
gas
chromatography
has
allowed
much
more
sophisticated
and
detailed
investigations
int.
0
the
volatile
composition
of
fruit
crops.
Neudoerffer
et
al.
(13)
identified
32
compounds
in
Concord
essence
using
gas
chromat.
ography:
Methyl
anthranilate
was
not
detected
because
it
did
not
elute
from
the
column.
Similar,
but
less
extensive
was
the
list
of
Stevens
et
al.
(201,
who,
$sing
gas­
liquid
chromatography­
mass
spec­
trometry,
sugg,
ested
methyl­
3­
buten­
2­
01
to
be
of
great
significance,
along
with
ethyl
acetate.
Sixty
components
of
Concord
essence
were
detected
by
gaschromato­
graphy­
mass
spectrometry
by
Stern
et
a/:
(191,
of
which
a
series
of
crotonate
esters
and
an
alkylthioester
were
viewed
as
being
important,
as
well
as
ethyl
acetat.
e,
MA,
and
several
other
esters.
For
the
facilitation
of'
selection
of
non­"
foxy"
culti­

14
',;
I
Am.
J.
Enol.
Vitic.,
Vot.
33,
No.
1,
1982
in
the
wine­
grape
breeding
program
in
Ontario,
;r7.­.>,
Li
(6,
r)
has
devised
an
index
for
t.
he
labrusca
flavor
acter
based
on
the
concentration
of
MA
and
total
,,,
latile
esters
(T\
'E)
in
the
fruit.
This
score,
known
as
\'ineland
Grape
Flavor
Index
(VGFI),
has
intro­
dltced
a
high
degree
of
speed
and
objectivity
into
the
celec'ti()
n
process.
The
derivat.
ion
of
this
index
was
based
i,,,
the
high
correlation
between
labrusca
flavor
charac­
ter
M
A
and
TVE
concentration.
The
inheritance
of
specific
flavor
characters
in
Craps.
most
notably
muscat
flavor,
have
been
investi­

Pa
ted
by
a
number
of
European
researchers
.i
n
.
V.
l
.i
,l
i
f
p
m
.
Wagner
(231,
using
organoleptic
evaluation,
,tlggel;
ted
t
h
a
t
a
t
least
five
dominant
complementary
plies
were
involved
in
muscat
flavor
inheritance.
Tsek­
nlislrenk<>
and
Filinova
(22),
considered
the
character
to
be
monogenic
and
dominant.
M'agner
et
ai.
(24)
p0st.
u­
laled
a,
mechanism
for
muscat
flavor
inheritance
based
o
n
the
heritability
of
the
five
major
terpenes
normally
Ihought
to
comprise
the
flavor.
It
was
found
that
four
of
the
five
were
inherited
identically
via
a
two­
gene
system,
ahereas
the
fifth.
linaloijl,
possessed
a
more
complex
inheritanre.
Later
studies
(25,26)
part.
ially
refuted
t.
his
hypothesis,
by
indicating
a
correlation
between
terpenes
previously
thought
to
be
unrelated,
and
a
lack
ofcorrela­
titm
between
terpene
concentration
and
intensity
of
muscat
flavor
as
perceived
organoleptically.

.­
..
Genetic
investigations
into
the
labrusca
flavor
char­

"
':
have
not
been
previously
carried
out.
The
objec­

"
of
this
research
were
to
elucidate
a
mode
of
mdritance
for
the
labrusca
flavor
charact.
er
based
on
31.4
and
TV­
E
content,
and
to
make
recommen'dations
on
its
minimization
in
wine­
grape
breeding
programs.
MATERIALS
AND
METHODS
Ayailable
genet.
ic
material:
All
mat.
eria1
for
our
investigations
was
grown
and
harvested
at.
the
Grape
Research
Station,
Vineland,
Ontario.
Cultural
methods
were
carried
out
according
to
Ontario
recommendations
(141,
and
those
of
Bradt(
1).
Two­
families
of
seedlings,
namely
V.
7218
and
V.
7219
were
chosen
for
investigation,
tn,
th
ofwhich
were
the
result
of
the
hybridization
of
one
high­
and
one
low­
VGFl
parent
carried
out
a
t
Vineland
in
1972.
Such
a
parentage
maximized
t.
he
probability
of
ohraining
a
significant
number
of
non­
labrusca
and
!at)
rusca­
flavored
seedlings
in
the
progenies.
Family
V.
7218
was
a
cross
between
V.
5407i
(de
Chapnac
X
Concord,
Yineland
1954;
MA
=
0.14
ppm,
TVE
=
21
ppm)
and
Bertille­
Seyve
5563
(S.
6905
X
B.
S.
:U­
4%
M
A
=
nil,
TVE
=
3
ppm).
A
total
of
229
seedlings
wab
available
in
the
autumn
of
1978.
Family
V.
7219
was
a
cross
between
V.
54077
and
V.
50061
(Alden
X
Lo­
manto,
Vineland
1950;
hlA
=
nil,
TVE
=
I
pprn).
In
the
autumn
of
1978,
226
seedlings
were
available
in
this
family.
Fifty
seedlings
were
chosen
randomly
from
each
fa2
,.
'..
for
our
investigations.
of
vigor
and
hardiness:
In
May,
1978,
ne^
in
both
families
were
rated
on
a
scale
of
0
to
4
r(
k
vigor
and
wint.
er
hardiness.
For
vigor,
shoot
length
and
number
constituted
the
criterion
for
the
rating,
Uihere
''4''
represented
the
most
vigorous.
I
n
assessing
METHYL
ANTHRANILATE
­
15
winter
hardiness,
the
amount
of
killed
buds
was
consid­
ered
such
that
a
rating
of
"0"
indicated
the
hardiest
vine.
H
a
r
v
e
s
t
i
n
g
a
n
d
s
t
o
r
a
g
e
:
M
a
n
y
s
t
u
d
i
e
s
(3,4,15,17,18)
have
indicat.
ed
t.
bal
MA
concentration
increases
with
advancing
fruit
mat.
urit.
y,
and
decreases
somewhat
as
the
fruit
becomes
overripe.
It
was
thus
imperative
to
harvest
each
seedling
at
it.
s
optimum
maturit.
g.
To
accomplish.
this,
preharvest
checks
were
initiated
on
September
1,
1978,
and
were
continued
twice
weekly
until
all
fruit
was
harvested.
The
fruit
was
considered
ripe
if
the
soluble
solids
content
attained
20"
Brix
or
greater,
or
was
t.
he
same
for
three
consecutive
sampling
dates,
as
measured
by
a
hand
refractometer.
Harvesting
took
place
between
September
15
and
Oct­
o­.
ber
18,
1978.
Once
the
fruit
of
each
genotype
was
harvested,
it
was
placed
in
a
plastic
bag
and
transported
to
the
Biochem­
istry
Laboratory,
Department.
of
Horticultural
Science,
University
of
Guelph.
Each
sample
was
then
washed,
.
destemmed,
and
replaced
int.
o
its
bag.
Subsequent
st.
or­
age
t.
ook
place
at
­31OC.
Sample
preparation:
Each
sample
was
ground
in
a
hand
mill,
and
50
g
of
the
homogenate
was
steam­
distilled
(SGA
Scientific,
Bloomfield,
N.
J.
No.
JD­
2115)
until
100
mL
of
distillate
was
collected.
This
procedure
was
duplicated
for
each
genotype.
A
50
mL
beaker
of
the
homogenate
was
also
retained
for
subsequent
soluble
solids
determination
using
an­
Abbe­
type
refractornet.
er.
Methyl
anthranilate
determination:
The
determi­
nation
of
MA
concentration
was
done
according
to
the
.

fluorometric
procedure
of
Casimir
et
d
l
.
(2)..
Fluoromet­
ric
determination
was
carried
out
on
a
Turner
Model
110
fluorometer
(G
.
K.
Turner
ASSOC.,
Palo
Alto,
Calif.)
equipped
with
an
attachment
for
accepting
glass
cu­
vettes.
A
narrow
pass­
(360
nm)
and
a
sharp
cut­
(415
nm)
filter
served
as
primary
and
secondary
filters
re­
spectively.
Sample
fluorometric
readings
were
related
to
hlA
concentration
through
the
use
of
st.
andard
curves.
using
a
series
of
aqueous
MA
solutions
that
ranged
in
concentrat.
ion
from
0.05
to
10
ppm.
Total
volatile
esters'
determination:
The
det.
ermi­
nation
of
TVE
concentration
was
carried
out
by
Hill's
method
(10)
as
described
by
Thompson
(21).
Spectro­
photometric
determination
was
done
using
a
Coleman
Junior
I1
Model
6/
20
spectrophotomet.
er
(Coleman
In­
struments,
Maywood,
Ill.)
set.
at
a
wavelength
of
540
nm.
Absorbance
was
related
to
TVE
concentrat.
ion
by
the
use
of
standard
curves,
using
a
series
of
aqueous
ethyl
acetate
solut.
ions
that
ranged
in
concent,
ration­
from
0
to
125
ppm.

RESULTS
AND
DISCUSSION
Variability
within
families:
The
variability
en­
countered
within
the
families
was
.
great,
but
specific
patterns
were
nonetheless
existent.
Hist.
ograms
(Figs.
1
and
2)
relating
seedling
frequency
and
MA
or
TVE
concentration
illustrate
this
variability,
suggesting
an
absence
of
simple
Mendelian
inheritance
in
either
sys­
tem.
The
lack
of
a
normal
distribution
in
t.
he
data
also
suggested
that.
a
typical
polygenic
system
was
also
not
Am
.I
Fnol
Vitir
16
­
METHYL
ANTHRANILATE
..

­f
0
V.
7219
SEEDLING
CLASS
PPm
M
A
x
100
Fig.
1.
Distribution
of
seedlings
in
families
V.
7218
and
V.
7219
according
to
their
methyl
anthranilate
concentration.

40.

>
V
2
Y
V.
7218
V.
7219
SEEDLING
CLASS
ppm
TVE
Fig.
2.
Distribution
of
seedlings
in
families
V.
7218
and
V.
7219
according
to
their
volatile
esters
concentration.

present.
Most
noticeable
vias
a
distinct.
bimodal
distribution
which
could
be
observed
for
both
characters
in
each
family,
whereby
the
majority
of
seedlings
lag
to
the
extreme
left
as
one
group,
and
the
others
were
scattered
to
the
right.
in
a
less
homogeneous
group.
This
distribu­
tion
conveniently
placed
the
seedlings
into
one
of
two
categories,
that
of
labrusca"
or
"non­
labrusca",
in
terms
of
MA
and
TVE.
The
line
of
demarcation
bet.
ween
the
two
categories
for
M
A
appeared
to
be
close
to
0.10
ppm
(Fig.
11,
a
value
suggested
by
Nelson
et
al.
(13)
as
being
the
compound7s
organoleptic
threshold
in
wine.
For
TVE,
the
point
of
delineation
bet.
ween
the
two
classes
was
found
to
be
about
12
ppm
(Fig.
2),
a
level
suggested
by
Fuleki
(5)
as
being
that.
where
labrusca
flavor
be­
comes
noticeable
organoleptically.
Number
of
genes
involved
After
placement
of
the
seedlings
into
either
the
%on­
labrusca''
or
"labrusca"

,/
'
class
i
n
terms
of
MA
and
TVE.
the
data
were
subjected
to
Chi­
square
goodness­
of­
fit
tests.
Table
1
indicates,
faF
MA,
the
ratios
were
5:
3
and
i
:l
for
families
V.
7218
and
Y.
7219
respectively.
These
ratios
corresponded
with
a
hypothesis
o
f
t
hre.
e
dominant
complementary
genes
for
t.
he
inheritance
of
this
compound.
The
large
variation
io
MA
concentration
in
the
"labrusca"
class
could
be
at.
trib­
uted
to
modifier
genes,
an
explanation
offered
by
Wag.
ner
(231,
for
the
control
of
muscat
flavor.
The
year­
to.
year
variation
in
the
concentration
of
MA
(and
TVE
well)
inmost
V.
labruscana
cultivars
could
also
be
explained
by
modifier
genes.

Table
1.
Chi­
square
goodness
of
fit
analysis
of
methyl
anthranila;
concentration
in
families
V.
7218
and
V.
72193
FamilyNon­
labruscab
Labruscac
Ratio
P
­

ObservedExpectedObservedExpected
V.
7218
32
31.25
18
18.75
5
3
.80­.
9<
V.
7219
44
43.75
6
6.25
7:
l
.90­.
95
a
Observedand
expected
valuesarebased
on
a
sample
of
50
b
Methyl
anthranilate
concentration
=
0
to
0.1
ppm.
c
Methyl
anthranilate
concentration
=
0.1
ppm
or
higher.
­
seedlmgs.

In
the
case
of
TVE,
5:
3
ratios
were
obtained
for
both
families
(Table
2).
These
corresponded
in
simplest
t.
erms
to
a
hypothesis
of
two
dominant
complementq
genes
for
the
control
of
this
group
of
compounds.
As
previously
indicated
for
MA,
modifier
genes
were
likely
responsible
for
the.
large
variation
in
TVE
concent.
ration
­
in
the
"labrusca"
class.

Table
2.
Chi­
square
goodness
of
ftt
analysis
for
total
volatile
ester;
concentration
in
familiesV.
7218andV.
7219a
.
`

Famity
Non­
IabruscabLabruscac
Ratio
P
ObservedExpectedObservedExpected
V.
7218
31
31.25
19
18.75
­513
.90­.
95
V.
7219
32
31.25
18
18.75
5:
3
.80­.
90
a
Observed
and
expected
valuesarebased
on
a
sample
of
50
b
Total
volatile
esters
concentration'=
0
to
12
ppm.
c
Total
volatile
esters
concenfration
=
12
pprn
or
higher.

It
must
be
stressed
that
these
dat.
a
possess
some
degree
of
weakness
due
to
the
small
sample
sizes.
Other
rat,
ios
teg.
3:
l
or
9
7
)
are
possible,
although
not
as
statistically
acceptable.
Insofar
as
gene
nomenclature
is
concerned,
three
genes,
henceforth
designated
as
M,
A,
and
F,
are
sug
gest.
ed
for
the
synt.
hesis
of
hlA
in
grapes.
Seedlings
in
the
"labruscar
class,
which.
were
characterized
by
rela­
tively
high
hfA
concentrations,
could
logically
result
from
all
three
loci
being
in
the
dominant
condition.
The
"non­
labrusca"
class
contained
seedlings
either
with
no
MA
or
with
concentrations
less
than
0.10
ppm.
Those
cont.
aining
no
MA
whatsoever
would
have
recessiveness
at
all
three
loci,
whereas
those
containing
minute
amounts
of
the
compound
could
have
at
least
one
dominant
locus.
In
the
case
of
TVE,
two
genes,
designated
as
V
and
E.
are
suggest.
ed
for
volatile
ester
synthesis
in
grapes.
A
seedling
possessing
both
genes
in
the
dominant
condi­
tion
would
be
one
characteristic
of
the
"labrusca"
class:
seedlings.

Am.
4.
Enol.
Vitic.,
Vol.
33,
No.
I
,
1982
Heterosis
=
­
x
100
F1
F,
+
P,
,,­
here
F,
is
the
averagcvalue
of
a
particular
character
in
the
F,
generation
and
P,
is
the
midparent
value
for
that.
The
high
values
displayed
for
both
hk4
and
T\
'E
i
i
the
two
families
suggested
that
dominant
genes
were
most
likely
present
(Table
3).

&e
3.
Transgressive
segregation
(heterosis)
of
methyl
anthranilate
a,
s
total
volatile
esters
over
the
parental
cultivars
In
families
V.
7218
and
V.
7219
"­
f
a
d
y
Heterosis
(0.
b)

Methyl
Total
volatile
anthranilate
v
7218
491.4
65.0
\'
7219
55.7
­65.5
.­

i
(i
:
High
broadsense
heritability
(h')
has
also
been
impli­
:
cated
as
a
criterion
for
dominance.
It
is
expressed
as
the
percentage
of
the
total
variance
of
a
progeny
that
can
be
1.
attributed
to
genotype,
and
can
be
summarized
in
the
1.
following
formula:

z
0,
'

3
ff,
'
+
4
2
..

4
hz
=
x
100
I
where
o­
i
is
variance
due
to
genot.
ype
and
u:
is
variance
i
due
to
environment.
The
quantity
u:
was
broken
down
I
­
into
ua
and
c;,?
which
represent
variance
found
between
duplicate
samples
wit.
hin
a
genotype
and
variance
due
to
error
respectively.
An
estimate
of
the
u,",
values
for
MA
i
and
TYE
were
obtained
from
an
analysis
of
variance
of
i
51.4
and
TVE
concentrations
in
several
1,
'.
labruscana
cultivars
over
a
nine­
year
period.
The
u3
values
were
t
ohtained
from
a
one­
way
model
LI
analysis
of
variance
of
'.
each
family.

1
..
The
h2
values
for
both
characters
viere
relatively
high
*
In
both
families
(Table
4),
with
the
exception
of
that.
for
2
,It.\
infarn'ily
V.
7219.
Such
information
uas
evidence
i
.'that
dominant.
genes
were
present.
t
I
t
8
­
.

Ta%
4.
Variance
components
and
broadsense
heritability
of
methyl
anthranilate
and
total
volatile
esters
in
families
V.
7218
and
V.
721ga
.
L
,>_

.z
I
Character
4
I
,
3rnnlly
METHYL
ANTHRANILATE
­
17
Parental
genotypic
formulae:
The
compilation
of
information
previously
reported
here
and
the
consider­
ation
of
the
ancestrx
of
the
parental
cultivars
allowed
the
postulation
of
genotypic
formulae
for
the
parents.
Since
the
seed
parent
V.
5407i
was
common
for
the
two
families.
any
statistical
differences
between
the
families
were
attributable
t.
o
the
pollen
parents
B
S
5563
and
Y.
50061.
Basic
stat.
istics
such
as
mean
and
variance
indicated
large
differences
between
the
families
in
terms
of
hfA
and
TVE
(data
not
shown).
Further
statistical
analysis
examining
coefficients
of
variation
and
equalit_
v
of
variances
confirmed
these
differences,
and
strongly
suggested
t.
hat
the
pollen
parents
were
not
the
same
genotypically,
at.,
least
in
terms
of
MA
(Tables
5
and
6).

Table
5.
Coefficients
of
variation
formethylanthranilateand
total
volatile
esters
concentration
in
families
V.
7218
and
V.
7219.
Family
Character
Methyl
anthranilateTotalvolatileesters
.
V
.7
2
1
8
5
7
3
.4
8
V.
7219
3336.62
27.93
5622.44
Table
6.
Equality
of
variancesformethylanthranilateand
total
volatile
esters
in
families
V.
7218
and
V.
7219.
Character
Methyl
anthranilate
24.87"
Total
volatile
esters
2.0342'

*
p
=
0.05
**
p
=
0.01
.­
­
With
this
in
mind,
genotypic
formulae.
were
'esta6­
lished
for
the
parental
cultivars
in
terms
of
MA
and
TYE.
A
collation
of
the
information
from
the
Chi­
square
t.
ests
(Tables
1
and
21,
the
statistical
analysis
(Tables
5
and
61,
and
the
ancest.
ry
of
the
parental
cultivars
(16)
.

provided
the
basis
for
this.
With
consideration
to
MA
in
family
V.
7218,
it
is
suggested
that
V.
54077
(MA
=
0.14
ppm)
is
of
the
genotype
MmAaFf,
while
B.
S.
5563
(MA
=
nil)
is
MM.
4aff
or
any
similar
genotype
cont.
aining
a
homozygous
dominant,
a
heterozygous,
and
a
recessive
.
.

locus.
For
family
V.
7219,
V.
54077
is
as
previously
noted,
while
V.
50061
(MA
=
nil)
is
rnrnaaff.
If
one
examines
the
ancestries
of
these
cultivars,
these
genotypic
formulae
can
be
qualified.
The
cultivar
V­
54O'ii
is
a
cross
between
Concord,
probably
a
pure
t'.
labrusca
cultivar
(and
likely
a
homozygote
in
terms
of
.

MA
and
TVE),
and
de
Chaunac,
whose
background
is
V.
labrusca­
free
for
OUT
generations,
(and
is
likely
ti,
be
a
homozygous
recessive).
An
F,
progeny
which
is
heterozy­
gous
at
all
three
loci
is
thus
probable.
For
B.
S.
5563,
the
parents
are
both
French
hybrids,
but
the
pollen
parent,
B.
S.
34­
45,
has
the
V.
lhbruscana
cuhivar
Othello
as
a
1
~~
~.
~~~
~~
~
~­
.~
.
.
~
~~
~
~
"
~~

­~
~

18
­
METHYL
ANTHRANILATE
grandparent
(8
).
Some
dominant
loci
for
this
cultivar
are
very
possible
in
light
of
this
informatinn,
despite
the
fact
that
i
t
contains
no
measurable
M
A
.
The
cultivar
V.
50061
does
have
some
V.
labrusca
in
its
ancest.
ry,
but
no
high
MA
o
r
TVE
seedlings
were
observed
in
the
family
from
which
it
was
selected
(Fuleki.
personal
communication),
so
a
completely
recessive
genotype
is
justifiahle
when
considering
t.
his
fact.
The
genotypic
formulae
are
also
consistent
with
the
segregation
ratios
obt.
ained
(Tables
1
and
2
)
and
the
information
derived
from
Tables
5
and
6.
Using
this
same
logic.
i
t
is
suggest.
ed
that,
for
TVE
in
family
V.
7218,
V.
54077
(TVE
=
21
ppm)
is
of
the
genotype
VuEe,
while
B.
S.
5563
(TVE
=
3
ppm)
is
Vuee
o
r
w
E
c
.
For
V.
7219,
V.
54077
would
b,
e
the
same
as
just
indicated,
while
V.
50061
(WE
=
1
ppm)
would
have
a
similar
genotype
to
B.
S.
5563.
The
significant
F­
value
in
Table
6
suggests
that
these
two
pollen
parents
may
be
genotypically
different
to
some
degree.
Correlation
between
these
and
other
characters:
Regression
analysis
indicated
that
concentrations
of
MA
and
TVE
were
not
correlated
in
any
way
(data
not
shown).
A
significant
r?
value
of
0.6780
for
family
V.
7219
could
be
explained
by
outliers
in
the
data.
Neither
character
could
be
correlated
with
"Brix
either,
al­
though
the
misconception
is
still
present
that
cultivars
of
labrusca
flavor
character
are
consistently
low­
OBrix,
high­
acid
t.
ypes.
A
correlation
could
also
not
be
found
between
either
flavor
component
.and
vigor
of
winter
hardiness
(data
not
shown);
this
too
has
oft.
en
been
the
concern
of
many
grape
breeders,
who
have
felt.
that
selections
with
labrusca
character
are
the
most
vigorous
and
most
winter­
hardy.

CONCLUSIO~
S
Our
results
have
shown
that
the
labrusca
flavor
character
is
a
complex
one
genetically,
involving
five
dominant
complementary
genes
plus
modifiers.
It
must
be
pointed
out
that
this
is
still
an
oversimplification,
since
many
other
compounds
are
likely
responsible
for
this
flaklor
character.
From
a
plant.
breeding
strindpoint,
it
is
a
difficult
trait
to
breed
against.
Knowledge
of
the
ancestry
of
the
potential
parental
cultivars,
coupled
with
efficient
selection
technigues
(such
as
the
utilization
of
the
VGFI)
appear
to
be
effective
weapons
in
combatting
this
problem.
This
study
was
enrisioned
as
being
a
preliminary
invest.
igation
which
could
provide
a
base
for
further
st.
udy
on
the
genetics
of
labrusca
flavor.
The
study
was
1imit.
ed
by
the
lack
of
availability
of
entirely
adequat.
e
genetic
mat.
eria1.
Future
researchers
in
this
area
would
do
well
to
hybridize
cultivars
extremely
rich
in
both
MA
and
TVE
with
V.
vinifera
cultivars,
which
contain
none
of
the
former
compound
and
little
of
the
latter.
The
segregation
ratios
obtained
from
such
crosses
should
yield
valuable
information.
The
examination
of
prog­
enies
from
selfed
high­
VGFI
cultivars
should
also
be
of
great
interest.
Finally,
much
work
is
still
required
on
the
biosynthe­
sis
of
MA,
TVE,
and
other
flavor
compounds
in
grapes.
Nnt
u
n
t
i
l
more
details
are
provided
on
the
biochemical
aspects
of
these
flavor
constituents
w
i
l
l
the
genetics
of
grape
flavors
be
comp1et.
ely
elucidated.

LITERATURE
ClTED
I
.
Rradt,
0.
A.
The
grape
in
Ontario.
Ont.
Min.
Agrir.
Food
2.
Casimir.
D.
.J..
d:
C.
hhyer.
and
L.
R.
Mattick.
Fluorometric
determination
of
methyl
anthranilate
in
C~
ncord
grape
juice.
J.
A.
O.
A.
C.
59:
269­
72
(1976).
Puhl.
467
(1972).

3.
Clore.
1%.
J..
A.
M.
Neuberl,
G
.
H.
tarter,
D.
W.
Ingalsbe,
and
V.
P.
Brummnnd.
composition
of
Washington­
produced
Con.
cord
grapes
and
juices.
Wash.
Agric.
Expt.
Sta.
Tech.
Bull.
48
(19651.

4.
Fuleki.
T.
Changes
in
the
chemical
composition
of
Concord
grapes
grown
in
Ontario
during
ripening
in
the
19'70
season.
Can.
J.
Plant
Sci.
52:
863­
7
(1972).

5.
Fuleki.
T.
Methyl
anthranilat.
e,
volatile
esters,
and
antho.
cvanin
content
of
grape
varieties­
grown
in
Ontario.
Can.
Hort,
Council:
Report
of
the
Cmt.
e.
on
Hort
Res.
153
(19741.

6.
Fuleki.
T.
A
new
objective
index
for
the
earl?
screening
of
grape
seedlings
based
on
flavor
character.
Can.
Hort.
Council:
Report
of
the
Cmte.
on
Hort.
Res.
179­
80
(1975).
7.
Fuleki.
T.
Vineland
Grape
Flavor
index
­
a
new
objective
measure
for
the
early
screening
of
grape
seedlings
based
on
flavor
character.
Can.
Hort.
Council:
Report
of
the
Cmte.
on
Hort.
Res.
1E2­
3
(1976).
.

8.
Galet.
P.
Precis
d'Ampelographie
Pratique.
Tome
H
I
.
De.

9.
Hedrick.
t'.
P.,
N.
0.
Booth.
0.
M.
Taylor.
R.
Wellington.
han,
hlonrpellier
(1976).
­

and
M.
J.
Dome?.
The
Grapes
of
New
York.
Fifteenth
Ann.
Report,
:

X.
T.
S.
Agric.
Expt.
Sta.,
Vol.
3,
Part
11.
.J.
B.
Lyon,
Albany.
N.
Y.!
(1908).

10.
Hill.
Lj.
T.
Colorimetric
determination
of
fatty
acids
and
11.
Holley.
R.
W.,
B.
Stoyla,
and
A.
D.
Hollev.
The
identifica­
'esters.
Ind.
Eng.
Chem.
(Anal.
Ed.)
1fX17­
9
(19461.

tion
of
some
volatile
constituents
of
Concord
grape
juice.
Food
Res.
2@
326­
31
(19.55).
12.
h'elson.
R.
R.,
T.
E.
Acree,
C.
Y.
Lee.
and
R.
M.
Butts.
hfethyl
anthranilate
as
an
aroma
constituent
in
American
wine.
J.
Food
Sci.
4257­
9
(19i7).
13.
Xeudoerffer,
T.
S.,
S.
Sandler,
E.
Zubeckis,
and
M.
D.
Smith.
Detection
of
an
undesirable
anomaly
in
Concord
grape
by
gas
chromatography.
J.
Agric.
Food
Chem.
13584­
8
(1965).

14.
Ontario
Crop
Protection
Committee.
Fruit
Production
Rec­
ommendations:
Ont.
Min.
Agric.
and
Food
Publ.
360
(1980).
15.
.Power,
F.
B..
and
V.
K.
Chesnut.
Examination
of
authentic
grape
juices
for
methyl
anthrani1at.
e.
J.
Agric.
Res.
23:
47­
53
(1923).
16.
Reynolds.
A.
G.
The
inheritance
of
methyl
anthranilate
and
total
volatile
esters
in
Vitis
spp.
MSc
Thesis.
Univ.
of
Guelph,
Guelph.
Ontario
(1980).

13.
Robinson.
W.
B.,
N.
J.
Shaulis,
and
C.
S.
Pederson.
Ripening
studies
ofgrapes
grown
in
1948
for
juice
manufacture.
Fruit
Prod.
J.
and
Amer.
Food
Manuf.
29(
2):
36­
7
(19491..

18.
Sale.
J.
W..
and
J.
B.
N'ilson.
Distribution
of
volatile
flavor
in
grapes
and
grape
juices.
J.
Agric.
Res.
33:
301­
10
(1926).
19.
Stern,
D.
J.,
A.
Lee,
W
.
H.
McFadden,
and
K.
L.
Stevens.
Volatiles
from
grapes
­
identification
of
volatiles
from
Concord
essence.
J.
Agric.
Food
Chem.
151100­
3
(19671.
20.
Stevens.
K.
L.,
A.
Lee,
W.
H.
hlcFadden.
and
X.
Teranishi.
yolatiles
from
grapes.
I.
Some
volatiles
from
Concord
essence.
J.
Food
Sci.
3
0
1106­
5
(1965).

Am.
J.
Enol.
'Vitic..
Vol.
33,
No.
1,
1982
METHYL
ANTHRANILATE
­
19
descendance
de
Vilis
tini/
rra.
Colloque
C.
N.
R.
S.
'Facteurs
et
Regulation
de
la
Maturiti.
des
Fruits.":
335­
9
11974).
.
25.
Wagner.
R.,
N.
Dirninger,
V.
Fuchs,
and
A.
Rronner.
Study
of
the
intervariet.
al
differences
in
the
concentration
of
volatile
constituents
(linalool
and
geraniol)
in
the
arnrna
of
the
grape.
Interest
of
such
analyses
for
the
appreciation
of
the
quality
of
the
harvest.
Int.
Symp.
Qual.
Vintage:
137­
42
(1977).
cw2
26.
Wagner.
R.,
N.
Dirninger,
V.
Fuchs.
and
A.
Bonner.
Premiers
resultats
concernant
]'etude
gPn6tique
de
ronstifuants
volatils
im­
portants
de
l'irome
des
raisins­
dans
deux
descendances
de
Vitis
L!
inift=
ra
L.
I
P
Symp.
CbnPtique
AmClior.
Vigne:
419­
34
(1978).

Am.
J.
Enol.
Vitic.,
Vol.
33,
No.
1,
1982
1024.
:E
programmed
to
165'C
at
2"
C/
min,
with
a
helium
gas
flow
of
5ml/
min
withoutsplitting.
%

5
.:
and
the
total
ion
trace
wasrecorded
at
20
eV,
whilst
the
mass
spectra
were
run
at
70
eV
and
50
p
~,
s
tea
stair
The
technique5­
6
did
not
employ
an
internal
standard.
Sel
del=
2.4.
Identification
of
components
const
The
compoundswere
identified
by
comparison
of
their
g.
c.
retentim
times
and
mas
spectra
with
I
more
those
of
commercial
samples
and
with
those
reported
by
other
authors.*+*
prese
i.
.
injector
valve
temperature
was
220°
C.
The
separator
and
m.
s.
s~
nce
were
maintained
at
2
6
0
%~
1
four
I
I
.__

..
..
­
....

4­

..
i
3.
Results
and
discussion
The
compounds
identifipd
are
listed
in
Table
I
,
together
with
the
number
of
the
chromatographic
peak
in
which
each
was
found.
The
relative
amounts
in
the
headspms
of
new
and
aged
wine
can
Table
1.
Compounds
identified
in
red
wine
heafspace
Identification
Peak
no.
Formula
Component
Rcfernce
r.
t.
ms.

i
CH40
Methanol
1
CZH~
O
Ethanol
1
X
C3H
a
0
n­
Propanol
I
CiHpO
i­
Propanol
I
CCHaOz
E
t
h
y
l
a
c
e
t
a
t
e
1
X
CsHmOz
n
­P
r
o
p
y
l
a
c
e
t
a
t
e
1
x
.
CSHIOOZ
­i
­P
r
o
p
y
l
a
c
e
t
a
t
e
1
X
C6H1202
Ethyl
n­
butyrate
1
X
C:
H1402
Ethyl
i­
valerate
.
I
X
CeHs
Indene
.
1
.x
X
3
C6HltO
n­
Hexanal
1­
X
C7H1402
.
i­
Pentyl
acetate
I
X
4
CsHlzO
3­
Methyl­
I­
butanol
I
­
x
X
CsHlzO
2­
Methyl­
I­
butanol
1
X
5
CaHlsOz
.
Ethyl
caproate
­.
1
X
X
X
:­
2
C4H
100
3"
ethyl­
I­
propanol
1
­
x
.

X
X
X
X
X
X
X
X
X
6
CeHlsOz
n­
Hexyl
acetate
1
8
C5Hl003
E
t
h
y
l
l
a
c
t
a
t
e
I
9
CsH14O
I
Hexanol
..
I
X
..
7
C;
HloOz
t­<
Ethoxymethyl)
furaa
X
C6H
1
2
0
trans­
3­
Hexeno
I
X
X
CsH
1
2
0
I1
cis­
3­
Hexenol
..
I
CaHtsO
n­
Nonanal
13
3
X
X
16
CloHzoOz
Ethyl
octanate
­
I
CTHIOO
17
Benzaldehyde
I
x
.
X
CIOHZOO
Decunal
3
19
I
X
X
CIIHZZOZ
Ethylpelargonate
.
.

22
I
CiH803
fihyI­
2­
furoare
X
23
1
GHnO
Aremphenone
.
.

24
1
CeH1oOz
Ethyl
Cenzoate
1
.
25
C1zH~
402
Ethyl
caprate
I
­x
26
CsH
1
4
0
4
Diethyl
sureinate
1
X
CI
nHs
Napthalene
29
I
32
T~
ethyldihydronaphthafene
CloH1202
EtIplpheny7aretate.
X
34
I
­
.x
ClnH~
rOz
IPhene~
hyl
acefure
X
.
35
1
­
x
CnHsNOz
Aicrhyl
attthranilate
36
4
C14HZeO:
Ethyl
lawme
37
1
CsHlnO
.'b~
hcny&?
thanol
.
.
1
X
X
X
X
1
.

.)8
CloH1sO
Linalool
x
.
X
x
X
X
X
X
X
x
X
X
28
.
C13H16
X
X
The
componenls
found
for
the
first
time
in
wine
by
headspace
tecknique.
are
given
in
italia.
The
components
found
for
the
fifst
time
in
wine.
are
in
bold.
I.
&<&
li
a
d
R.
\'&
ni
Sithout
splitting.
Thc
maintained
at
26O.
c
n
at
70
eV
and
50
FA,

md
mass
spectra
with
'
the
chromatographic
:w
and
aged
wine
can
entitication
?x
sd'
x
X
X
x
X
X
X
X
X
X
X
.x
­x
X
'
X
X
x
X
X
x
'
X
X
,x
..

.
'
,
x
x
msoc
Rdesco
1973
I
i
­2
!
i
i
Rirbesco
1966
"J
Figure
2.
G.
c.
traces
of.
1973
and
1966
wines
headspaces.
Among
&e
constituents
identified
I
,I
.6­
trimethyl­
l
,2­
dihydronaphthalene
(dehydroionene)
and
2­(
ethoxymethyl)
furan,
were
found
for
the
first
time
in
wine
aroma.
1,1,6­
TrimethyI­
l,
2­
dihydro­
naphthalene
had
been
previously
described
in
strawberry,
l2
peach
13
and
tobacco
leaves,
14
and
in
.

rum,
15
passion
fruit,
I6
and
peach
fruit.
';
Since
we
were
unabie
to
detect
it
in
Swiss
Garnay
and
Pinot
Noir
red
wines,
18
its
presence
in
the
aroma
together
with
other
leaf
constituents
such
as
cis­
and
frans­
3­
hexenols,
could
be
explained
by
differences
in
some
operations
of
wine
making.
The
ethyl
ether,
2­(
ethoxymethyl)
furan:
already
known
in
white
bread
ar0rna1~.
20.
has
a
fruity,
aromatic
and
slightly
pungent
smell.
The
confinement
of
this
compound
io
aged
wine
suggests
that
ageing
processes
involving
ethanol
reactions
occur
not
only
by
esterification
of
carboxylic
acids,
as
other
authors
have
observed6J1*
21.22
hut
also
by
etherification
of
Maillard­
formed
alcohols,
such
as
2­
furfuryl
alcohol.
Further
work
is
in
progress
on
the
genesis
of
these
compounds.
i
f
4.
Conclusion
i
Headspace
analysis,
in
which
one
measures
only
the
volatile
constituents
present
in
the
atmosphere
of
the
glass
in
their
natural
equilibrium
conditions,
could
become
the
technique
of
choice
for
1
following
the
evolution
of
wine
"bouquet"
and
thus
supplement
the
information
on
the
total
aroma
obtained
by
standard
solven't
extraction
techniques.
i
Acknowledgements
1
We
thank
Professor
G.
Montedoro
for
useful
discussions,
and
Dr
G.
Philippossian
for
the
synthesis
of
ethyl
furfuryl
ether.
.
j
References
1
1.
2.
3.
4.
.5.
6.
7.
8.
9.
10.

,
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
c;
TAB
C.
USlFDA
REGISTRATION
OF
METHYL.
ANTHRANILATE
AS
A
GENERALLY
RECOGNIZED
AS
SAFE
(GRAS)
MATERIAL.

TABLE
OF
CONTENTS
1.
21
CFR
1182.60
­
Synthetic
flavoring
substances
andadjuvants;

2.
21
CFR
184.1021
­
Benzoic
Acid
Methyl
anthranilate
(methyl­
2­
aminobenzoate)
8
182.40
21
CFR
ChA
(4­
1­
91
Edition)

§
182.40
Natural
extractives
(solvent­
free)
used
in
conjunction
with
spices,
seasaninp,

Natural
extractives
(solvent­
free)
used
in
conlunction
with
spices.
seasonings.
and
flavorings
that
are
generaflyrecognized,
as
safe
for
their
intended
use,
within
the
meaning
of
section
409
of
the
Act,
are
as
follows:
and
flavorings.

­~

canmon
name
Botankal
m
of
plant
saum
W
t
kemel
(persic
oil)
PlUOUS
MneniBC8
L
Peach
kernel
(persic
air),
.........­........".­...
".
f"
penica
%b.
81
%
Peanut
stearine
.­
...............................
..~."...­..~
Arachis
hvpooa~
L
Persic
o
i
l
(s
e
e
apicot
kwnel
and
peach
kemdL".
~uince
seed
........................................
......"­
cvdonia
OMOnpa
Mil&.

E42
F
R
14640,
Mar.
15.1977.
as
amended
at
47
FR
47375.
Oct.
26,19821
I182.50
Certain
other
spices,
seasoninge.
essential
oils,
oleoresins,
and
natural
extracts.
Certain
other
spices,
seasonings,
essential
oils,
oleoresins.
and
natural
extract8
that
are
generally
recognized
8s
d
e
for
their
intended
use.
within
the
meaning
of
section
409
of
the
Act,
are
as
follows:

B
182.60
Synthetic
flavoring
substances
Synthetic
`flavoring
substances
and
adjuvants
that
are
.generally
recog­
nized
as
safe
for
their
intended
use,
within
the
meaning
of
section
409
of
the
Act,
are
as
follows:
Acetaldehyde
(ethanal).
Acetoin
(acetyl
methylcarbinol).
Anethole
(parapropenyl
anhole).
Benzaldehyde
(benzoic
aldehyde).
N­
Butyric
acid
(butanoic
acidl.
d­
or
I­
Carvone
tcarvol).
Cinnamaldehyde
(cinnsmtc
aldehyde).
Citral
(2.6­
dimethyloctadien­
2.6­
aZ­
8.
gem
Decanal
(N­
decylaldehyde,
capraldehyde.
nial.
nerd).

c­
10).
capric
aldehyde,
caprinaidehyde.
aldehyde
and
adjuvants.

Ethyl
acetate.
Ethyl
butyrate.
3­
Methyl­
3­
phenyl
glycfdic
acid
ethyl
ester
(ethyl­
methyl­
Dhenul­
fclucidate.
so­
called
strawberry
aldkhyde.
6
1
6
aldehyde).
Ethyl
vanillin.
Geraniol
(3.7­
dfrnethyl­
2,6
and
3,6­
octadien­
1­
02).
Geranyl
acetate
(geraniol
acetate)..
Limonene
(d­,
I­,
and
dl­).
Linalool
(linalol,
3.7­
dimethyl­
1.6­
octadien­
3­
02).
Linalyl
acetate
(bergamol)..
Methyl
anthranilate
tmethyl­
2­
aminoben­

Piperonal
(3.4­
methylenedioxy­
be&
d*

Vanfwn.
E42
FR
14640.
Mar.
15.
1977,
as
amended
at
43
FR
47724.
Oct.
17.1978:
44
FR
3963.
Jan.

51907,
Nov.
15.
1983:
54
FR
7402.
Feb.
21,
19,
1979:
44
F'R
20656.
Apr.
6.
1979;
48
F'R
19891
ff
182.70.
Substances
migrating
from,
cotton
and
cotton
fabrics'
used
in
dry
food
packaging.
Substances
migrating
to
foodfrom
cotton
and
cotton
fabrics
used
in
drY
food
packaging
that
are
generally
r
y
ognized
as
safe
for
their
intended
use,
­within
the
meaning
of
section
409
of
the
Act.
are
as
follows:
Beef
tallow.
Carboxymethylcellulose.
Coconut
oil,
refined.
Cornstarch.
Gelatin.
Japan
wax.
Lard.
Lard
oil.
Oleic
acid.
mate).

..
hyde,
heliotropin).

404
Food
a
Pearrut
c
Potato
s
sodium
sodium
I
sodium
I
sodium
1
SOY­
Talc
Tallow
(1
T
~O
W
n
Tapioca
8
Tetrssod
me!&
st
zinc
m
a
E42
FR
1
43
FR
1
May
IS,
FR
27801
Nov.
7.
1'
FR
51909
52445,
5;
May
7.1s
f4
18290
.
fram
s
u
m
in
food
paper
a
recogniz
use,
wit1
of
the
A
Alum
<do
12SAlum
moniun
Aluminlu
Aluminm
Cellulose
CaSeil3.

Diatomac
cornstarc
Ethyl
cell
Ethyl
van
Glycerin.
oleic
add
Potsssiun
Silicon
d
i
t
sodiuni
al
sodium
cl
Sodium
hj
Sodium
h:
sodium
PI
Sodium
si
sodium
sc
Sodium
tr
SorbitoL
Soy
protei
Starch
ac
Starch.
pr
starch.
ut
Talc.

Zinc
hydn
Vanillin
Zinc
sulfa
E42
FR
141
0
184.1011
tc)
The
ingredient
is
used
as
a
flavor­
ing
agent
as
definedin
H
170.3(
0)(
12)
of
this
chapter:
leavening
agent
as
de­
fined
in
8
170.3(
0)(
17)
of
this
chapter:
and
pHcontrol
agent
as
defined
in
0
170.3(
0)(
23)
of
this
chapter.
(d)
The
ingredient
is
used
in
foods
at
levels
not
to
exceed
current
good
man­
ufacturing
practice
in
accordance
with
0
184.
UbXl).
Current
good
manufac­
turing
practice
results
in
maximum
levels,
as
served,
of
0.05
percent
for
baked
goods
as
defined
in
g
170.3tnM1)
of
this
chapter;
0.005
percent
for
non­
alcoholicbeverages
as
defined
in
0
170.3(
n)(
3)
of
this
chapter;
5.0
per­
cent
for
condiments
and
relishes
as
de­
fined
in
§
170.3tn)(
8)
of
this
chapter:
0.45
percent
for
dairy
product
analogs
as
defined
in
0
170,3tn)(
lO)
of
this
chapter;
0.3
percent
for
fats
and
oil
as
defined
in
H
170.3tn1(
12)
of
this
chap­
ter;
0.0004
percent
for
frozen
dairy
desserts
as
defined
in
f
170.3tn)(
20)
of
this
chapter:
0.55
percent
for
gelatin
and
puddings
as
defined
in
8
170.3tn)(
22)
of
this
chapter;
0.1
per­
cent
for
gravies
as
defined
in
§
170.3tn)(
24)
of
this
chapter;
0.3
per­
cent
for
meat
products
as
defined
in
P
170.3tn)(
29)
of
this
chapter:
1.3
per­
cent
for
snackfoods
as
defined
in
§
170.3(
n)(
37)
of
this
chapter;
and
0.02
percent
or
less
for
all
other
food
cate­
gories.
(e)
Prior
sanctions
for
adipicacid
different­
from
the
uses
established
in
.this
section
do
not
exist
or
have
been
waived.
E47
FR
27810.
June
25.19821
I
184,1011
Alginicacid.
(a)
Alginic
acid
is
a
colloidal,
hydro­
philicpolysaccharide
obtained
from
certain
brown
algae
by
alkaline
extrac­
tion.
(b)
The
ingredient
meets
the
specifi­
cations
of
the
Food
Chemicals
Codex,,
3d
Ed.
(1981).
p.
13.
which
is
incorpo­
rated
by
reference.
Copies
are
avail­
able
from
the
National.
Academy
Press,
2101
Constitution
­Ave.
NW.,
Washington,
DC20418,
oravailable
'
for
inspection
at
the
Office
of
the
Fed­
.
eral
Register,
1100
L
St.
NW.,
Wash­.
ington.
DC
20408.
­k
)
In
accordance
with
8
184.1(
b)(
2),
the
ingredient
is
used
in
food
only
within
the
followingspecific
limita.
'

tions:

soup
and
mlxea
?P
5
170.3(
n)
(40)
Of
this
chapter.

(dl
Prior
sanctions
for
this
ingredi­
ent
different
from,
the
use
established
in
this
section
do
not
existor
have
been
waived.

C47
F'R
41375,
Oct.
26.19821
I
184.1021
Benzoicacid.

zenecarboxylicacidtC7He0,).
occur­
ring
in
nature
in
free
and
combined
forms.
Among
the
foods
in
which
ben­
zoic
acid
occurs
naturally
are
cranber­
ries,
prunes,
plums,
cinnamon,
ripe
doves,
and
most
berries.
Benzoic
acid
is
manufactured
by
treating
molten
phthalic
anhydride
with
steam
in
the
presence
of
a
zinc
oxidecatalyst.
by
:

the
hydrolysis
of
benzotrichloride,
or
by
the
oxidation
of
toluene
with
nitric
acid
or
sodium
bichromate
or
with
air
in
the
presence
of
a
transition
met@
salt
catalyst.
(b)
The
ingredient
meets
the
specifi­
cations
of
the
"Food
Chemicals
Codex."
3dEd.
(
19811,
p.
35,
which
is
incorporated
by
reference.
Copies
may
be
obtained
from
the
National
Acade­
my
Press,
2101
Constitution.
Ave.
NW..
Washington.
DC20418,
or
mayex­
amined
at
the
Office
of
the
Federal
Register,
1­
100
L
St.
'
NW..
Washington,

tc)
The
ingredient
is
used
as
an
anti­
microbial
agent
as
defined
in
170.3(
0)(
2)
of
this
chapter,
and
as
a
flavoring
agent
and
adjuvant
as
de­
fined
in
0
170.3(
0)(
12)
of
this
chapter.
(d)
The
ingredient
is
used
in
food
at
levels
not
to
exceed
good
manufactur­
ing
practice.
Current
usage
results
in
a
maximum
level
of
0.1
percent
in
food.
tThe
Food
and
Drug
Administration
(a)
Benzoic
acid
is
the
chemical
ben­
.

DC
20408.

422
Food
has
cant1
woul
(e)
ent
lishe
that
ter,
142
F
49
m
§
184
CAS
(a
cal
I
ered
fatt
Pare
fern
tion
in
c
(b
cati
Cod
incc
be
c
Wa
ami
Reg
DC
(C
ing
0
1:
((

acc
not
Pra
ser
go(
Ch:
fin
0.0
fin
f
0
1
0
1
tin
5
1
pr'
t
h
f
i
I
O.(
f
iI
fo
er
OUS
pra
ar
­91
Edition)

cific
limita­

his
ingredi­
established
st
or
have
?mica1
ben­
3A
occur­
combined
.which
ben­
re
cranber­
.mon,
ripe
snzoic
acid
ng
molten
?a
m
in
the
ltalyst,
by
:1loride.
or
with
nitric
3r
.".­;$
h
air
..
which
is
'opies
may
.la1
Acade­
Ave.
NW.,
.lay
be
ex­
le'
Federal
uslhington,

a,
an
anti­
fined
in
.d
n
d
a
s
a
nt
as
de­
5
qhapter.
in1
food
at
.Wfactur­
3
S
~l
t
s
in
a
nibtration
­t.,
b
food.
Food
end
Qrug
Administration,
HHS
has
not
determined
whether
signifi­
cantly
different
conditions
of
use
would
be
GRAS).
(e)
Prior
sanctions
for
this
ingredi­
ent
different
from
those
uses
estab­
lished
in
this
section,
or
different
from
that
set
forth
in
part
181
of
this
chap­
ter,
do
not
exist
or
have
been
waived.

142
mZ
14653,
Mar.
15.
1977,
as
amended
at
49
FR
5610.
Feb.
14.19841
I181.1025
Caprylic
acid.
(a)
Caprylic
acid
CCH,(
CH,),
COOR,
CASReg.
No.
124­
07­
21
is
the
chemi­
cal
hame
for
octanoic
acid.
It
is
consid­
ered
to
be
a
short
or
medium
chain
fatty
acid.
It
occurs
normally
in
vari­
ous
foods
and
is
commercially
pre­
pared
by
oxidation
of
n­
octanol
or
by
fermentation
and
fractional
distilla­
tion
of
the
volatile
fatty
acids
present
in
coconut
oil.
(b)
The
ingredient
meets
the
specifi­
cations
of
the
"Food
Chemicals
Codex,"
3d
Ed.
(1981).
P.
207.
which
is
incorporated
by
reference.
Copies
may
be
obtained
from
the
National
Acade­
my
Press,
2101
Constitution
Ave.
NW..
Washington,
DC
20418,
or
may
be
ex­
amined
at
the
Office
of
the
Federal
Register,
1100
L
St.
NW.,
Wasfiington,
DC
20408.
<e)
The
ingredient
is
used
as
a
flavor­
ing
agent
and
adjuvant
as
defined
in
8
170.3(
03(
123
of
thischapter.
(dl
The
ingredient
is
used
in
foods
in
accordance
with
§
184­
1(
b)(
l),
at
levels
not
to
exceedgood
manufacturing
practice.
Current
good
manufacturing
practices
result
in
maximum
levels,
as
served,
of:
0.013
.Percent
for
baked
goods
as
defined
in
0
170.3(
n)(
l)
of
this
chapter;
0.04
percent
for
cheeses
as
de­
fined
in
§
170.3tn)(
5)
of
this
chapter;
0.005
percent
for
fats
and
oils
as
de­
fined
in
H
170.3(
11)(
12)
of
this
chapter,
for
frozen
dairy
desserts
as
defined
in
9
170.3tnX20)
of
this
chapter,
for
gela­
tins
and
puddings
as
defied
in
H
170.3(
n)(
22)
of
this
chapter.
for
meat
products
BS
defined
in
9
170.3tn)(
29)
of
this
chapter,
and
for
soft
candy
as
de­
fined
in
P
170.3tnX38)
of
this
chapter;
0.016
percent
for
snack
foods
as
de­
fined
in
5
170.3(
n)(
37)
of
this
chapter;
and
0.001
percent
or
less
for
all
other
(e)
Prior
sanctions
for
this
ingredi­
ent
different
from
the
uses
established
'
food
categories.
in
this
sewon
a0
noz
exist
or
nave
been
waived.

[43
FR
19843.
May
9,1978,
as
arnt
FR
5611,
Feb.
14.19841
I
184.1027
Mixed
carbohydrase
w
e
enzyme
product.
fa)
hlixed
carbohydrase
and
protease
enzyme
product
is
an
enzyme
tion
that.
includes
carbohydrase
and
the
culture
filtrate
resulting
xrom
a
pure
culture
fermentation
c
*
~
~~
~

1
pathogenic
strain
of
B.
lichen
(b)
The
ingredient
meets
the
specifi­
cations
of
the
Food
Chemici
3d
Ed.
(1981).
p.
107.
which
is
incorpo­
.
rated
by
reference.
Copies
are
avail­
able
from
the
National
Academy­
­

Press,
2101
Constitution
A"­
""

I
Washington,
DC
20418,
or
available
eral
Register,
1100
L
St.
NV
ington.
DC
20408.

the
ingredient
is
used
in
fooc'
1
limitation
other
than
currt
manufacturing
practice.
The
tion
of
this
ingredient
as
generally
rec­
ognized
as
safe
as
a
direct
human
food
ingredient
is
based
upon
the
following
current
good
manufacturi"
conditions
of
use:
(1)
The
ingredient
is
used
as
an
this
chapter,
to
hydrolyze
protelns
or
carbohydrates.
(21
The
ingredient
is
used
in
the,
fol­
lowing
foods
at
levels
not
to
exceed
current
good
manufacturing
­
­­
­
~

alcoholic
beverages,
as
defined
in
0
170.3(
n)(
2)
of
this
chapter,
candy.
nutritive
sweeteriers,
and
protein
hy­
drolyzates.

148
F
R
240,
Jan.
4.19833
8
184.3061
Lactic
acid.
(a)
Lactic
acid
(C3H603.
CAS
Reg.
Nos.:
DL
mixture,
598­
82­
3;
L­
isomer,
79­
33­
4;
0­
isomer,
10326­
41­
7).
the
chemical
2­
hydroxypropanoic
acid.
occurs
naturally
in
several
foods.
It
is
produced
commercia""
mentation
of
carbohydrates
glucose.
sucrose,
or
lactose,
or
by
a
protease
activity.
It
is
obtai:
­
J
I
for
inspection
at
the
Office
of
"

1
(c)
In
accordance
wnn
3
18%
1(
b)(
i),

­­­­*
'­
­
I
enzyme,
as
defined
in
§
170.­
I
­lL'­
­­
­~
e
..
TABLE
OF
CONTEWS
1
.
Fenaroli'sHandbook
of
FlavorIngredients,
Second
Edition,

2.
Perfume
Synthetics
and
Isolates
Volume
2
I
\

,

!
t
FENAROLI'S
HANDBOOK
FLAVOR
INGREDIENTS
Second
Edition
Volume
2
Edited,
Translated,
and
Revised
bY
THOMAS
E.
FURIA
d
NICOU)
BEUANCA
QVtl47p­
1
Palo
Alto.
Culifortzia
Published
by
Adapted
from
the
Italian
language
works
of
PROF.
DR.
GIOVANNI
FENAROLI
Director,
Center
for
Srudies
of
Aromatic
Substances
University
of
Milano.
Milano.
Italy
CRC
PRESS.
Inc.
18901
Cranwood
Parkway
Ckveland.
OhioU128
..
.
..

..

e"

.
'
,
I'
€46
Fenaroii'r
Handbook
of
Flavor
Ingredients
Stmture
Pbysical,
'chemial
characteristicsJ
APP=
iUKC
*=
Y
Moleculu
weight
Melting
point
Wii
point
Congealing
point
SpeciBc
gravity
Refractive
index
Cresol
mtmt
Solubility
Organokpiic
char­
acteristics
SyMwsis
Natural
OCCII~~
CLKC
Regulatory
status
P
c
m
~l
methyl
ether
pMethoxy
toluene
Methyl
pcrcsol
Methyl
ptolyl
ether
Methyl
2­
aminoknzarre
Methyl
u­
aminoknzoate
(?"
CH,
.

CH,

Colorless
liquid
122.17
0.96do.
970
st
i5­/
25'C2
1.5100­
1.5130
at
20'C2
Not
more
than
0.5
0iz
Fungent
odor
suggestive
of
ylang­
ylang
By
methylation
of
pcresol
Reported
found
in
the
oils
of
ylang­
ylang
cananga,
and
0
t
h
~~
­
.

Non­
alcoholic
beverages
1.7
ppm
Ice
crram.
ices
etc.
2.7
pprn
kked
goods
7.6
Ppm
jehtins
and
puddings
0.
W.
o
ppm
hdiments
2.0
ppm
­A
121.1164;
FEMA
No.
2681
h
M
i
Y
4.8
pprn
iYrUPS
8.0
pprn
REFERENCES
,

For
References
1­
5,
Jee
end.
of
Part
111.
6.
Mattick
et
d..
1.
Agric.
fmd
Chrm..
4.
331.
1963.
7.
Roger,
Food
Technof..
6.309.
1961.
v
t
Colorless
to
pale­
yellow
Itquid
uith
bluish
98O,
mln'
151.17
24­
25
C
i32'Cat
I?
mm
Hg
.
23.8.
C
(24
C
)

t.
161­
1.169
at
IS
.25
C:
'
1.16.$
0
at
25
6
1.5820­
1.5810
at
20
C:
'
1.5802
at
I5
C
fluorescence
Characteristic
orange­
flower
odor.
and
dlghtly
brtter.
pungent
taste
By
heating
anthranilic
acid
and
meth>
i
and
subsequent
distiliatlon
alcohd
in
the
preKnce
of
sulfzric
s::
d
Reported
found
inxw­
al
essential
oil*:
neroli.
orange.
bergamot.
lemon.
man­
darin.
jasmine.
tuberose,
gardenla.
champaca.
ylang­
ylang.
and
o
t
h
m
:
also
in
the
juice
and
oil
of
I
:1t5
iabrusrrr*~
'

Non­
alcoholic
beverages
16
ppm
Alcoholic
beverapcs
lce
cream.
ices.
etc.
0.20
ppm
Candy
II
ppm
Baked
goods
56
pprn
Selatins
and
puddings
13
ppm
IO
ppm
Cheuinggum
2.200
ppm
FDA
GRAS:
FEMA
No.
2682
.
*

I
'
.
..

a
I
and
IsoZates
BY
PAUL
2.
BEDOUK14W,
PH.
D.
Conlpagnic
Parenlo,
Inc.

D.
VAN
XOSTRAND
­
COMPANY,
INC.
*NEW
VORK
,
p
NEW
TORK
I).
Van
Nodrand
Company,
Inc.,
250
Fourth
Avenut,
New
Yort
3
TORONTO
D.
Van
Sostrand
Company
(Canada),
Ltd.,
a8
Bloor
Street,
Toronto
.3f~
crnillan'&
Company,
Ltd.,
St.
Martin's
Street,
London,
W.
C.
2
LOXWN
ANTHRANILATES
Methyl
Anthranilate
N­
Methyl
Methyl
Anthranilate,

CfJHfrO2N
3101.
Weight
151.08
Dimethyl
Anthranilate
C*
II,,
O~
X
Siol.
Weight
165.09
Although
Fritzsche's
discovery
of
anthranilic
acid
dates
hack
to
1841,
the
importance
of
its
esters,
especially
the
methyl
ester,
was
not
r
e
a
l
h
a
until
1899
dien
Walbaum
noted
its
occurrence
in
neroli
oil.
Methyl
anthra­
nilate
has
since
been
found
iu
.a
number
of
other
essential
oils
and
bas
attained
!onsiderableimportance
as
a
perfumery
material.
Although
it
possesses
a
powerful
and
pleasant
odor,
the
use
of
anthraniiic
acid
and
its
esters
in
perfumes
is
limited,
however,
because
the
amino
group
reacts
most
readily
with
various
aldehydes,
giving
highly
colored
Schiff
bases
which
are
undesirable.
Large
amounts
of
methyl,
anthranilate­
or
other
esters
a
n
n
o
t
therefore
'
b
e
used
in
most
cases,
as
for
example
in
cosmetics
and
soaps
where
discoloration
is
a
serious
problem.
Derivatives
of
anthranilates
where
the
amino
group
ia
bound­
say,
with
an
acetyl
group­
are
usetes
for
perfumery
purposes,
since
they
are
either
extremely
weak
in
odor
or
odorless.
Large
quantities
of
methyl
anthranilate
are
manufactured
annually
for
the­
flavor
industry,
most
popular
grape
sodas
depending
mainly
upon
­methyl
anthranilate
for
the
characteristic
grape
flavor.

Occurrence.­
XethyI
anthranilate
has
been
found
in
a
nunlber
of
essential
oils,
being
first
rrported
in
neroli
oil
by
Walbaum.
1
This
oil
is
obtained
from
the
flowers
of
the
bitter
orange
tree
(Citrus
biguradiu
R.).
It
has
also
been
identified
in
the
oil
from
jasmine
.flowers
cultivated
in
France.
2
Practically
all
jasmine
oils
from
flowers
cultivated
in
other
countries
contain
SIWIII
quantifies
of
methyl
anthranilate.
Other
source
include
:
f
Mol.
Weight
165.09
Oil
of
rcrcir
Bowers
(Robiniu
psc*
doocacia
L.)
8
Oil
of
tuberoe
(Polyunlhcr
tutcrotu­
L.)
'
oil
of
nalfbov;
en
(Chciranfhw
cheiri
L.)@
oil
of
gardenia
dowen
*
Oil
from
the
leaves
of
the
bergamot
tree'
Orange
oil
'
3fetlq­
l
antbranilat;
has
also
been
found
in
grape
juice.)
X­
methyl
methyl
antbranilate
is
known
to
occur
in
mandarin
mendah
leaf
oil,
ll
oil
of
Koenlfctia
efhclia,"
oil
of
rue,!!
and
in
the
oil
of
hyacinthflowers
(Byacinlhtcs
oricnfulis
L.)."

Preparation
of
Anthranilic
Acid.­
Anthranilic
acid
was
discovered
in
the
course
of
studies
on
indigo.
In
1841,
Fritzsche
treated
indigo
with
concentrated
solution
of
potassium
hydroxide
at
elevated
temperatures
and
obtained
a
greenish­
yellow
solution.
This
solution
on
acidibcation
gave
a
dark
brown
precipitate
which
was
later
identified
as
chrysanilic
wid.
The
latter
decomposed
on
warming
with
mineral
a&,
and
one
of
the
products
crystallized­
out.
This
product,
which
remairied
unknown
to
the
perfume
industry
for
more
than
fifty
years,
was
anthranilic
acid.
Nethyl
anthranilate
was
prepared
synthetically
shortly
after
its
dis­
covery
in
various
essential
oils.
The
literature
on
methods
of
preparing
anthranilic
acid
is
qnik
volum­
inous
and
a
fd
discussion
of
the
various
rnethocb
cannot
be
undertaken
here.
The
acid
has
been
prepared
from
various
bewnt
derivatives
having
an
alkyl
and
nitro
grouping
in
the
ortho
position.
For
example,
it
can
be
obtained
by
treating
ortho­
nitro
benzyl
alcohol
with
sodium
hFdroxide
solution,
1'
ortho­
nitro
benzoic
acid
with
zinc
and
hydrochloric
acid?
'
or
by
reducing
orthonitro
benzaldehgde
with
zinc
dust
and
alkaliJ8
­
An
unusual
q­
ntbesis
of
anthranilic
acid
consists
of
treating
ortho­
nitro
"

8
Elre,
Ckm.
2rg.
34,
814
(1910).
4
El­,
Bcr.
36,
1465­
(1903).
6
Kummcrt,
Ckm.
ttg.
35,
667
(1911).
*
Pmroor.
Bod.
d
i
m
.
form.
41,
489
(1902).
7
GuDii
.t%
Chcmut
and
mgki
60,
1995
(1m).
8
Pam.
Chcmuf
0­
d
Druggut
56,
462
and
2
2
(1MO).
0
Power,
J
.
Am.
Chtm.
Sor.
13.
375
(1921).
10~
Tdbtom,
J
.
pdt.
a
r
m
.
I?].
62,
136
(lOU0).
n"
anbot,
Coappt.
red.
US,
580
(IS=).
I*
Goddig
mud
Roberta,
J
.
C
h
m
.
SOC.
101,
316
(1915).
18
Shimme1
t
Ce.
Report,
October
(1901).
47.
14Hoejenbor
and
Coppenr,
Rev.
morquu
parfum.
rt
(1931),
588.

I
C
c
l
r
r
e
,
BUZZ
roc.
dim.
Pranm
131,
33.1161
(190%
1?
Belktain
and
Kublberg,
AU
163,
138
(1872).
I*
Frr~
ndlcr.
BUR
me.
d
i
m
.
Prance
[SI,
31,
450
(1904).
1sFlit+
e,
1.
p
0
L
­C
.
C
b
t
­2
3
,
67
(1841).
'
56
11
I
nilic
acid.
One
molecular
quantity
of
the
phthalimide
is
dissolved
in
three
molecular
quantities
of
aqueous
alkali
solution.
To
the
cooled
solution
are
added
two
molecular
proportiolls
of
sodium
hypochlorite
with
constant
agitation.
After
several
hours
the
oxidation
'is
brought
to
completion
by
heating
the
mixture
for
half
an
hoar.
The
mixture
is
then
acidified
and
tbt
precipitated
anthranilic
acid
subjected
to
purification.
The
yields
obtained
by
this
process
are
of
the
order
of
'IO
per
cent,=

of
anthranilic
acid
by
the
reduction
of
ortho­
nitrobenzoic
acid
with
sulfuric
acid
and
iron
o
t
copper
metal.
This
process,
however,
does
not
seen1
ta
haye
fannd
commercial
application.
.,
More
recently
a
patent
has
been
obtainedm
describing
the
production
I
­
AXTRRc
with
alkali.
The
re
common
method
Y
V
I
.
­I
50
PERFUYERF
sY1`
STIIETICS
A
S
D
ISOLATES
'

f
The
above
procedures
u
l
alcoholysis
do
not
work
well
with
tertiarr
i
slcohols
and
an
interesting
variation
is
emplopd
ill
the
preparation
of
such
esters
as
Iinalyl
anthranilate
{IX).
In
this
case,
linatyl
formate
is
i
sodium
alcoholate
of
linalool.
Thus,
1
mole
of
litlalyI
formate
(VIII),
1
1
mole
of
methyl
anthranilate
(IV),
0.05
mole
of
linalool,
and
0.05
atom
of
sodium
are
heated
for
several
boun,
and
the
methyl'
formate
formed
dlowd
to
distill
O~
T.~
'
The
followingreaction
takes
place.
1
reacted
withmethyl
anthranilate
in
the
presence
of
a
small
amount
of
:
i
IX
AU
of
the
above­
mentioned
derivatives
were
esters
02
the
carboxyl
group
in
anthtaniIic
acid.
The
other
active
group,
namely,
the
amino
group
of
anthranilic
acid,
is
equally
capabie
of
undergoing
reactions
and
giving
interesting
derivatives.
.
8­
Methyl
methyl
anthranilate,
which,
(LS
mentioned
previously,
wcum
in
se~
cral
essential
oih
can
be
prepared
by
treating
anthranilic
acid
(I)

Congding
Point
18%
1S.
C.
1cc.
I2.
C.
1O.
C.
60
PERFD'YERP
SrNTBETICS
AKD
ISOLATES
Chemical
Properties.­
Bn~
hranilic
acid
reacts
with
simpler
aldehydes
in
a
definite
manner.
Thus,
1
mole
of
formaldehyde
condenses
with
2
moles
of
anthranilic
acid."

XI
rY
XI1
The
amine
group
can
be
acetylated
by
treating
the
acid
with
acetic
anhydride.=­
Although
esters
of
anthranilic
acid
are
stable
to
heat,
anthranific
acid
itself
decomposes
to
aniline
and
carbon
dioxide
on
heating
to
210'
C.
a
It
h
interesting
to
note
that
in
nature
methyl
anthranilate
and
indole
are
very
often
found
together.
Skatole
(XIII)
can
be
oxidized
to
acetyl
anthranilic
acid
(SIV)
uith
potassium
permanganate.
N
XI11
I
I
XIV
....
7
.:
ao
H
e
6
and
Fieuelmnnn,
ARK
324,
119
(1902).
81
Ticdke,
Bcr.
42,
611
(1909).
32Kaufmann,
Bcr.
42,
3455
(1909).
as
Fritrrhe,
Inn.
39,
86
(1511).

as
Qtr.
Pat.
137,008,
Norembcr
29,
190?,
;I
8
,
Jackson,
Ber.
.14,885
(1881).
A3Th
/

........
.............
..
.
...
......
..
...
.....
..

..........
.........
­,
...".
..
.I
...~
Ifany
6UCh
interrelationships
haw
been
studied
in
detail
since
they
rre
of
considerable
importance­
in
the
spthesis
of
indigo
and
other
dpstuf!
r
I
1.68
8
1.76
6
5.11
S
~

­
The
anthranilates
are
shipped
in
glass
or
aluminum
containers.
Tb­
lined
containers
are
also
used
occssionally.
They
should
be
stored
away
from
light;
otherwise
they
undergo
considerable
disco~
oratioa
...
.....................
__
.:,.
..........
..
.......
.
.
­j
.
­
...
I...................
I
__
...
......
.......
.........
,
:>
.,_.­..._
~

......
:.,:
.........
..
..
p
t
y
TAB
E
U.
S.
DEPARTMENT
OF
COMMERCE
SCIENTIFIC
\
'k­
b'
LITERATURE
REVIEW
OF
ANTHRANILATES
IN
FLA?
7QR
USAGE.
VOL.
I.
fntrouduction
and
Summary
Tables
of
Data,
Bibliography.

TABLE
OF
CONTENTS
1.
2.
3.
4.
5
.
..

6
.
7.
8.
Toxicity
to
mouse,
rat
and
Guinea
pig.
Chemical
identity
and
physical
properties
Pharmacological
and
toxicological
ef€
ects
Natural
occurrance
Flavor
and
Extract
Manaufacturers'
Association
(FEMA)
and
Addendum
to
Table
IV­
2
Bibliography
Data
guide.
National
Acadamy
of
Science
(NAS)
use
levels.
Scientific
Literature
Review
of
Anthranilates
in
Flavor
Usage.
Volume
1.
.Introduction
and
Summary
Tables
of
Data,
Bibliography
flavor
and
Exltod
Manufactunrs'
Association
of
.the
Unitd
States,
Washington,
DC
fwd
and
Drug
Administration,
Washington,
DC
Nor
10
291112
SCIEMIFIC
LITERATURE
REVIEW
'
OF
AMMRANILATES
IN
FUVOR
USAGE
.
..

..
SCIWlFIC
LITERATURE
REVIM
OF
ANlliRANIUTES
IN
FLAVOR
=AGE
BIBLIOGRAPHY
bY
SCIEhTIFIC
LITERATURE
REVIEW
OF
A8TIiRAs
1
LATES
IN
FLAVOR
US
ACE
PIgC
1­
3
4­
8
sEc?
IcN
1
.

A.
S.
..

.,
,

9­
12
13­
17
SECTION
11.
SUBSTANCES
REVIEWED
Ncrrsriul
Designation
urd
Chemic81
Identity
.
(Table
11­
1)
Alphabetical
Cnus­
Reference
List
of
Nbms
and
Synonym­
(T
r
b
l
~.
11­
2)
Physical
Proporties
l.
fable
11­
5)
A.
E.

C.
18­
24
.

SECTICH
111.

A.

SECTIW
IV.

A.
1.
39­
42
4
5­
73
SECffON
V.
74­
79
SECTIQJ
VI.
60­
96
V
O
W
11.
COPIES
OP
.WICLES
CInD
IN
SIwI(
ART
SECTION
SEcTIOH
I
.A.

This
revieu
is
a
prssentatioa
of
det?
p
e
r
t
i
n
e
n
t
t
o
the
safety
evalua­

ti=
of
mthraailrtss
wed
as
flavor
ingredients,
fhe
14
substances
uere
selected
for
renew
Y
8
gmup
because
of
their
close
chemical
relationships
on
tho
asrumption
that
the)
would
f
o
l
l
w
s
i
d
l
u
metabolic
3rthwryr,
and
havo
sim5la.
r
physiological
effects
in
tho
mamnialian
organism.
The
list
of
sdrtmces
rovicrwtd
urs
compilerr
fma
sevtrlj
sources.
I
t
includes
flavoring
caapo\
pl&
tistad
by
tho
Food
urd
k
g
Admiairtratiacr
(FM)
in
,Ticlo
2:
of
the
Cod.
or'
Fed0181
Regulations
,.
md
rdrtaaces
clurifird
u
Gonorrlly
Recognirsd
u
Safe
(GRS)
by
the
Expert
Pmal
of
the
Flavcr
8ad
Extract
Mmufacturers'
k
s
o
d
a
t
i
o
n
(­
1
urd
recognAred
by
FpA;.
md
ury
additional
flavor
rdsturces
roportod
by.
rtsponden:
r
t
o
a
aatiaul
s
w
r
y
conducted
by
RUA
Bnd
the
Natian.
1
Acat3r.
r
of
Sciences/
riational
Resuucb
Council
!NAT;
lNNRC)
in
1970­
1971.
C;
uid.
linor
for
the
typos
and
ammt
o
f
data
t
o
be
gatherod
yet.
developed
.
.

in
consultrtioa
utth
the
EqorC
Puul
o
f
RClA
(4
5
1,
a
d
in
ucordutc6uitb
tho
ctftorir
pub:
irhed
by
tho
bod
Proteetian
Colrittw
o
f
.the
Natiaul
A
u
d
y
of
Sciences/
Nationrl
Res­
Cormcil
(WRVRC)
(%
1.

Ih8
bulk
of
the
infomtioa
on
biologic81
properties
YU
&t8ined
froll
a
soarch
of
the
sciearific'literrturi
frolr
3930
thm­
1977,
hy
articles,
revim
md
texts
wet.
well
Lnavn
urd
had
been
utilized
in
oulier
evalwtionr.
Using
t
h
i
chemical
aIyr
(u
wtll
u
the
c­
n
ae1#
.md
ryno­
nyrs
of
gach
r\
rbrtanw)
as
key
wor&,
ca
independent
orgmixatisn
urrid
out
t
h
i
s
search
by
locating
a
l
l
refe~
encer
t
o
litersturn
pertaining
t
o
phyriologiui,
yhWCOiOgiC8ls
toxicologfal,
metabolic
or
o
t
h
t
r
b
i
o
l
o
l
i
u
l
.
fniorvtion
or.
&use
substmces.
This
litemture
s
w
d
)
oxtended
back
t
o
1920
for'the
aeuch
p
u
r
a
+t
e
r
r
t
o
s
i
c
i
t
y
and
metrboliu.
hppmximteLv
40
nformci8
uom
10cated
in
this
reuth,
but
only
thwe
a
r
t
i
c
l
e
s
conttidq
drtr
relevant
t
o
safety
o~
alurrtlon
r
o
n
selected
for
inclwioa
in
this
=v
i
r
.
S
w
i
u
of
a
l
l
OC
tho
rrlovmt
articles
Y
e
n
then
w
r
i
t
t
e
n
urd
are
presented
in
Section
111.
A
.corrpleto
bibliogrtphy
o
f
a
l
l
references
lwsted,.
whuther
or
not
the
articles
were
usod,
1s
also
included
(Section
v),

1
Iho
informtion
included
on
the
natural
0ccurrcn;
e
of
:he
reviared
sd­
c:
uIces
i
n
food
*PI
provided
thxwgh
food
i
n
b
t
l
v
sources,
and
includes
I
I
W
~O
U
references
t
o
the
open
l
i
t
e
m
t
u
r
e
.
This
natura:
occurrence
infor­
s
t
i
o
n
is
pxcsanted
i
n
table
foi­
t
in
Section
:V.
A.
Physical
data
we=
a
l
s
o
obtainod
ftor
the
industry
as
v
e
l
1
as
v
a
t
L
o
u
coapendia.
Ihesa
data
are
pre­
santod
in
S
r
c
t
i
k
I
I
.
0.
t.
on
the
usage
of
thcso
srrbrturce
u
flavor
ingre­
d
i
e
n
t
s
uoro
0ttuxa.
d
fro8
tho
a8tioul
surveys
o
f
wrgcs
cmducted
by
both
FEW
and
by
N
W
N
R
C
in
1970­;
1.
Theso
data
aert
coPqri1.
d
by
the
M
I
N
X
,
a
d
revoml
calculations,
includirg
ai:
e
s
t
h
t
e
d
possible
daily
intake
value,
are
pr*
rurted
and
d
w
c
r
i
k
d
in
dotril
ia
Soctioa
1V.
B.

Ihr
uniqueness
of
the
types
m
d
amorat
of
data
presented,
u
uoll
u
the
f
o
n
u
t
for
iu
p
n
s
o
a
t
r
t
i
o
n
,
i?
i
d
i
c
t
a
t
e
d
by
the
f
a
c
t
thst
flrvorinp
sub­

stances
c
o
a
r
t
i
t
u
t
o
I
spodal
s
d
o
r
o
u
~
a
i
chemical
c
a
u
t
i
t
r
u
n
t
r
of
fwd
which
1
this
problem
has
been
tpprlached
by
coruidering
the
substances
i?
r
gxyups
or
classes
of
structurally
related
c
~~o
u
n
d
s
,'
a
s
well
as
individwlly.
nit
approach
is
j
u
t
t
i
t
i
e
d
becaucsr
o
f
t
h
e
i
r
r
e
l
a
t
i
v
e
l
y
simple
che8ical
nature
and
the
fact
that
the
great
o
r
j
o
r
i
t
y
f
i
*
r
e
r
d
i
l
y
i
n
t
o
w
e
l
l
e
r
t
d
l
i
r
h
c
d
pathways
of
metabolis8
describd
proviowly
i
n
W
'
s
Scientific
Litcraturo
k
v
i
u
of
Alipbtic
P
r
i
p
r
r
y
Alcohols,
AlMydes,
ki.&
md
Estozr
t
14
1.
For
theso
muons,
uch
of
the
r
e
i
o
n
t
i
f
i
c
1
itar8turo'
is
8
b
O
org8nixed
by
groups
of
s
t
n
a
c
t
u
n
l
l
y
r
t
l
r
t
e
d
coapounds.
A
l
l
o
f
tho
above
arterial
can
be
fomd
as
indicated
ia
t
h
o
Table
of
Catents
md
is
arranged
as
follows.
F
i
r
s
t
,
imedi8toly
follawing
this
Intro­
&tion
is
8
rupury
of
tho
biologic81
properties
and
other
irponrnt
data
fros
tho
oatirc,
reVi."

Slrctiorr
11
of
t
h
i
s
roviw
iacludos
tho
fallwing:
Tablo
11­
1
lists
ouh
subst.
aco
includod
i
n
tho
ropolct,
statu
i
t
s
chemical
i
d
m
t
i
t
y
,
u.
d
pro­
vides
8
SpMifiC
~~r
i
C
8
1
d~
si@
UtiOA
for
*&;
Tab1811­
2
Cmt8im
.­
alpha­
betical
cmrs­
mfenr;
ce
list
of
n­
r
and
sy"
3nyu
for
o
r
h
coqomd;
.
.

TJ1o
11­
3
givor
tho
physical
p
t
o
p
o
r
t
i
~
o
f
tho
substances
unhr
roviw.
Section
.I
t
!
contains
tha
dotailod
biological
data
for
each
iadividurf
compourd,
p
r
r
r
o
n
t
e
d
i
n
tdlw
containing
abstracts
o
f
all
the
individurl
paprs
frcm
which
thae
dat8
wore
t8.
k.
n.
h
b
l
o
111
preseats
i
d
b
r
u
t
i
o
a
on
~u
m
c
o
l
o
g
i
c
8
l
a
d
t
o
x
i
c
o
l
o
j
f
u
l
offrctr,
md
.st&
o1ism
of'the
substaaces.
Sctioa
IV
is
devoted
t
o
tho
pFuaF.
ce
urd
amounts
of
08ch
of
the
CM­

potads
i
n
tho
food
supply.
Table
IV­
1
i
s
drvotod
t
o
natural
occurrwtce
urd
frblo
IV­
2
p
k
v
i
b
r
infomtioa
c
o
l
f
o
c
t
~d
by
R34A
urd
NASORC
with
respect
t
o
­

tho
US8g.
10VOlS
O
f
0.
d
SubSmCO.

Section
V
is
caaplotr
biblioarrphy
of
a
l
l
articles
and
papers
con­
'
s
i
b
r
e
d
'for
us0
i
n
this
rwim,.
including
a
r
t
i
c
t
u
aot
deemed'
refovaat
u
well
u
r
r
t
i
c
l
o
r
f
r
o
m
uhidr
d8ta
wen
r
c
t
w
l
l
y
takea.
n
o
l
a
t
t
o
r
8­
noted
by
ut
asterisk
precodily
tho
ontry.
Tha
find
section
of
tho
report,
Section
VI,
pr0vid.
r
quick
nfer~
nco
to
tho
ammats
and
typu
of
d8tr
pxormtrd
for
each
s&
st.
nco
in
this
r
e
v
i
a
.
:
"
.
"

SECTION
I.
B.

Eleven
simple
esters
o
f
o
n
t
h
r
m
i
l
i
c
u
i
d
,
2
esters
of
H­
methylurthnnilic
acid
and
othyl
N­
cthylaathrurilic
acid
are
reviewed
here
for
thc*.
r
flavor
use,

All
14
substances
are
used
a
t
lcru
levels
i
n
food
a
t
average
maximum
we
.levels
.

(50
pps,
with
the
exception
of
UJO
i
n
chewing
gum
(where
wen
high
we
!evels
r
e
s
u
l
t
i
n
very
lw
ingestion)
as
follous:
methyl
m
t
h
m
n
i
h
t
c
(No,
1)
­
1583
ppm;
ethyl
anthrmilate
(No.
2)
­
116
ppm;
.ethyl
X­
methylanthranilate
(No,
12)

­
91.
S
ppm.
The
"possible
rvozqe
daily
intakes''
for
these
throo
rdstmces
f
r
o
m
cherinp
gum
US.
havo
bwa
calculated
(seo
Tablo
IV­
2)
t
o
be
0.08,
0.91
and
0.005
q,
respectively.
nesa
l
o
r
usage
levels
are
reflected
in
the
lw
mnua1
volumes
for
flavor
US.
which
rage
fmm
9
pounds
for
isobutyl
N­
ncthylmthrurilate
(No,
13)
t
o
2056
pounds
for
mthyl
'X­
nethylmthmilate
(No.
12)
and
75,300
paurds
for
wthyl
anthranilrtr
(No.
1).
Iho
calculated
(sea
intruduction
to
Table
IV­
2)
per
wit0
intakes
aro
0.02
rg/&
y
or
~w
o
r
for
811
except
for
0.7
mg/&
y
for
eethyi
anthra­
nilrt..

ents
of
Iood
($
eo
Tab10
IV­
I).
Wrthyl
m
t
h
r
u
r
i
l
a
t
r
(No,
11
occuA
in
a
d
e
r
of
f
r
u
i
t
s
:
reported
1ovels
tcing
17A
ppa
i
n
­grape
juico
and
33
p
p
i
n
Concord
grapes.
Theso
levo18
r
n
coqu8blo.
to
tho$@
resulting
f
t
a
flavor
use.
Ethyl
rnthr8nilrto
(NO.
2)
i
s
also
8
component
of
grrprs.
&thy1
N­
8athyl.
nthrdlate
(No.
12)
has
boen
roportd
t
o
c
o
a
s
t
i
t
u
t
o
0.7s)
of
bergamot
o
i
l
urd
O
.X
\
of
bitter
oranzr
oil.
The
th.
rro
largest
voluw
substaacer
covered
hen
are
811
natural
constitu­

iaTABOLISM
k
h
v
boon
discussed
befon
[F.
E.
M.
A.
1974.
Ref.
No,
141,
tho
14
esters
dcid
O?
N­
8ikyl
i
n
t
h
m
i
l
i
c
studies
on
methyl
anthrrnilato
be
hydrolyze4
t
o
an
alcohol
and
.tither
anthranilic
acid.
This
sssmption
is
supported
by
in
Vik.
0
(No.
i)
vhich,
*hilo
only
slowly
hydrolyzing
i
n
4
.

a
r
t
i
f
i
c
i
a
l
g
a
s
t
r
i
c
or
pancreatic
juice,
is
readily
hydralyted
i
n
r
a
t
liver
homogenate
(50%
i
n
27
ainutes)
and
rapidly
hydro1y:
ed
i
n
r
a
t
small
intestinc
mucosa
(SO\
in
2
.5
minutes)
itongland,
e
t
a
l
.
1977.
Ref.
­So.
34
1.
F'ethyl
anthrani.
late
has
also
be
en^
show
t
o
be
coqlercly
hydrolyzed
(,
99%
in
2
hours)
i
n
p
i
g
,
liver
homogenate
but
is
more
r
e
s
i
s
t
a
n
t
t
o
hydrolysis
i
n
pig
jejunum
homogenate
and
completely
unhydro1y:
ed
by
pancreatic
solution
[CnmCschober.
1977.
Hef.
!a.
23).
I
n
a
d
d
i
t
i
o
n
,
lpcthyl
K­
sethylmthranilate
(Yo,
12)
has
been
shown
t
o
be
hydrolyzed
on
oral
adair.
ia.
tration
to
a
hwur
and
a
r
a
t
[F.
D.
R.
L.
ulpubl.
Wep.
1963.
Ref.
No.
18
J.
A
small
amount
of
N­
Canethylatian
occurs
i
n
both
c
a
e
s
.
In
an
i
n
:u
i
t
m
StUdyD
methyl
S­
rwhylanthrrnilatt
uas
resistant
to
hydrolysis
by
p
a
n
c
r
e
a
t
i
n
o
r
p
i
g
jejuI.
um
homqenatc
but's993
hydxlyted
i
n
2
.
,

hourr
by
p
i
g
l
i
v
e
r
homogenate
[Cnmdschober.
1977.
Ref.
So.
23
1.
I
t
has
also
been
s
h
w
n
t
h
a
t
methyl
&­
8cthylanthr&,
il8te
w
a
s
ra2121Y
rbso*
d
f
r
o
m
the
small
intestin8
of
guine8
pigs,
corpletely
hydrolyzed
a
t
10u
cimcmtntionr
8nd
par­
t
i
q
l
l
y
hydrolyzed
at
higher
cor.,
entrationr
[Pelting,
e
t
a1.
Unpubl,
Rep.
'
!?
ef,.
so.
46
1.
Prompt
ditappearsrce
f
r
o
m
the
blood
of
.these
guinea
Figs
indicated
rapid
Met8bolfS8.
In
supypxy,
it
appears
t
h
a
t
t
h
e
a
n
t
h
m
i
l
a
t
e
s
are
not
hydrolyzed
t
o
any
a
p
p
r
e
c
i
a
b
l
e
e
l
t
e
n
t
i
n
the
StOmaChD
but
eXtenSiYe
if
not
COBplete
hydrolysis
takes
place
o
n
a
b
s
o
r
p
t
i
o
n
t
h
r
o
u
g
h
t
h
e
i
n
t
e
s
t
i
n
a
l
u
r
l
l
s
a
t
tht
concentrations
at
which
these
substances
aro
used.
Any
remaining
uahydmlyrd
u
t
e
i
t
a
l
can
be
hydrolysed
by
t
h
e
liver.
Excretion
of
anthranilic
acid
occurs
primarily
as
o­
minahippuric
8cid
and
t
o
8
lesser
extent
u
anthmnilic
reid
glucuronide
i
n
u.
n
[Brown.
and
Price.
1956.
Ref.
80.
5
1,
rabbits
and.
rats
(Oarrcoanct­
tiarJing,
e
t
81.
1933.
R
e
f
.
80.
8
1.

nJXICIl7
­

!n
l
i
@t
of
the
above,
it
is
obvious
that
8
discussion
of
the
toxicity
of
the
8
n
t
h
r
u
r
f
h
t
e
esters
m
u
s
t
also
i
n
c
l
u
d
e
o
n
t
h
r
8
n
i
l
i
c
a
c
i
d
d
a
t
r
,
k
r
t
h
r
8
n
i
l
i
c
r
c
i
d
is
a
nom1
Betrbolite
i
n
man
and
is
c
x
c
r
e
t
d
i
n
t
h
a
urine
u
0­
88inohip­
p
u
r
i
c
acid
(27
u#
les/&
y)
and
&&
ranitic
acid
alucutonid+
(6
vwXes/
day).
Ihir
is
equiV8h1t
t
o
tot81
rvetrge
d
8
i
l
y
excretion
of
&art
f
a#
urthturilic
a
c
i
d
p
e
r
day
[Ria.
e
t
81.
19%.
bf.
NO.
47
1.
.
hlr
liritcd
d8t8
a
r
e
8
v
t
i
l
.b
l
e
on
t
h
e
toxicity
of
a
n
t
h
r
a
n
i
l
i
c
a
c
i
d
;
however,
the
Or81
tbso
of
4S49­
mglkg
i
n
r
a
t
s
1R.
T.
E.
C.
S:
1977.­
Ref.
SO.
48
]
indicates
low
,toxicity.
It
ha
been
reported
5
..

[Et­
and
Strombeck.
1949.
Ref.
Xo.
I?]
that
O.?\
in
the
diet
of
rats
pro­

duces
blodd..
r
papilloma^
on
long
feeding.
The
oral
iDjO
values
for
the
esters
are
also
quite
h
i
f
i
for
t
h
i
s
class
o
f
compounds,
vllues
ranging
fnPl
2250
m&/
kg
for
methyl
S­
methvlanthrsn:
'atc
(SO.
1:)
13
guinea
pigs
t
o
>SG30
mdl'kp
in
rats
for
several
o
f
the
&t
e
s
(see
Table
A­:).

The
two
anthranilates
uith
highest
eaposurc
have
botl;
h
e
n
studied
more
extensively.
Methyl
anthranilate
(ertinroted
as
?xplained
in.
the
Introduction
t
o
Table
I
V
­2
t
o
have
a
l
e
r
qitct
daily
intake
o
f
0.31
8g/
kg
for
a
t4
kg
person)
h*
s
been
added
t
o
t
h
e
d
i
e
t
of
r
a
t
s
at
levels
;rf
?OOO.
and
10,000
ppm
trpproximtely
equivalent
t
o
an
averape.&
rfly
intake
of
SO
and
500
mg/
kg
for
an
adult
rat).
f
o
r
.
13
ueeks
u
i
t
h
no
Ldverst
effects
.(
Haoan,
e
t
81.
1967.
Ref.­
So.
27
j.
'kthyl
X­
methylanthranilrtcr
(No.
12)
(estimated
per
qitc:
daily
intake
of
.C.
OOO,
t
ag/
kg
f
o
r
a
60
kg'pcrson)
added
t
o
the
d
i
e
t
of
rats
a
t
levels
o
f
300,
1230
and
3604
ppm
(approximately
1s.
LO
and
120
ag/
kg/&
y
for
8n
a
d
u
l
t
r
a
t
)
for
90
days,
[Cauirt,
e
t
81.
1970.
Ref.
No.
23
f
resulted
i
n
8
s
l
i
g
h
t
b
u
t
s
i
g
n
i
f
i
c
a
n
t
leukocytopenia
and
anemia
a
t
t;.
e
2
higher
levels
urd
i
n
c
r
r
u
c
d
kidnc;
'
w
i
g
h
t
s
i
n
tLe
u
l
e
s
.

No
effects
were
seen
at
300
?pm.
A
drily
intake
of
19.9
q
/k
g
(males)
and
22.2
&kg
(femles)
by
.rats
for
90
days
caused
no
8dvem.
e
effects
[Oscr,
e
t
a
l
.
1965.

Ref.
So.
441.

*
Both
methyl
anthranilate
(No.
I
)
urd
einnuyl
anthrrailate­(
No,
IO)
in
tricapxylin
solution
uem
infoetad
intrrperitonerlly
3
tines
per
week
for
I
weeks
a
t
the
saxfur
tolemted
des.
(WD)
8nd
a
t
0.2
tire
the
WfD
in
me
mice
uhich
were
maintsined
for
15
rdditiorvl
uetks
betom.
sacrifice
and
autopsy,
Methyl
a
n
t
h
n
n
i
h
t
e
w
u
injected
i
n
repante
doses
of
0,09
d
k
g
(0.2
m)
a
d
0.47
g/
bg
(WTD)
for
8
­total
doso
of
e
i
b
r
2.25
or
11.2
g/
kg
t
o
20
f
c
u
l
e
i
a
t
s
'
.

each.
A
t
t
h
e
end
of
t
h
e
24
ueek
test
period
(I
week
administration
and
16
ueek
observation)
3
of
tho
18
survivors
(ln)
at'
tho
1­
dose
md
S
of
the
19
sw\*
ivon
(26%)
a
t
t
h
e
m3
had
developed
lung
tumors.
C
i
n
n
q
l
a
n
t
h
r
a
n
i
l
a
t
e
MU
injms­
d
i
n
separate
doses
of
0.1
g/
kg
(0.2
MTD)
and
0.5
g/
kg
(MD)
for
(L
total
dose
of
.
e
i
t
h
e
r
2.4
or
12.0
o/
kg
t
o
IS
male
urd
IS
ftmale
tats
each.
In
t
h
i
s
case,
a
l
l
male
rats
8nd
IS
f
e
u
l
o
r8ts
survived
a
t
either
dose.
Sev?
males
(47E3
and
6
f
e
u
l
e
s
(468)
a
t
tho
low
dose
urd
14
v
l
c
s
~~(Y
3
t
)
and
7
f
d
e
s
(S4%]
a
t
the'wTD
developed
lung
ttrwr,.
O
f
the
80
m
l
a
urd
8C
femslc
controls
injected
similarly
with
tricaprylin,
77
8810s
and
t?
females
survived­
the
test
petioa.
Of
these
z8t
of
tha
males
md
20\
.of
the
fe=
lm
developed
lung
tmm
[Stoner,
et
1973.
Ref.
No.
531.
.c
...­
,>

6
..
..­

PI:
,""
.
.
.
"
e..­­
."
.
~

L
l
k
1.
,/

.,

1
,

With.
tha
exbcpticns
of
isobutyl
S­
methy1anthr:
milatc
(So.
13!
and
ethyl
.N­
e:
hylanthranilat~
(No.
14).
which
have
never
bees
submitted
to
the
Pane!,
311
o
f
the
anthranilates
discussed
here
nave
been
reviewed
by
t
h
e
Exper,
oanel
of
F.
E.
N.
A.
and
found
to
be
Generally
Recoyrlired
as
Safe.
(CRAS)
under
the
condi­
tions
o
f
intended
use
as
flavor
ingredients
(F.
E.
M.
A.
1965­
1978.
Ref.
No,
15
1
.
lhis
determination
w
a
s
based
on
a
review
of
a
l
l
available.
data
as
well
as
the
rruonable
analogies
which
the
Panel
juciged
could
be
drawn.

.#
..

..

c!
':
.
se+^
so
"
S
O
Substance
Spec
ios
Rout
e
­
­
L
Werhyl
anthrazilatc
hb
us
e
Xntrapeti
toneal
Rat
Intraperitoneal
).
bcs
e
Ora
1
kat
(fasted)
Oral
Guinea
pig
Oral
(fasted)
­
17
30
1
7
30
30
Oral
Oral
2
Ethyl
anthrmilate
Mouse
Rat
17
42
4
Sutyl
anthranilate
Rat
Oral
38
Oral
­
4
1
I!
.
..
'..?.
...
<..
..
Phenethyl
anthranilate
Rat
Oral
43
Oral
>S
g/
kg
39
10
Cinnamyl
anthranilate
Rat
12
Methyl
N­
mtthyl­
Rat
(fasted)
anthranilate
,.
,Rat
Or81
Oral
23
2.2s­

3.7
ml/
kg
3
.3
g/
kg
40
8
SECTIN
11.
A.

NUMERICAL
DESIGNATION
AND
CHEMICAL
IDENTITY
TAEE
11­
1
This
table
lists
the
prirpary
names,
chemical
structures,
molecular
formulae
and
molecular
weights
of
the
substances
included
i
n
this
review.
These
Corupoun~
8­
listed
by
chmical
stxucture
as
explained
bclw,
and
am
assigned
I
numberby
which
they
a
b
referred
thmughout
th8
review.
In
order
t
o
mom
easily
sake
comparisons
between
metabolically
nlated
substances,
the
compounds
are
organired­
md
n
d
e
r
o
d
according
t
o
chemical
structure
pro­
gressing
f
r
o
m
r
t
q
l
e
s
t
t
o
.
more
complex.
.
A
cross­
rsference
of
the
restances
and
t
h
e
i
r
s
p
o
n
p
s
is
listed
alphabetically
­in
Table
11­
2.
Additional
inclusions
in
Table
11­
1
are:
a)
'the
n
h
e
r
assigned
t
o
t
h
e
substance
by
the
Flavor
and
Extract
Manufactu~
e~
s~
cIJsoci8tion
(€
Em)
in
its
prtblicatioas
I
f
]
listing
substances
judged
by
the
Expert
Psnel
of
FEMA
t
o
be
generally
recognized.
u
safe
conditions
'of
intanded
use;
b)
the
section
o
f
2
1
Cod8
o
f
Federal
R
e
p
l
s
t
i
o
n
r
i
n
which
tho
r
r
~s
t
a
n
c
c
l
have
been
l
i
s
t
e
d
by'
the
Food
and
Drhg
Mn$
strrtion
'(
FDA)
;
c)
the
QIdca1
Abstracts
registry
number.
I
hl
rr)

bn
bn
N
b
c.

..

L.
l
rr)

m
L.
l
0
r
4
CJ
0
Y
L1
"

e4
0
11
c;
m
N
..
e
5
.4
=m
L.
l
m
u
Y
r(
e
a
Y
c
.a
c.

c
x
w
Y
10
N
..
.:

L
m
m
hl­
c4
.II
R
0
2
Q
v
s:
Fi
Q
1
I­
wl
(Y
Q
hl
N
2
Q
wl
n"
yr
LI
*!
'
4
u
n"
h
=­
i
c
­.
u­
u"

c
I
CI
e
Y
d
e
Y
8
C
L.
4
t
s
5
0
*,

d
a
*
a
a
"
.d
4
&
CI
Y
rl
3
x
­3
L
0
d
LI
2
x
Q
i
a
9
Q
Y,

cy
t­
I
I
I
I
I
2
­m
9
L.
.I
n
N
m
­
N
rn
N
9
8­

N
0
n"
2
*a
U
U
?­
h
8"
In
=e4
U"
r.

­0
Y
L..

r.
r.
SECTION
11.
b".

ALPHABkTIUL
CROSS­
REFERENE
LIST
OF
SAME5
AND
SySoNy).
Ls
TAB&
11­
2
Thrr
tablo
.lists
a
l
p
h
i
b
e
t
i
d
l
y
the
principal
names
(i
n
'
c
a
p
i
t
a
l
e
t
t
e
a
)
a
d
synonyms
[in
louer
case
lotters)
of
the
substances
i
n
t
h
i
s
roviw.
A
l
l
synonyms
am
l
i
s
t
e
d
a
f
t
e
r
o
a
A
principal
name
u
uoll.
Iho
ndcrs
refor
to
tho
rtnrctunl
1is.
tiag
i
a
T
d
l
o
11­
1
and
indiuto
tho
ordering
of
tho
tub­

ttmccu
i
a
a11
othor
tdlos.

mi3
cmrr­
nteGacr
list
inc1dws
tho30
synonyms
most
cauDnly
found
in
tho
literatwo
md
synonym,
including
soma
trado
names,
frequently
wed
in
tho
flavor
indwtry.
In
a11
c
u
.3
thr.
rtder
is
rearred
t
o
the
cqitalired
nasa
u
h
i
d
is
the
n
l
w
used
throug.
ho*
t
h
i
s
review.
No
­

3
4
LO
i
6
TABLE
11­
2
AX
nt
tu.\
I
LATE
s
Substvrse
Si;
ron
\IS
.

~l
l
y
l
2­
rpinobentoate
[ALLYL
&W
I
L
I
\T
E
­­n
o
.
31
A
l
l
y
l
o­
iuninobntoate
[A
l
l
Y
L
"HRNIUTE­­
no.
31
ALLY
t
A
S
l
I
i
i
i
X
I
UTE
A
I
l
y
l
o.­
uninobcururrr
Allyl
2­
aminobcnrootc
2­
Propenyl
?­;
lrPinobentoatt
:­
Propenyl
anthranilate
Vinyl
carbinyl
anthruri:
rta
6­
Amino
methyl
benzoate
[WTHYL
A!!
WIUTE­­
no.
11
Butyl
o­
uinobsnzocrte
[BLIIYL
~1
L
A
l
E
­­n
o
.
41
iw'­
htyl
o­
aminobenroato
[ISOSUlYL
MTHUNiUTE­­
no.
5;

Butyl
2­
rninobcnzonte
[BUIYL
MMWILATE­­
no.
41
..

BUTYL
AKMRANIUTE
Rttgl
2­
aminobcntoate
Wltyl
o­.
cuinolenrorte
b
­k
t
y
l
&n
t
h
~i
h
t
.
1
I
S
8
V
n
L
AMMRANIUTE­­
M).
S]

CiMIII)
Il
o­
adnoburroat.
iC1­
t
NIH~
IU'TE­­
T~
O.
10)

Cinnuyl
2­
ainobmzoate
[CIMUNfL
AMHRANIUTE­­
no.
10)

c
INNAMYL
mRAN1
UTE
C
i
n
n
u
y
l
2­&
imbennrorto
Cinnuyl
o­
ainobcnroate
3­
Phenyl­
2­
propenyl
2­
amino­
benzoate
3
­P
h
~y
I
­t
­p
m
p
~t
8nthIWlil8te
Cyclohexyl
2­
uinobentoato
[CTC­
XYL
AKIHIUNfUTE­­
m.
6j
CYCLOHEXYL
ANTHRANIUTE
Cyclohexyt
2­
aminobcntorte
..

14
­

i_
No
"
Subrtmc.
Synonps
Diethyl
unthranihtc
[E
M
L
X­
LTHk'UhiIWIUTE­­
no.
141
Dimethyl
anthranilate
(WTHYL
S­.
WMU,
XWlUTE­­
no.
121
.

3,7­
DiP~
thyl­
l,
O­
octa~
ien­
3­~
1
2­
ullincbur:
oate
JUUAYL
ApiTHaASlUTE­­
no.
7
)

$,
7­
Pi~
thy1­
1,6­
octrCica­
J­~
l
urthrrnilato
~L
l
W
Y
L
AN'MWlUTE­­
no.
t
]

Ethyl
o­
dnobenzoatr
[ETHYL
ANWXLATE­­
no.
:!

Ethyl
2­
dnobrnroate
[ETHYL
~l
U
T
'
E
­­n
a
.
2)

Ethyl
o­
adnobenzoate
Ethyl
2­+
miaobmrolitr
5
Diothyl
.nthturilrto
Ethyl
2­
ethyluinobenroate
Ethyl
(2­
ethyluiaophcnyl)
­
8ethuxnto
Isobutyl
2­
uinobratorto
(ISWJlYL
A
~'
M
I
U
T
E
­­n
o
.
5)

Isobutyi
2­~
ethyluinobenrmtr
[
ISO%
VTrt
N­
IIEMYUMHRANIUTE­­
ao.
131
Isobutyl
2­
methylmino­

24ethylpmpyl
2­
wthyl­
kntoatr
uinobaitmt~

tinalyl
2­
uinobenroato
(UN&
Yt
ANTHRANIIATE­­
no.
71
7
1
it
11
Subs
t
mce
Synonyms
Linalyl
c­
rminobcnroati
[LIsMYL
ANHMNItAI'F.­­
no.
?]

LINALYf.
&T1iRA!
iILATE
3,~­
Diwrhyl­
1,6­
octadicn­

3.7­
Dimethyl­
i
,+
oitadien­

Linalyl
2­
aminobenzoate
t
i
h
~&1
y
.
c­
aninobtnroatr
3­
yl
2­
ainoben:
oate
3­
71
rnth1urilate
p­)(
mthhr­
l­
m­
tyl
2­
uinobonroate
ja­
TERPINYL
&7lWh(
IUTE­­
no.
81'

p­
l4enth­
l­.
rr­
l­
y1
~thraniIgte
[a­
TERPINYL
N7HRANI'JTE­­
no.
81
2­
lbthtluriao
wthyl
bonroate
[)
4EIliYL
N­
lETHY~
RMIUTE­­
no.
12)

m
Y
L
AKIHRANIIAX
o­
kino
w
t
h
y
l
bcnroa:
e
kbthyl
o­
uinobenzoate
Methyl
2­
uinobcnroate
Dimethyl
8nthr.
nilrto
2­
libthyluino
rtthyl
bontoate
Matby1
2­
rwthyluinobentoate
Wthyl
o­
methyluinobenroate
2­
Naphthyl
o­:
rinobaroato
[#­
NAPWHY1
ANMRANItAfE­­
no.
111
2­
Naphthyl
6­
uinobenzoato
2­
Nqhthyl
mthrmilrto
2­
Naphthyl
m
t
h
r
u
r
i
l
a
t
o
f8­
U"
HYI.
uJTHRAxIUTE­­
no.
111
16
.
No
Substance
­
Synonyms
6­
Phencthyl
o­~
minobcn:
o;
rte
2­
Pharylethyl
mthrmilate
2­
Phenylethyl
anthranilate
(PHENETHTL
Ah'l?
lRAi.
YIUTE­­
no.
01
3­
Phrnyl­
2­
propenyl
2
­u
i
n
o
t
a
t
o
o
t
e
[CINNQIAHI'L
ANMRMfUTE­­
rro.
101
8
a­
TERPINYL
ANTHWIUTE
­
"_.
."
.
.­
"

1
SECTION
I
i
.C.
­
..
I
I
I
t
should
further
be
erqhuird
that
the
data
and
descri?
tion
of
sub­

stances
contamed
within
this
table
r're
those
lasted
in
t
h
t
referenced
source
and
no
o
t
t
e
q
t
Lss
been
made
t
o
c
l
a
r
i
f
y
these
listings.
;or
eurnplc.
although
sone
substances
ore
listed
as
color!
esr;
i
t
should
not
be
asslmpcd
that
com­
pounds
not
so
designated
are
colord.
As
another
exaprple,
the
d,
ifference
between
sOlubl8
and
8isciblr
w
u
not
aluays
clearly
defined.

Abbreviations
Used
in
This
Tablt
alc
I
ethyl
alcoho:

s.
6
solubl.
far
insoluble
rls
slightly
soluble
l
i
q
­
liquid
mr
6
melting
point
bp
=
boiling
point
rpg
=
specific
gravity
d
=
density
doc
6
decowsu
19
I
i
I
"
"
"
­
1
i
L
I
+

I
­
.*
­
9
4
1
"

"­
b
i
a
3
..
a
­
f
2
.
3
f
"'
I
..
I
c)

t
4
YI
22
..................
........
.............
......
...
....
.......
.
.
................
.............
..
........
:
>.:..­
,
.:.
....­,
..._
I_
".
i
i
z
t
P
W
..

2.4
SECTION
I
If
TAELE
111
lhis
tablo
contains
inforvtion
en
tho
biological
yroperties
and
.stabolism
of
the
substances
included
in
this
review.
Tho
bulk
of
this
inferntion
uu
derived
f
r
o
m
t
r
t
i
c
l
o
s
gt*
rined
­Cram
t
coaprshcnsive
survey
of
tho
s
d
o
n
t
i
f
i
c
l
i
t
o
r
r
t
u
m
published
betkcen
1.920
and
1977.
This
survey
v
u
carriod
out
by
an
indepeaant
orgurizstioa,
bfomticsD
lac.
of
RocLVillU,
Cluy1­
d.
..

hfomatics,
Inc.
u
r
d
the
chaical
rima,
c
o
m
n
naaos
md
syaonym
of
each
substance
as
key
uords
t
o
locato
reforcrncer
t
o
tho
publish&
l
i
t
e
r
a
t
u
r
e
pertaining
t
o
tho
p
h
r
r
k
o
l
o
g
i
u
l
urd
toxicological
daQ
on
thoso
substances.
Summaries
of
a11
the
portinont
articles
fro.
this
survey
worn
then
written
and
are
pnsentod
i
n
this
tal..
Othor
data
c
o
l
l
e
c
t
e
d
f
m
a
o
u
l
i
o
r
litera­

two
md
privrtr
industry
SOUIICOS
8ro
also
s­
izod
.and
includod
h
m
i
n
.
Iho
tablo
is
orgaairod
by
subttaac.
with
the
simplost
c
h
u
i
c
a
l
stnac­
t
u
o
s
procoding
a0
.on
C­
l.%
D
u
in
oqor
roctions
of
this
rovim.
me
nuberr
rftor
tho
nmo
of
oach
substurco
uo
the
same
f
o
i
a11
the
other
tablos
in
the
riviou.
rithia
or&
substurco.
gmup.
tho
entries
ue.~
orpnized
<.­.

!
25
..
bferencc
KO.
:
3.1
(Longland,
W.
C.,
e
t
01.
Toxicology.
1>
77)

b
t
i
I
y
1
a
n
t
h
r
a
n
i
l
a
t
e
uas
i
n
c
u
b
a
t
e
d
w
i
t
h
a
r
t
i
f
i
c
i
a
l
g
a
s
t
r
i
c
juice
aJ
a
r
t
i
­
ficial
Fancreat
ic
j
u
i
c
e
at
37.
for
up
to
J
hours.
The
time
required
for
SO\
hydrolysis
bas
5950
and
4150
minutes,
respectively.

i
n
SO$
hydrolysis
in
27
and
2,
s
minutes,
respectively,
Incubation
hith
rat
l
i
v
e
r
homogenate
and
small
i
n
t
e
s
t
i
n
e
mucosa.
resulted
Reference
No.:
25
(Gradschober,
F.
Toxicology.
1977)

Methyl
a
n
t
h
r
a
n
i
l
a
t
e
a
t
a
cowentrarion
of
100
or
250
u
l
/l
w
a
s
incubated
i
n
0.5M
phosphate
buffer
(pH
7.5)
at
37.
w
i
t
h
e
i
t
h
e
r
p
a
n
c
r
e
a
t
i
n
,
pig
jejunum
homogenate
o
r
p
i
g
l
i
v
e
r
homogen8te
for
2
hours.
C
U
u
d
l
y
s
i
s
r
e
v
e
r
l
d
0,
1s
urd
,998
hydrolysis,
respectively,
of
t
h
e
ester.
I
,
N
m
of
Substance:
!EB'zHyi
AKl?
iRANXLATf
(1)

Reference
No.:
17
(F
k
Unpubl.
Rep.
1977)

Species:
Nouse
No.
/Croup
:
10
Duration:
Acute
Route
:
Intraperitoneal
Vehicle:
Sot
Specified
Control
0
Sot
Specified
Tho
intraperitonoa:
LD
f
o
r
methyl
anthtanilato
i
n
l
i
c
e
u
u
reported
t
o
be
1.04
el/
kg
(958
C.
L.
Oy­
1.19
W
k
g
).

Referonce
No.:
30
(Janner,
Z
.M
.,
e
t
81.
Food
Cosaet.
loxicol.
1964)

Spocies
:
.House
No./
Group:
Not
Specified
[kration:
Acuto
(14
day
obremtiou)
Route:
otrl
(intub8tion)
\'ahicle:
Nert
Control:
Not
Sprcifid.

Tho
om1
LD
(Litchfield­
Wtlcoxon)
for
methyl
anthnnilato
in
mice
w
a
s
calculattd
t
o
beSg90O
8
d
k
g
(OS8
C.
L.
3260­
4680
&kg).
Symptom
of
toxicity
included
depression
and
death
uithin
4­
18
hours.

i
i
.
c
*
k&
J
\

Roforenco
No.
:­
53
(Stoner,
C.
D.,
e
t
a1.
Cancer
Res.
1075)

Species:
)louse
(W
e
)
Rout.
:
Xntrrp.
riton&
l
No./
Crwp:
20
(F)
Vehicle:
Trieapxylin
Duration:
8
w
e
+L
t
(24
week
obsonation)
Control
:
Tricqrylin
alone
Mica
wore
injected
inttrperiteneally
uith
24
d
a
m
,
cithar
0.09
g/
kg
­or
0.47
g/
kg.;
of
methyl
a
n
t
h
r
8
n
i
l
i
t
o
i
n
t
r
i
c
r
p
x
y
l
i
n
3
tims
per
week
for
8
weeks
f
o
r
8
t
o
t
a
l
dose
of
eithor
2.23
or
11.2
fig.
A
t
the.
higher
dose,
19/
20
feaalos
sunivad
the
treatment,
while
a
t
the
lower
dam
.the
survivors
wero
18/
20.
A
t
the
ond
of
the
24
week
test
period,
3
(268)
had
develoyd
luly
tumrs
a
t
tho
high
dose
and
5
(In)
had
a
t
t
h
e
lower
dolo.
Y
i
t
h
tricapryfin
alone
them
utre
77/
60
male
and
77180
female
survivort
of
which
28*
of
the
males
and
20%
of
the
females
developed
lung
tmors.

!27
Species
:
bat
.io.
/Croup
:
10
Duration:
Acute
Kefertnce
So.:
1
7
(Hapan,
E
.C
.,
e
t
a
l
.
Food
CoSmct.
Toxicol.
1967)

Species
:
Kat
(Osborne­
Sendel)
Route:
Oral
So./
Croup:
20
(10M
L
10F)
Vehicle:
Diet
Duration:
13
weeks
Control:
Diet
810m
'
.
Methyl
anthranilate
was
administered
a
t
diotrry
concentrations
of
1,000
*qd
10,
COO
ppm
(approximately
equivalent
t
o
8
daily
intake
of
50
and
500
q
/k
g
i
n
a
d
u
l
t
r
a
t
s
)
t
o
weanling
Osborne­
Mendel
r
a
t
s
f
o
r
13
weeks.
Xo
effect
ras
seen
on
g
n
n
t
h
or
hesatology,
.IS
determined
by
an
examination
of
white
and
("?<
I.
.
red
blood
c
e
l
l
count,
Iwrtoglobin
8nd
hematocrit,
i
n
e
i
t
h
e
r
group.
So
gross
;$
tissue
changes
uere
found
i
n
e
i
t
h
e
r
group,
nor
uere
any
microscopic
changes
d
d
observed
i
n
t
h
o
high­
dose
gmup.

28
Reference
No.
:
30
(Jenner,
P.
M.,
e
t
a
l
.
Food
Coraet.
Toxicol,
1964)

Species
:
Rat
(Qsborne­
libndal)
Route:
Oral
(intubation).
No.
'Group:
10
(SM
6
SF)
Vehicle:
Seat
Duration:
Acute
(14
day
Obsenration)
Control:
Not
Specifid
f
u
t
d
r
a
t
s
up3
8
h
l
a
t
.d
to
bo
7910
&kg
(9S\
C.
L.
1500­
3400
'mgjkg).
Syaptom
of
toxicity
includod
depression,
c
o
u
upd
death
vithia
1­
2
days.
The
oral
LD
(titchficld­
WilcoxQn)
for.
methyl
anthranilato
in
18
hour
Reference
No.:
.
30
(Jenner,
P.
N.,
e
t
a1.
F
c
o
d
Cosset.
Toxicol.
1964)

Tho
oral
LD
(tiidfield­
Nilcoxon)
for
­thy1
mthnnilato
in
18
hour
fast&
pucnea
piat
YU
calculated
t
O
be
2780
rg/
kg
(9SI
C.
L.
.
2210­
3500
8g/
kg).
SpptoM
of
toxicity
included
depression,
gasping,
rapid
respiration,
sutro­
i
n
t
e
s
t
i
n
a
l
i
r
r
i
t
a
t
i
o
n
and
death
vithin
4
hourr
t
o
4
days.

29
Sme
of
Substance:
E
W
L
ILYIHRAh'liAfE
[2
]

Reference
KO.:
17
(FW
Cnpubl.
Rep.,
1977)

Specjer:
Nouse.
.

ro./
Cloup:
10
(W
6
F)
Durrtron:
Acute
&l
i
t
e
:.
Oral
(intubation)
Vehicli.
Not
Specified
Control
:
Not
Specifrd
The
oral
LD
for
ethyl
anthranilate
in
mice
w
a
s
reported
t
o
be
3.37
ml/
kg
(958
C.
L.
ao2.78­
4.60
ml/
kg)
.

Referencr
No.
:
42
(Opdykc.
D.
L.
J.
F&
d
Cosoct.
l
o
x
i
c
o
l
.
1976)

spiciss
:
Rat
Route:
Oral
No./
Group:
Not
Specified
Vehicle:
.
Sot
Specifid
Ouration:
Acute
.,
Contrcl
:
Sot
Specif
id
&'kg
(958
C.
L.
­'g.
Y­
4.18
g/
kg).
The
oral
LD
for
ethyl
mthrmilote
i
n
r8:
s
YU
reported
to
be
S,
7t
,.
3
0
'.
X'irme
of
Substance:
ETHYL
AXRiRAI;
I!
ATE
{2
)

Reference
KO.:
1
7
(FDA.
Unpubl,
Rcp.
1977)

.Species:
PWse
%o./
C,
roup:
10
(N
6
F]
Durrtlon:
Acute
Route:.
Oral
(intubation)
V2hicI.
a.
Not
Specified
Control
:
Not
S
p
e
c
i
f
i
d
The
or81
LO
for
ethyl
anthranilate
in
mice
u
u
reported
t
o
be
3.57
d
/k
g
(958
C.
1.
3°
2.78­
1.60
81/
kg).

Referenca.
No.:
42
{Opdyke,
D.
L.
J.
FoodXosaet:
Toxicol.
1976)

spocia:
Rat
Route:
O
r
a
l
No./
Croup:
Not
SpUlfied
Vehicle:
Sot
Spwified
(kntion:
Acute
t
o
n
t
x
l
:
Sot
Sptcif
id
me
oral
LD
for
achy1
urthrmilate
in
r8:
s
v
u
reported
t
o
be
3.75
&kg
(95%
C.
L.
­sg.
32­
4.18
g//
kg).

30
i
N
.u
of
%brturco:
S
u
n
&
AMHRAN:
uT€
[4]

Reference
No.:
38
(Opdyke,
D.
L.
J.
Food
6
%
Spec
ics
:
Rat.
No./
Crouy:
Not
Specified
Owation:
Acute
~h
r
oral
for
butyl
anthranilate
than
5
p/
kg.
Cosmet
.
Toxicol.
1975)

Rout
e
:
Crol
Vehicle:
trot
Specified
,
Control
:
Not
Sp­
ified
in
rats
vi&
reported
t
o
be
greater
Samo
o
f
Substance:
LIMLYL
LiNlXIHRASIUTE
[7
j
Reference
tio.
:
41
(Opdyke,
D
.L
.3
.
Food
Comet.
foxicol.
19'6)

Species:
Rat
SG.
iftoup:
>kt
Specified
Dura
ion
:
Ac
ut
e
Route:
Oral
Vehicle:
,Uot
Specified
Control:
Nor
SyccifirZ
'.
L'

I
32
I
Naae
of
Substmci:
PHENRhYL
ANTHRASIUTE
191
Reference
No.:
43
(OyrtyLc,
D.
L.
J.
Food
Cosmet.
Toxicol.
1976)

Species
:
Rat
No./
Group:
tiot
Specified
&ration
:
Acute
Route:
Or01
Vehicle:
"
Kat
Specified
.Control
:
Sot
Specified
oral
LD
for
phenethyl
anthturilateain
rats
w
a
s
reported
t
o
be
greater
than
5
24,.
I
I
1
Nsme
o
f
Subrtuce:
ClNNAHYL
AlSTHSWIUTE
[lo)

Reference
do.:
S
j
(Stoner,
G
.D
.,
etoal.
Cexer
b
s
.
1973)

Species
:
Mose
(NHe)
Route:
Intraperitoneal
Duration:
8
weeks
(24
week
observation)
Contml:
Tricaprylin
alone
N
o
.i
C
~p
:
:#
O
(194
I
1SF)
Vehicle:
Triuprylin
Mice
were
injected
intraperitoneally
with
24
doses,
either
0.1
g/
ky
or
0.
S
g/
kg,
of
cinnllpyl
ant).
zonirate
i
n
t
r
i
c
a
p
x
y
l
i
a
3
times
Fer
week
for
8
weeks
ior
a
t
o
t
a
l
dose
of
either
2.4
or
12.0
dkg.
At
the
higher
dcse,
1S/
15
mala
and
13/
15
fe­
les
sunived
the
treatment,
bhile
a
t
the
lower
dose
the
survivors
wero
13/
15
males
and
13/
15
females.
A
t
the
end
of
the
24
week
test
period,
I4
males
(93t)
and
7
females
(S48)
had
developed
lung
twrs
a
t
the
high
dose
and
7
(47%)
moles
1nd.
6
(468)
­females
had
af
t
h
e
lower
dose.
K
i
t
h
t
r
i
c
a
p
y
l
i
n
­alone
there
were
77/
80
male
ann
77/
80
'female
survivors
of
which28%
of
the
.

des
urd
20%
of
the
females
developed
lung
tumors.

Reference
No.:
.39
(Updyko,
D.
C.
J.
Food'
Cosmef.
Toxicol.
1975)

Species
:
Rat
No./
Croup:
Not
Specified
Ruation:
Acute
Route:
Oral
.
Vehicle:
Not
Specified
Controi:
Not
Specified
I
Name
of
Substance:
WEIIML
I\"
MET"
niRAKIUTE
[12]

Reference
No.:
18
(F.
D.
R.
L.
'hpubl.
Sep.
19s;)
I
I
One
a
d
u
l
t
r
a
t
each
was
adainistercd
.orally
(stomach
tube)
a
1.0,
5.0
or
SO
mg
dose
of
methyl
N­
methylar!
tnranilate.
Analysis
of
the
24
hour
urine
revealed
unsytcified
amounts
0''
S­
merhylanthranilic
acid
ar.
d
anthranilic
acid
i
n
a
r
a
t
i
o
of
approximately
:O
i
l
.
7he
authors
concluded
tfiet
ingestlot
c
f
methyl
N­
mathyl.
anthnnilote
is
followed
promptly
b)
d
e
e
s
t
e
r
i
f
i
c
a
t
i
o
n
w
i
t
h
t
h
e
urinary
elimination
principally
of
the
X­
methylated
acid.

Reference
No.
:
46
(Pelting,
e
t
01.
Unpuol.
Rep.
)

Metnyl
H­
ut­
thylanthranilate
a
t
concentrations
ranging'
from
25
t
o
­
260
p
p
~
i
n
physiological
saline
nas
i
n
j
e
c
t
e
d
i
n
t
o
t
h
e
duodenal
lmcn
of
male
guinea
pigs
a
t
a
dose
volume
o
f
5
.ml/
kg
boiyweight
a
t
o
rate
o
f
6
mllmin.
A
ligature
was
t
i
e
d
around
t
h
e
duodenvnr
n
e
a
r
t
h
e
p
y
l
o
n
s
t
o
p
r
e
v
e
n
t
r
e
g
u
r
g
i
t
a
t
i
o
n
i
n
t
o
the,
stoaaach.
Samples
of
portal
blood
were
taken
a
t
2,
5
,.
?O.
20
and
30
minutes
and
analyz­
ed
for
unhydrolyzed
e
s
t
e
r
.
TBe
r
e
s
u
l
t
s
i
n
d
i
c
a
t
e
t
h
a
t
.
methyl
fz­
methyl­
a
n
t
h
r
m
i
l
a
t
e
is
rapidly
.absorbed
a
t
a
l
l
c
o
n
c
e
n
t
r
a
t
i
o
n
s
and
i
n
a
conp:
ctcly
hydrolyzed
form
at
25
ppo.
So
unhydrolyzed
e
s
t
e
r
wits
detectable
10
minutes
a
f
t
e
r
i
n
f
e
c
t
i
o
n
of
the
40
pym
solution
and
20
ainutes
after
injection
of
*he
120
ppm
s
o
l
u
t
i
o
n
.
w
i
t
h
t
h
e
260
ppm
solution
a
peak
concentration
of
?.?
6
ug
ester/
rpl
o
f
blood
uas
r
e
t
c
h
e
d
a
t
t
h
e
3
minute
sampling
and
a
­concentration
of
0
.~6
ulJm1
remained
a
t
t
h
e
30
minute
sampling.

Reference
No.:
25
(Grundschober,
F.
Toxicology.
1977)

­
Methyl
N­
methyla&
lirurilate
at
a
concentration
of
15
o
r
250
ul/
i..
was
i
n
c
u
b
a
t
e
d
i
n
0.
SU
phosphate
buffer
(pH
7.5)
at
with
tither
pancreatin,
p
i
g
3ejunwn
homogenate
or
p
i
g
liver
homogenate
for
2
hours.
G
U
x
a
l
y
s
i
s
revealed
0
,
IS
and
,99%
hydrolysis,
respectively,
of
t
h
e
ester.

35
.
­.

Name
of
Substance:
MERfYL
N­
CIE'MYWJRRANILATE
[I
t
]

Reference
Yo.
:
18
(F.
Q.
R.
L.
U~
publ.
Rep.
1963)

One
human
volunteer
vas
administered
orally
a
single
150
mg
dose
of
methyl
X­
methylanthranilate
and
urine
w
a
s
c
o
l
l
e
c
t
e
d
a
t
7
hours
following
treatment.
Analysis
revealed
unspecified
amounts
of
N­
methylantkranilic
acid
and
anthra­
n
i
l
i
c
a
c
i
d
i
n
a
r
a
t
i
o
of
approximately
20
:l.
The
authors
concluded
t
h
a
t
inges­
t
i
o
n
of
methyl
N­
methylant5ranilate
is
foliowed
promptly
by
deesterification
with
the
urinary
elimination
principally
of
the
N­
mccnylated
acid,

.
.
36
Reference
No.:
4
(8ar,
F.
and
F.
Griepetltrog.
Xed.
Ernachr.
,196;)

Species:
Rat
Ho./
Group:
Not
Specified
Duration:
12
weeks
Rout
e
:
Oral
Vehicle:
Sot
Specified
Conrro
1
:
tiot
Specif
id
For.
12
weeks,
rats
uere
administered
daily
a
20.3
mg/
kg
dose
of
methyl
N­
methylanthranilate
by
gastric
intubation.
No
adverse
toxic
effects
uere
aoted.

Reierence
No.:
23
(Gaunt,
I
.F.,
et
a
l
.
FoodCosmet.
Toxicol.
1970)

Species:
Rat
(CFE).
.Route:
Oral­
No./
Croup:
30
(15H
6
15F)
Vehicle:
Diet
Duration:
90
days
Control
:
Ciet
alone
MetSyl
N­
met?
tiylanthrar.
ilate
w
a
s
added
t
o
t
h
e
d
i
e
t
of
rats
at
levels
of
either
300,
l?
5U
or
3600
ppa
(apprOXimiitely
equivalent
t
o
a
J8ily
intake
of
15,
bo
or
180
mgjkg)
f
o
r
90
days.
Measurements
of
bodyweight
urd
food
intakes
were
recorded
regularly
and
no
significant
differences
between
t
e
s
t
and
con­
t
r
o
l
s
were
noted.
ilematological
examination
revealed
a
slight
but
significant
leukocytopenia
and
anemia
i
n
animals
receiving
1200and
3600
ppo
a
t
week
6,
!.
ut
not
at
90
days.
The
r
e
s
u
l
t
s
o
f
u
r
i
n
a
l
y
s
i
s
conducted
a
t
week
4
and
again
with
blood­
Chemical
deteminations
8t
week
13
were
within
the
normal
limits.
Measurements
of
organ
weights
revealed
a
statistically
significant
increase
i
n
the
kidaey
weights
i
n
aales
receiving
1200
and
3600
ppm.
Gross
findings
showed
no
evldencer
of
agent­
nlated
lesions.

,,

'
I
Reference
No.
:
23
(Gaunt,
I.
F.,
e
t
81.
Food
Cosmet.
Toxicol,
1970)

Species:
Rat
(CFE)
Route:
Oral
.
No./
Group:
4
(F)
Vehicl­
e:
Weat
'

Duration:
Acutr
(7
day
observ8tior,)
Cantml:
Not
Specified
The
Or81
LD
f
o
r
methyl
N­
mtthylanthranilate
in
fasted
rats
w
a
s
repottcd
t
o
be
2.25­
3.38
39kg.
Symptoms
of
intoxication
included
increased
exploratory
behavior
for
15
minutes,
decreased
motcr,
activity
at
4­
24
hours,
non­
responsive­
ness
to
painful
stimuli,
piloerection,
t~
loody
nasal
discharge,
loss
of
con­
sciousness
and
death
within
18­
48
hours.
Cross
cxaminaticu
raveold
3
slight
reddening
o
f
pulmonary
tissue.
..
I
Saae
of
Substance:

Reference
No.:
40
(%
dyke,
D.
L.
J.
Fcod
Cosmet.
Toxicol.
1975)

Species:
Rat
R
c
u
t
e
:
C
r
a
l
So./
Croup:
Sot
Specified
Vehicle:
No:
Specified
Puration:
Acuto
Control
:
Not
Specified
3.7
ml/
kg.
The
oral
LOso
f
o
r
methyl
N­
methylanthrPnilatt
in
r
a
t
s
v
u
'
e
y
o
r
t
d
t
o
be
Reference
No.:
44
(her,

Species:
Rat
(FDRL)
No./
Group:
30
(ISM
4
1SF)
&ration
:
90
days
B.
L.,
e
t
01.
Fwd
Cos
met
.

Rout
e
:
Vehicle:
Control:
foxicol.
.196S)

Oral
Diet
Net
alone
Nethyl
N­
methylurthrrnilate
v
u
added
t
o
t
h
e
d
i
e
t
of
r
a
t
s
a
t
:cvels
cal­
c
u
l
a
t
e
d
t
o
r
e
s
u
l
t
i
n
approximate
d
r
i
l
y
i
n
t
a
k
e
s
o
f
19.9.
mg/
kg
(M)
a
d
22.2
mg/
kg
(F)
for
90
days.
Neasurunents
of
bodyweight
and
food
consumption
uere
recorded
regularly
and
nc
significant
differences
between
test
and
control
r
a
t
s
were
seen.
Hematological
examinations
and
blood­
chemical
determinations
.conducted
a
t
weeks
6
and
12
revealed
noma1
values.
Liver
and
kidney
weights
at
autopsy
were
n
o
r
u
l
,
and
.hirtopCthology
revealed
no
dose­
related
lesions.
I'
I
I
SECTION
1V.
A.

~r
b
1
e
IV­
I
sumaarites
references
in
the
scientific
literature
t
o
the
natural
occurrenee
of
­the
substuncas
i
n
foods.
For
each
substance
which
YU
found
t
o
occur
i
n
foods,
the
tab10
indica%­
the
food
source,
.h
o
t
h
o
d
of
detoctiar,
level
of
concentratioa
i
n
t
h
o
food
(tf
ruportodl
,
and
other
portinent
coIMllts.
Ihe
nfonnco
nudor
folloring
oath
entzy
i
n
tho
tablo
raforr
t
o
the
bibliography
nubeg
usignod
t
o
the
article
i
n
which
that
d8ta
m
u
.
fomd.
Copies
of
the
articlos
citod
w
i
t
h
English
tru~
shtiont'
Of
foreign
lmgurgo
8fliClM
are
in­
cluded
i
n
Voluw
111.
As
previously
s
t
a
t
e
d
i
n
S
o
c
t
i
o
a
I
,
tho
natural
occulldnce
of
a
substance
in
fwds
indicrtes
that
tho
substurco
has
bean
cogmod
b)
humans
for
centuries.
Where
q
u
m
t
i
t
a
t
t
n
d
a
t
a
on
t
h
o
hV.
1
of
COnCentr8tia1
are
awilrble,
tho
level
of
CoWrnptiOU
CIO
bo
~~t
i
m
t
e
d
;
hheta
80
qrturtit8tiVO
&t8
UOZG
f
m
d
,
the
o
c
c
m
n
c
a
of
a
substance
in
a
variety,
of
foods
usually
indicates
that
mom
than
.
traco
q&
titios
have
bem
eonsrrwd.

tho~
flavor
indusuy.
Altho­
the
tables
do'not
necessuily
represent
8
em­
plcte
l
i
s
t
i
n
g
of
a
l
l
references
t
o
n
a
t
u
r
a
l
ocrurrencs,
which
e
x
i
s
t
i
n
t
h
e
l
i
t
e
r
a
­
ture,
8n
a
t
t
e
q
t
u8s
asdo
t
o
include
roferenccr
t
o
n
a
t
u
r
r
l
occurrence
of
each
substmco
in
u
.m
n
y
d
i
f
f
e
m
t
food
sources
u
possible.
Tho
d
a
t
~
i
n
t
h
i
s
t
a
b
l
e
wero
gathered
primarily
f
r
o
m
inforution
pmvided
by
When
it
is
s
t
a
t
e
d
t
h
r
t
a
substance
wu
detected
by
isolation,
t5is
means
t
h
r
t
the
rCtw1
SUbStWC,.
Or
8
S
i
q
i
e
derivrtive,
W
U
isolrted
i
n
8
relrtivcly
pure
torr.
TIm
torr
"chdcaily
characterized"
mew
that
identificr;
ion
YU
obtdnod
by
UI
`rppropri@
to
dtcdcal
dotaction
method.
For
conveni~
cr
in
pre­
paring
tho
tablo,
8
e
t
h
e
using
physical
wuu
of
deteectim,
such
u
mixed
mltiw
­points,
wen
included
under
t
h
i
s
heading.

39
.
..
.
Abbreviations
Used
In
l
h
i
s
fable
i
:
CLC
gas
liquid
chromatography
IR
infrared
spectroscopy
W
=
ultraviolet
spectroscopy
HHR
*
nuclear
magnetic
resonance
spettmcopy
TLC
thin
layer
fixautography
PC
.=
paper
chromatography
E6
=
mass
spectroscopy
40
1
u
?
8
P
0
I
0
R
."
c4
a
._
U
c
u
U
­0
u
e
o
*c
c
o
o
n
"
C
0
.­.

x
L
!

h
*­
0
L.
0
O
n
L.
6
B­
ul
f
4
1
.

,
I
42
.

t
SECTIQN
I
V.
6.

.
USAGE
EVES
FRM
SURVEYS
BY
fE)
lrr
Ah0
NU
TABLE
IV­
2
­

This
tablo
is
primrily
a
c
o
l
l
o
c
t
i
~
of
the
most
ponincnt
results
published
in
a
series
o
f
tables
by
tho
Nationul
&a&
q
of
Sciencm
md
avail.
blo
frorr
the
National
Technics1
Infomation
Semi.­.
T
h
e
tables
contain
results
calculated
from
data
obtained
f
r
o
m
several
sources,
princi­
pally
f
r
o
m
tho
1970­
71
surveys
of
usage
conducted
by
the
Flavor
and
Extract
"frctumrr'
Associatioa
(=.
MI
m
d
t
h
e
Nation81
Ac­
of
Scioncos,
Utimal
Rosoarch
Council
(WINRC).
The
mothods
of
compiletion
md
coqutation
aro
oxplainad
in
dotail
i
n
tho
Addendum
t
o
Tab12
IV­
2
b
d
in
sources
roforrtd
t
o
i
n
that
Addendum.

'Ihcso
documents
8%.
8vril.
hlo
f
r
o
m
the
National
Tochnic8l
Infom8tion
Sewice.
..

Tho
notes
below
uhich
explain
the
entries
i
n
this
t
a
b
l
o
r
e
f
e
r
t
o
tho
~

Not..
1
­
This
colran
lists
she
nuL.
of
tho
substmco
and
i
n
brackets,
srople
page
that
follows.

tho
nubor­
by
dich
t
h
o
sub,
stmco
i
s
l
i
s
t
e
d
in
all­
othor
t&
hs
in
this
rrviw.
Bonoatb
tho
n
u
o
i
s
tho
t
o
t
a
l
urnwl
poundage
us&
in
flavorr.
u
reprtod
on
both
tho
FEN4
survey
ud
the
NM/
NRt
survey.
Tho
NECSINRC
sub­
c
o
d
t
t
e
e
o
s
t
.i
r
t
8
d
thrt
tho
datr
reported
ropnsentcd
between
608
ud
I
O
t
of
.tho
actus1
pomd8ge
rdd8d
t
o
tho
nition's
,food
supply
8nausllym
To
astipate
per
argita
daily
intako
in
mg
for
any
r3st.
nce.
mw
the
docinl
point
of
the
pom&
p
figure
five
places
t
o
the
left.
TU
trtrrr
into
r
c
c
o
~t
conwrsian
factors
and
rn
estimated
b
o
%
coverage.
Ihm,
19
annual
­age
of
73,300
l
b
s
f
o
r
=thy1
m
t
h
m
i
l
a
t
e
is
r
p
p
r
~x
l
u
t
e
l
y
equivalent
t
o
a
daily
p
8
r
+tu
intake
of
0.73300
ag.
.
.

Note
2
­
Tho
food
crtegorisr
for
which
.usage
u
l
t
.
reported
aro
listed
Noto
3
­
Tho
n\
Pbcr
of
fim
raportinp
usage
in
ea&
foal
eitaaozy
it
in
this
colraa.
Sea
thq
Adden­
t
o
t
h
i
s
table
for
further
eqlurrtiarr.
1
..

l
i
s
t
e
d
hem..
An
asterisk
(*)
indicates
3
OY
f­
or
firrr
reported
us8gom
i
"

Note
4
­
These
columns
l
i
s
t
:he
weighted
means
of
t
k
*usual
a
d
r
u
x
i
u
levels
of
use
reported
on
the
survey
of
usage.
'
Usage
'levels
were
reighted
wag*
level
reported
by
each
f
i
m
on
a
given
substance
w
1
s
muJ.
tiplied­
by
a
ratio
ca:
culated
by
dividing
the
t
o
t
a
l
pounds
reportad
by
the
firm
by
the
total
reported
by
a
i
l
fims.
n
o
levels
thus
weighted
uere
s~
lppyd
t
o
ob­
t
a
i
n
tho
weighted
mans.
"occoadiny
t
o
t
h
e
annual
poundage
reported
as
being
used
i
n
foods.
Each
Note
5
­
This
lists
the
age
gmup
for
uhich
food
consumption
data
were
Note
6
­
These
colums
list
the
possible
urd
potential
intakes
of
the
wad
t
o
calculate
tho
possible
drily
intake
values.

designnrrrd
substance
for.
each
category
by
aye
group.
The
t
o
t
a
l
f
o
r
a
l
l
categories
­is
presented
a
t
the
top
o
f
each
c
o
l
m
on
a
level
w
i
t
f
r
the
nue
of
the
substance.
These
intake
lovels
WIT
calculated,
a
t
described
in
tho
Addenb,
from
usage
lavolt
and
food
COnSullptim
data
obtained
h
m
surveys
conductod
by
the
Ynrket
Research
Corporation
of
h
r
i
c
a
and
United
States
Deputnent
of
Agriculture
and
are
highly
inflated
as
8
r
e
s
u
l
t
of
the
m
l
t
i
p
l
e
exaggercrtions
b
u
i
l
t
into
the
calculations.
first;
the
(wslmrption
is
made
,,.~..
..
\:
~i?
4
that
if
a
substance
i
s
used
i
n
one
food
i
n
I
cotegozy,
it
is
used
i
n
a
l
l
lurri
foods
in
that
categozy.
.
For
cxnqle,
if
one
mvfacturer
reported
the
w
e
I
YU
i
n
a
l
l
bread,
cakes,
cookios,
etc.
in
the
baked
goods
crtegozy
a
t
the
,
of
a
substance
i
n
cheese
blintzs,
i
t
would
be
assumed
t
h
a
t
the
,substance
s
a
n
lovel.
Secondly,
i
t
i
s
usurred
that
the
8verrpc
person
eats
food
fmm
.;
1
food
categories
rvry
diy.
This
would
result
.in
a'
d
a
i
l
y
i
n
t
&
of
oyer
.
.

5,000
Calories,
obviously
an
exrggoration.

of
thr
usage
data
and
the
inherent
ikccurrcies
that
exist
in
the.
tdle.
To
fully
understand
and
apprtcirte
the
finitations
of
this
data,
thd
Addendum
should
be
nvisved
c
u
e
f
l
l
l
y
,
I
Iho
foregoing
discussion.
is
only
a
brief
sumnary
of
t
h
t
significance
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i
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I
fie
purpose
of
t
h
i
s
ciiscussion
iS
to
revlev
in
8
cursory,
m
n
­t
s
h
i
c
r
l
way,
tire
background,
purpose,
VIIUCS,
and
l
i
n
i
t
a
t
i
m
s
of
a
w
j
o
r
e
f
f
o
r
t
to
gather
data
relating
to
the
w
e
of
food
ingredients
wtrich
are
generally
recognized
as.
safe
('*
GUS*
').
Not
only
t
h
i
s
sumnary
discussion.
but
the
f
u
l
l
report­
should
'be
read
carefully
by
those
intending
to
d
e
.any
use
of
the
data.

1.
Background
and
Purpose
A
review
of
the
safety
and
appropriate
classification
of
substances
'
~e
n
e
r
a
l
l
y
flecognized
2s
fafe"
.(
'',,,,
'I
('I
for
their
intended
use
in
food,
re@
res
(1)
identifying
each
substance,
A
d
(21
gathering
for
each
substance
all
available
data
on
its
toxicity
and
occurrence
in
food:.
goth
kinds
of
dot8
are
necessary,
since
the
safety
of
8
substance
is
detemined
3y
consid­
ering
both
its
toxicity­
how
little
or
grtat.
its
inhertnt
capacity,
to
cause
h
m
­a
n
d
the
amount
of
the
substance
t
o
vhich)
uc
are
exposed.
Determining
exposwe
requires
two
primry
c.
ypes
of
informtion:
First,
,

hos
arch
of
each
GRAS
substpnca
is
wed
in
each
clearly
defined
food
category?
Second,
how
mch
is
eaten
of
each
food
category
in
chich
thc
CFUS
s&
stance
is
wed?
­

2.
Survey
Procedun
AS
part
o
i
thm
GRAS
review,
a
special
subcorittcc
of
the
htion.
1
Academy
'of
Sciences
conducted
a
.survey
designed
t
o
provide
informtion
an
the
probable
and
possible
intake
of
such
food
ingredients.
The
subcornittee
US&
both
its
own
f
a
c
i
l
i
t
i
e
s
and
those
of
sever81
ifkiustxy
groups..
nit
took
the
forr
of
a
co.
plex,
comprehensive
survey
of
the
chemical
and
food
processing
industries
t
o
deterrino
the
exact
identity
+nd
specifications
of
the
subttances
aided
to
various
foods.
The
suney
obtained
confidential
inforntion
fro.
each
participating
firm
on
erch
CRM
ingredient
used,
the
f
a
d
categories
in
which
~

used,
and
the
usual
a
d
8aximm
levels
of
use
in
e8ch
food
category.
Respoading
firms
kdic8tcd
the
importance
they
attached
t
o
each
ingredient,
the
errliest
date
of
use,
and
the
pomd8ge
of
the
ingredients
they
eolritted
to
the
food
(')
See
Glossary
of
Tcru
f
o
r
d
e
f
i
n
i
t
i
o
n
of
underlined
words.
­2­

supply.
The
survey'provlded
much
additional
data
or1
toxicology,
natural
Occurrence,
and
other
information
relevant
to
the
evaluation
o
f
safety.
n,
e
Comaittee
made
use
of
other
surveys
bhich
covered
specis1i:
ed
ingredient
areas.,
such
as
flavors,
the
ingredienrs
used
in
chewing
gwn
and
certain
candies,
and
brewed
oevtrayes.
A
separate
survey
covered
infant
foods.
Thew
usage
data
vera
reviewed
for
consistency,
accuracy,
and
appropriate­
ness,
referred
t
o
the
originators
for
correction
or
:orfirmatier,
*here
necessary,
tabulated
i
n
a
way
t
o
preserve
confidentiality
and
collated
to
provide
informa­
tion
of
more
direct
value
in
safety
evaluation.
In
order
that
information
on
the
amounts
of
each
ingredient
used
in
food
be
applied
to
the
­estimation
of
possible
human
intake,
the
Committee
obtained
from
t
h
e
Market
Research
Corporation
of
Anri'ca
(bRCA)
data
from
a
survey
of
frquency
of
consumption
of
a
l
l
food
i
t
e
m
eaten
In
or
away
froa
home.
nese
data
were
r5classified
to
f
i
t
wizhin
the
food
categories
crployed
in
the
Academy
survey
of
ingredient
usage
by
the
food
industry.
Data
on
portion
size
obtained
from
a
US9A
survey
were
employed
t
o
complete
t
h
e
c3lculation
of
..

"average"
and
"high"
coalsumption
of
each
fooci
cakcgory­
used
in
the
survey.
me
data
finally
present.
ed
on
each
GUS
substance
included
weighted
Lcans
of
the
wuai
and
maximm
&e
in
each
food
category,
potential
"average,"
and
high
intakes
from
each
focd
category
and
the
tot81
diet
by
age,
total
yorradage
used
in
the
food
supply,
technical
effects
for
which
used,
and
industrial
importance.

3.
Scope
of
the
Survey
Within
the
limits
of
8vailible
tire
and
funds,
the
Comittee
concludcd
t
h
a
t
t
h
e
survey
achieved
its
intended
purpose.
Consistent
and
apparently
valid
information
was
obtained
from
firms
responsible
f
o
r
60
to
70
percent
of
the
nat.
ion's
processed
food
supply
(andeven
a
higher
percentqe
for
flavors
utd
certain
specialty
foods:.

4.
Valuer
­
mere
s
w
e
y
s
provided
the
first
n
l
i
8
b
l
e
d
a
t
a
on
the
levels
and
r
b
c
r
of
use
of
oost
ingredients
on
the
original
FD4
'White:
Listn
and
those
used
by
prior
sanction.
I
t
generated
ve­
ry
useful
detailed
information,
including
re­
.

checks,
on
:he
highest
wages
of
ingredients,
and
the
foods
in
which
such
@".
,.

k
j"
­.
r*.+$
>,
high
levels
ocrwrred.
­

.?"­
'.
L.
63.

I
.

.I
I
I
Ir
­_
.
­
­,.
­

The
survey
shows
uhether
a,
ri
ingredient
was
used
by
m
y
f
i
m
s
o
r
only
a
few;
or
used
i
n
many
or
few
food
categories.
A
related
concluslon
to
which
the
surveys
point
is
the
familiar,
but
frequently
ignored,
need
to
achieve
variety
and
balance
in
and
t
o
avoid
an
excess
The
surveys
also
on
t
h
e
amounts
used
in
use
can
be
estimated.
is
an
extremely
useful
of
other
data.
the
diet
as
t
h
e
best
way
to
insure
adequate
nutrition
of
any
ingredient,
whether
naturally
occurring
or
added.
provided
data
on
the
importance
of
each
ingredient,
and
the
total
f&
d
supply
fromwhich
average
per
capita
While
not
directly
valuable
in
safety
evaluation,
this
bench
mark
against
which
to
compare
the
reasonableness
These
efforts
provided
inforubatio;
l
which
is
less
than
pcrfsct;
but
the
best
available
on
food
consumption
by
current
categories
of
processed
food
andby
age.
Furthermore,
these
new
results
2rovide
consumption
data
for
those
individuals
who,
"eating
in
a
rut,"
consume
a
?articular
food
category
more
often
than
90
percent
of
the
population.
The
proccdtxes
wed.
in
these
surveys
have
resulted
in
several
'benefits.
Among
these
are
a
consistent,
comprehensive
clas.
sification
of
"technical
effects"
for
which
GRAS
substances
and
food
additives
are
&.
xi,
and
the
,
definition
of
food
categories
mre
adapted
t
o
modern
processed
foods
than
the
traditional
nutritional.
or
commodity
classifications.
The
results
of
the
survey
will
be
of
substantial
value
in
providing
for
industry
guides
t
o
good
manur'act\
uir,
g
practice
i
n
ingredient
usage.
Finally,
it
is
c
l
e
a
r
t
h
a
t
t
h
i
s
aust
be'a
recurrent,
orderly
process,
and
these
sur­

veys
have
set
a
useful
pattern
for
the
future.
.
,

5.
Limitations,
Cautions,
and
Restraints
Though
valuable,
these
survey
data
a
r
e
cocsplex
and
subject
t
o
a
number
of
uncertainties.
To
mderstand
their
values
and
'limitations
and
avoid
mi$­

,interpretation
and
invalid
conclusions,
those
who
wish
to
use
the
information
in
the
exhibits
or
the
tables
should
first
read
thoroughly
the
report
urd
exhibits
prepared
by
the
Committee.
The
most
inportant
caution
t
o
be
observed
is
t
o
k
g
n
i
z
e
t
h
e
cumulative
effect
of
the
several
conservative
assumptions
mdt
in
the
coqilation
of
t
h
e
­
data.
The
result
is
a
substantial
over­
estimate
of
possible
intake.
The
reported
data
indicate
usage
in
some
food­­
rarely
a11
foods­­
in
a
food
cotegQry.

i,,
'­

I
64
.
­4­

Such
d
a
t
a
a
r
e
v
a
l
i
d
o
n
l
y
when
and
i
f
the
ingredient
is
acxual!
y
used
i
n
­
t
h
a
t
category;
yet
the
calculations
conservatively
assume
usage
app;
iet
to
a
l
l
foods
in
a'
category.
Furthermore,
an
ingredient
may
be
partially
lost
in
processing
due
to
evaporation,
washing
away,
or
other
causes.

The
survey
can
only
indicate
indirectly
whether
a
particular
ingredient
is
used
i
n
many
foods
within
a
given
food
catsgory,
in
uhich
case
there
would
usually
be
mjny
responses
on
that
ingredient,
or
is
*sed
only
in
a
feu
foods
(usually
specialty
foods).
Fdr
example,
the
use
of
sulfur
dioxide
is
conftned
t
o
sow
d
r
i
e
d
f
r
u
i
t
s
.
I
t
is
not
used
in
fresh,
canned,
o
r
f
r
o
z
e
n
f
r
u
i
t
s
­or
juices.
Caramel
is
used
o
n
l
y
i
n
some
gravies,
not
in
cream
and
other
sauces;
yet
its
level
qf
use,
only
in
those
fords
in
which
it
is
used,
could
be
taken
t
o
a
p
p
l
y
to
the
whole
sauce
and
gravy
category.
Such
e
r
r
o
r
s
should
be
avoided.
Uhether
an
ingredient
is
used
i
n
many
or
feu
foods
within
a
category
depends
upon
t
h
e
a
v
a
i
l
a
b
i
l
i
t
y
of
alternative
ingredients.
NO,
or
few,
a
l
t
e
r
­
natives
mean
wide
use.
Many
alternatives
rean
g
r
e
a
t
l
y
r
e
s
t
r
i
c
t
e
d
we,
although
t
h
i
s
is
not
clearly
apparent
since
the
reported
level
of
use
sppears
:o
apply
t
o
a
l
l
f
c
o
d
s
i
n
a
category
e
v
a
though
t
h
i
s
would
seldom,
if
ever,
be
the
east.

A
d
i
s
t
i
i
l
c
t
i
o
n
must
be
drawn
between
those
tables
based
on
the
frcquency
with
which
foods
are
consumed
by
the
average
of
the
total
populatiar
('Total
Sample")
as
compared
with
those
tables
("
Eaters­
o
n
)
which
are
based
solely
on
those
who
consume
the
food
a
t
some
time
withir.
the
suwey
period.
The
l
a
t
t
e
r
d
i
s
r
e
g
a
r
d
s
t
h
o
s
e
i
n
the
population
who
d
i
d
n
o
t
consume
the
food
during
t
h
a
t
p
e
r
i
o
d
.
(Consumption
of
any
one
food
category
reduces
the
consumption
of
other
food
categories
due
t
o
r
e
s
t
r
i
c
t
i
o
n
s
rn
t
o
t
a
l
a
l
o
t
i
e
intakt.)
­Thus,
levels
of
conslmtption
based
on
"Eaters
Only"
can
be
valid
only
for
s
i
n
g
l
e
o
r
a
few
food
categories.
"Eaters
Only"
intake
figures
cannot
b,
e
added
across
a
l
l
categories,
since
no
one
eats
everything.
Such
an
assumption
rould
r
e
s
u
l
t
in
a
wholly
impossible
total
calorie
intake.
n
e
same
l
o
g
i
c
makes
the
levels
in
t
h
e
"E
a
t
e
r
s
W
y
"
t
a
b
l
e
s
n
o
t
u
s
u
a
l
l
y
a
p
p
l
i
c
a
b
l
e
even
to
a
l
l
foods
within
a
single
food
category.
From
t
h
e
l
e
v
e
l
o
f
each
ingredient
used
in
at
least
.some
foods
within
each
food­
category,
the
frequency
with
which
each
category
is
consumed,
and
the
portion
sire
when
consumed,
the
tables
calculate
the
possible
d
a
i
l
y
i
n
t
a
k
e
of
the
ingredient
through
each
food
category.
The
sua
of
such
­possible
intakes
fTW
a11
food
categories
Frovides
an
estimate
of
possible
­

65
.
..
t
o
t
a
l
d
a
i
l
y
intake.
For
the
reasons
just
discussed,
t
h
i
s
estimate
is
u
s
u
l
l
y
hizhly
inflated.
The
extent
of
such
inflation
can
be
gathcred
by
comparing
that
figure
of
possible
d
a
i
l
y
intake
with
the­
per
capita
daily
intake
obtarned
fTOP
the
.poundage
figures
committed
to
food,
the
coqletencss
factor
of
t.
le
survey
(a
t
l
e
a
s
t
60
percent),
and
the
population.")
Tht
ratio
is
low
i
.e
.,
­

.
tha
exaggeration
of
possible
daily
intake
compared
with
the
per
capita
is
satall)
uhcre
an
additive
is
broadly
used
and
where
there
are
feu
alternatives.

The
r
a
t
i
o
(the
exaggeration)
is
high
where
an
additive
is
u
s
e
d
.
infrequently,
as
only
in
specialty
foods,
o
r
where
there
are
many
alternatives
for
obtaining
the
same
technical
effect.
h
e
cannot
take
the
possible
or
­potential
daily
intake,
even
khat
is
perhaps
misleadingly
called
the
flaverage,
l'
from
the
"total
sample"
table,
a
l
t
i
p
l
y
by
the
population
and
the
days
per
yeor,
and
expect
to
obtain
the
annual
food
us0
of
an
ingredient.
This
is
an
impermissible
us8
of
the
data
because
of
the
multiple
exaggerations
reviewed
above.
The
intake
tables
inevitably
reflect
s
a
c
p
o
i
n
t
a
t
the
upper
tail
of
the
distribution
curve.
From
thi's
it
follows
that
the
intake
values
labeled
"high"
or
'%
e.:,

high"
could
be
obtained
only
as
a
result
of
a
highly
improbable
combination
of
circumstances;
selection
fror
food
Category
of
arly
those
foods
contain­
ing
a
particular
ingredient,
no
­toga
or
loss
o
f
the
ingredient,
cons­­
tion
of
the
food
category
at
m
t
u
w
l
l
y
high
frequency
QI
high
levels
of
llsa
of
the
ingredient
or
both,
and
no
1st
of
alternative
ingredients.
Irrprobable
in
itself
for
one
fwd
categor/,
this
is
virtually
inpossible
for
more
than
a
feu
food
categories
simltaneowly.
Because
one
could
not
possibly
seek
a11
possible
technical
effects
in
.

a
single
hod,
and
because
of
the
multiple
alternatives
frequently
available,
one
cannot
add
a11
substances
used
in
any
f+
category
to
obtain
the
totof
amount
of
substances
potentially
occurring
a
t
one
time
in
any
t00d
or
category.

lack
s
t
a
t
i
s
t
i
c
a
l
s
o
l
i
d
i
t
y
and
refiect
very
limited,
therefore
mrnetim.
S
extreme,
we.
The
data
base
for
some
food
categories
and
Copulation
GOUPS
is
also
narrower
than
­the
i
d
u
l
.
The
figures
indicated
by
asterisk
t
o
signify
three
responses
or
less
This
is
the
NAS
11/#
2
ratio
in
Exhibit.
58.
....
..
......

"7'
'7
......

­0­

Is
is
clear
there
is
nc!
final
Lay
to
be
absolutely
certain
of
either
accuracy,
or
completeness
i
n
t
h
e
survey.
Overall,
the
survey
is
as
complete
as
uas
practical
because
of
the
diminishing
returns
€or
add'itional
effort
expended.

6
.
Qgortmities
Tkis
survey
points
out
the
desirability
of
having
(1)
better
food
eon­

sumption
rather
than
use
or
disappearance
data,
(2
)
more
exrensive
analysis
of
foods
to
establish
what
colPponents
remain
i
n
them
r
e
l
a
t
i
v
e
t
o
what
was
introduced,
(3)
determination
of
the
effects
of
preparatim
and
p
l
a
t
e
s
a
s
t
e
on
ingredient
loss,
(4)
better
understanding
of
'food
category
use
by
specific
population
groups,
(­
5)
estimates
o
f
intake
from
a
l
l
sources,
natural
and
added,
:
and
(6
)
future
surveys
t
o
r
e
f
i
n
e
amd
*date
these­
data,
The
present
inforiation
deserves
substantial
additional
analysis.
A
cursory
inspection
reveals
food
c
o
n
s
q
t
i
o
n
patterns
in
processed
foods
uhich
are
age­
dependent
o
r
o
f
n
u
t
r
i
t
i
o
n
a
l
i
n
t
e
r
e
s
t
.
I
t
seems
probable
t
h
a
t
t
h
e
.

ingredients
used
for
different
technical
effects
tend
to
group
within
definable
wage
and
t
o
t
a
l
a
n
n
u
l
poundage
levels.
As
examples,
salt
m
d
t
h
e
t
h
r
e
e
,
sueeten.
ers­­
sucrose,
dextrose,
and
corn
syrup­­
make
up
the
four
most
.u
s
e
d
GRAS
substaxes.
Flavors
constitute
the
bulk
of
those
as&
i
n
least
voluae.

A
better
appreciation
of
such
patterns
of
w
e
could
lead
to
an
aderstanding
.

of
where
it
would
be
desirable
from
e
i
t
h
e
r
.
an
economic
or
a
public
health
point
of
view
to
concentrate
research
on
new
alternatives.

67
(;
LOSSXRY
ACCEPTAPLE
DAILY
INTALE
.(
AD?)

Daily
dose
of
a
chemical
that
appears
to
bc
without
appreciable
risk
on
the
basis
of
a
l
l
the
facts
knom
a
t
thc
time.
"Without
appreciable
risk"
is
taken
t
o
mean
the
practical
certamty
t
h
a
t
injury
will
not
resuit
even
a
f
t
e
r
a
lifetime
of
exposure.

ANNUAL
POUNDAGE
The
t
o
t
a
l
number
of
pounds
used
per
year
by'each
participating
firm
or
ih
t
o
t
a
l
by
a
l
l
r
e
p
r
t
i
n
g
firms.
..

AVERAGE
DAILY
INTAKE
>

Estimated
amount
of
a
($
AS
substance
that
may
be
ingested
daily
through
consumption
of
foods
to
which
the
substance
has
been
added.

CONSWION
LEVEL
The
level,
i
n
grams
per
QY,
of
a
food
category
consumed
by
the
rveragh
person
("
total
sample")
or
consumed
only
by
those
who,
within
the
sur­
vey
period,
ate
food
from
that
category
at
least
once
("
eaters
only").

DAILY
INTAKE
See
CONSfp(
PTI0N
LEVEL.

FAO/
nm>
JOINT
EXPERT
'
C
W
f
l
T
E
E
A
committee
of
toxicologists
and
o
t
h
e
r
s
c
i
e
n
t
i
s
t
s
.appointed
for
their
individual
qualifications
and
not
representing
government
agencies
or
industries.
Brought
together
=der
the
joint
sponsorship
of
the
Food
and
Agriculture
'Organization
'and
the
World
Health
Organization
for
the
purpose
of
defining
specifications
for
food
additives,
appraising
their
safety­
in­
we,
and
setting
such
limitations
or
requirements
for
further
information
as
seem
appropriate
t
o
them.

FEMA
Flavor
and
Extract
Manufacturers'
Association.
..

FpW
GRAS
LIS
Set
GRAS
LIST.
..
a
.
f"
­8­

The
food
at
consumed
distinguished
fro8
intermtdiate
and
incoaplete
stages
of
processing
or
preparation,
such
as
dry
atixes,
concentrated
syrups,
ray
meat
or
vegetables,
etc.

FLAVORING
ADJUNCT
A
substance
which
does
not
itself
contributc
flavor,
but
which
is
used
in
association
u
i
t
h
flavoring
ingredients
t
o
improve
their
effectiveness
in
use.
This
includes
.solvents,
fixatives,
hntioxidantt,
etc.

FLAVORING
ADJWAHT
Synonym
for
FLAVORING
ADJUNCT
FUVORINC
INGREDIENT
oft&
simply
called
a
"flavor,"
is
any
substatce
added
to
food,
;Irugs,
or
other
products
taken
i
n
the
mouth,
the
clearly
predominant
purpose
and
effect
of
which
is
to
prqvide
a
particular
flaror
in
the
final
product.

"1.
a:
miterial
consisting
essentially
of
protein,
carbohydrate,
a
d
fat
used
in
the
body
of
an
organism
t
o
sustain
growth,
repair,
and
v
i
t
a
l
processes
and
t
o
furnish
energy;
also:
such
food
together
l
l
f
h
supple­
mentary
substances
(as
minerals,
vitamins,
and
condiments)"
This
is
used
here
in
the
sense
of
human
fodd.
In
the
law,
the
term
**
food**.
includes
beverages,
chewing
p,
and
the
componenu
of
ail
food
articles.

FOOD
ADDITIVE
General
Definition
~n
y
m
i
i
g
r
e
d
i
c
n
t
added
t
o
food,
or
residues
ofwhich
are
found
in
fwd,
resulting
fro.
its
use
to
achieve
a
particular
technical
effect.
(2)­
.
.
Leg81
Definition
­
**
Any
substance
the
intended
use
of
which
results
or
m
y
reasonably
bc
expected
t
o
r
e
s
u
l
t
r
d
i
r
e
c
t
l
y
o
r
i
n
d
i
r
e
c
t
l
y
,
in
its
becomiag
8
corponcnt
or
othemise
affecting
the
characteristics
of
YIY
focd
(including
my
substance
intended
f
o
r
use
in
producing,
manuf8cturing,
packing,
pro­
cessing,
preparing,
treating,
packaging,
transporting,
or
holding
f&;
and
bcluding
any
source
of
radiation
intended
for
any
such
use),
if
such
substance
fr­
not
generally
recognized,
among
experts
qualified
by
scientific
training
and
experience
t
o
evaluate
its
safety,
as
having
been
adequately
shown
through
sciemific
procedures
(or,
in
the
case
of
a
stance
used
in
food
p
r
i
o
r
t
o
January
1,
­1958,
through
either
s
c
i
a
i
t
i
f
i
c
procedures
OT
experience
based
on
co­
n
use
in
food)
t
o
be
safe
under
the
conditions
of
its
intended
use;
...
.'*

(l)
Webster'$
Seventh
New
Colltgiate
Dictionary,
p­
324.
Springfield,
Massachusetts:
G.
6
C.
Merriaa
Company
(1969).
­9­

AJI
act
of
Congress,
parsed
in
1958.
amending
the
federal
Food,
Drug,
acd
Cosmetic
Act.
The
amendment
established'
requirements
for
premarket
clear­
a?:=;
f
o
r
safety
and
functionality
o
f
.
a11
substances
intended
for
use
in
food
h
i
t
h
certain
exceptions.
The
exceptions
include
substances
that
qualifjcd
scientists
generally
recognize
as
safe,
other
substances
covered
by
a
'.
prior
sanction"
(q.
v.
1,
color
additlves,
and
pesticidal
residues,

f
OCQ
cATE03RY
One
of
34
categories
into
which
a
l
l
foods
haye
been
classified.
for
the
purpxe
of
t
h
i
s
survey.
Categories
consist
of
foods
closely
related
i
n
purpo,~,
structure,
process,
composition,
01
propcrties.

GOOD
MANUFACTJRXNG
PRACTICE
Procedures,
facilities,
quipment,
and
personnel
training,
whichtoken
togethex
and
properly
applied,
define
the
conditions
for
safe
and
effec­
tive
processing
of
food.

GRAS
The
acronym
for
*'generally
recognized
as
safe."
it
is
a
s
l
i
g
h
t
ad;
pta­
tion
and
incomplete
rendering
of
the
language
of
the
Food
Additives
hzdment
of
1958
to
the
Federal­
Food,
Drug,
and
Cosmetic
Act.
I
t
r
e
i
r
r
r
to
I*.
.
.
t
substance
.
.
.
generally
recognized,
among
experts
qualified
by
scientific
training
and
experience
to
evaluate
its
safety,
w
having
been
aCequacelY
shown
.
.
t
o
be
safe
mder
the
conditions
of
its
incended
use;
.
.
."
'17s
total,
much
longer
definition.
is
gwen
i
n
Section
201
of
the
Act.
(See
also
FOOO
ADOITIVE.)

GRAS
LIST
A
a&,
used,
but.
rnaccurate
term
f
o
r
all
substances
used
in
food
an
the
basis
that
they
are
GRAS,
rather
than
regulated
additives.
There
has
Tot
been
.any
single
"GRAS
List."
The
FDA
published
two
intentionally
incoaplete
lists
('White
Lists'3;
the
flavor
and
chewing
gum
industries
pub1
i
s
h
4
8
series
of
lists
(the
"FEMA
GX4S
?.
ist*
'),
the
brewing
industry
pmpared
a
coqrehensive
unpublished
list,
the
FDA
presmed
some
sub­
stances
t
o
be
CRAS
without
publication
urd
issued
"no
objection
letters,"
usually
tmpub1ishcd.
m
feet
other
substances,
and
thert
undoubtedly
were,
as
the
law
allows,
some
private,
rnputlished
determinations
that
a
use
of
a
particular
substance
was
WS.
Aside
from
these
groups,
USDA
and
FDA
had
i
n
earlier
y
e
n
s
approved
Dy
"prior
sanction"
a
large
number
of
substances
not
necessarily
covered
by
thr;
e
l
a
t
e
r
GUS
actions,

IMPORTANCE
Commercial
importance
in
the
sense
o
f
how
unique
are
the
properties
of
the
ingredient,
and
how
easily
can
it
be
replaced
by
another
ingredient
or
change
of
process.
\
......
I
INGRED
I
ENT
A
component
Or
constituent
3f
food.
The
term
is
usualiy
U
J
~
in
the
sense
of
an
ingredient
vhich
is
mtentionally
made
a
part
of
food.
.

Cozstituent
often
mans
a
naturally
cxcurrkg
component.

WIMJN
USAGE
LEVEL
Highest
level
used
in
any
product.

WEA?
'
INSPECTION
ACT
kr
act
of
Congress
originally
passCp
in
1907,
amended
and
extend4
to
permit
continuous
inspection
of
neat
slaughtering.
and
processing
est&­
l
i
s
b
e
n
t
s
by
employees
of
the
Dcpar­
tmnt
of
Agriculture.

HEDUN
USAGE
LEVEL
The
usage
level
of
a
particular
ingredient
in
a
food
categor),
uhich
f
a
l
l
s
in
tho
middle
of
the
reported
uses;
i.
e..
half
the
wes
are
reported
a
t
a
lower
level
and
half
the
uses,
a
t
a
higher
level..
.
"

NAS
,.

National
Academy
of
Sciences.

NDN­
USER
FIRMS
Firms
uhich.
mnufacture
or
distribute,
but
do
not
cornit
the
kyr&
ieat
t
o
food.

Letters
issued
by
FDA
after
1958,
by
which
FDA
indicated
that
there
would
be
no
objection
posed
at
the
ti­
of
the
l
e
t
t
e
r
to
the
w
e
t;
food
of
the
ingredient
or
ingredients
named
therein
in
accordance
­
.
with
the.
uses
outlind.

Offensive
to
the
sense
of
tsttt
or
me11
and
therefore
inedible;
usually
usel
t
o
refer
to
1
food
which
is
spoiled
or
overflavord.
..

PER
CAPITA
DAILY
INTAKE
An
average
figure
derived
by
simply
dividing
annual
national
food
use
of
an
ingredient
by
the
population
and
days
in
a
year.

PORTION
SIZE
A
figure
derived
from
estiuutes
of
nean
food
consumption
using
"total
­le'*
(see
def'nition
o
f
CONSUMPTIC3
LEVEL)
and
average
food
additive
use
for
each
foot
category,
described
in
Tables
i3A
and
B
IS
"aver:
lge
intake
level.
''
Bccause
or'
a
series
of
conservative
assumptions
(see,
discussion
in
report),
t
h
i
s
reprtsents
J
pgssible
intake
that
would
only
infrquently
be
achieved.

POTENTIAL
MILY
INTAKE
A
figure
dcrivcd
from
estimates
or'
food
c
o
a
s
q
t
i
o
n
based
on
"eaters
only"
[see
definition
of
COPISUHMION
LEVEL),
or
the
90th
percartile
frequency
of
consurption,
or
higher
levels
of
hod
additive
use.,
or
some
co.
bination
of
these
for
each
food
category.
These
are
described
in
tat
tables
as
''High
A,"
''High
B,"
or
"Very
High,"
or
"Eaters
Only.**
In
addition
t
o
t!
w
conservative
assumptions
inherent
in
­the
calculation
.

intskes
mlikely
for
.ore
thar.
one
or
a
few
food
categories.

WL'TRY
PRODOCM
INSPECTIW
A
r
c
f
.
.
of
'possible"
if.
tOkCS,
these
additional
factors
=de
the
potential
1
This
­5
M
8
C
t
O
f
COngrCSS
passed
in
1957
t
o
provide
for
the
inspec­
tion
of
poultry
and
poultry
products
and
otherwise
rcgulatiag
the
pro­

.n.­"
cessing
and
distribution
of
such
articles
in
interstate
copmerce.
;=.><

Ld
f
\.,
PRIOR
SANCTION
Action
prior
to
19S8,
by
the
FDA
or
USDA.
to
p
e
d
t
8
substance
to
bo
usd
in
food
­der
the
Food,
Drug,
and
Cosme:
ic
Act,
the
Poultry
Products
Inspectiat
Act,
or
the
M
e
a
t
Inspection
Act.

SPECIALTT
FO0OS
Foods
of
iiritrd
appeal,
usually
to
particular
ethnic,
economic,
or
,
geographic
groups,
often
mote
expensive
and
limited_
in
distribution
and
conswption.­
Th.
tern
irplles
lack
of
broad
appeal
or
frquant
.

US..

The
rchitvicg
of
8
desired
characteristic
in
a
food
or
food
manufactw­
ing
process.
For
euPpla,
8
desired
nutritional
content
or.­
antioxidant
in
a
food
produet;
ws*
of
removal
f
r
o
m
a
baking
pan;
or
ease
of
extm­
sion
for
pasta
product.

see
cotlsw1oN
LEVEL.

Capacity
of
the
substance
to
produce
injury.
'&%
e
term
includes
capacity
t
o
induce
teratogenic,
mutagenic,
and
carcinogenic
effects.

72
­
12­

Refers
t
o
"Food
Intake
and
Nutritive
Value
of
Ilietr
of
h,
b
m
,
a
d
chihirea
in
t
h
e
United
States,
Spring,
1965,
'*
a
preliminaq
report
by'
the
Gaasuvr
and
Food
Economics
Research
Division,
b
r
i
e
u
l
t
u
r
i
l
&sar&
Service,
hit4
States
Department
of
Agriculture
(sea
Exhibit
328).
,.

USER
FIM
Nom1
or
average
concentration
roughly
weigh4
by
amur1
prodwt
VOIUC.

YEIQITU)
MEAN
USAGE
LEVEL
'The
average
level
a
t
which
a
p
r
t
i
c
u
l
a
r
ingredient,
if
it
is
~4
,
is
added
to
foods
within
a
fwd
category
weighted
in
proport'
&on
to
the
VOIWC
of
the
additive
used
by
each
firm;
e.
g.
,
a
firpl
Vhia
wed
a
mil
lion
porurb
of
an
additive
coats
in
the
average
one
Wrtd
t
i
a
c
t
=re
thrn
8
fim
which
uses
o
l
y
lO,,
OOO
pounds.
The
bibliograpky
presents
a
c
'
q
l
e
t
c
l
i
s
t
i
n
g
o
f
a
l
l
l
i
t
e
t
q
t
u
n
a
r
t
i
c
l
e
s
and
references
used
in
the
preparation
of
t
h
i
s
r
c
i
e
n
t
t
z
i
c
l
i
t
e
r
a
t
u
n
r
e
v
i
a
.
me
e
n
t
r
i
e
s
i
n
t
h
i
s
s
e
c
t
i
o
n
ire
l
i
s
t
e
d
i
n
alphaLetica1
order,
by
nane
a
f
the
principal
author:
The
bibliography
include
a
l
l
a
r
t
i
c
l
e
s
1cca:
lti
i
n
the
literatwe
sear&,
whether
o
r
n
o
t
i
n
f
o
n
n
a
t
i
m
f
r
o
m
those
arti:
lrss
w.
u
included
in
the
literature
review.
Those
references
denoted
by
a
tingle
asterisk
(*)
are
a
r
t
i
c
l
e
s
c
i
t
e
d
within
the
literature
review,
ad
references
denoted
with
a
double
asterisk
(0
0
)
are
a
r
t
i
c
l
e
s
cited
within
the
S
m
r
y
portion
of
the
review,
in
Section
I.
Y
d
u
m
e
t
11
and
111
contain
copies
of
a
l
l
c
i
t
e
d
a
r
t
i
c
l
e
s
,
inclvding
­.
.
.

English
trans!
o:
ions
o
f
f
o
r
e
i
k
language.
articles.

..
1.

2.

3.

4.

+*
5.

6.

7.

,.
+*
8.

'
9
,

10.

11.

**
12.

13.

+­
14.
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B.
D.,
A.
J
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Poyaton,
and
I.
0.
Rat.
1972.
Aromatic
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i
d
e
r
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Steric
hiadrance
to
hydrogen
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Auat.
3
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Cheg.
15(
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Arctander,
S.
1969.
Perfume
and
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chedcalr,
S.
Arctander,
Moatelair,
N.
J.
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Barbaa,
S.
I.,
N.
S.
Karavya
and
U.
S.
Ilifnavy.
1971.
Study
of
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peel
ailr
of
lemoa,
lima
and
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i
n
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Ip.
Prrfm.
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86:
53­
56.

Bar,
?.,
and
e'.
Gricpcatrog.
1967.
Vherc
&
rtaad
coacrrning
the
evalurtioa
of
flavoring
rubrtanccr
frcm
the
viewpoint
of
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Ktd.
Lruaehr.
6:
264­
251.

Brom,
P.
E.,
m
d
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X.
Price.
1956.
Quantitative
studirr
ah
metabolite~
of
tryptophaa
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t
h
e
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o
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a
t
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and
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i
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t
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l
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u
r
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e
t
i
t
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r
a
i
n
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i
l
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derrrd
bergamot
petitgrain
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3143.

Carcoavt,
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and
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of
­the
uma
of
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Helv.
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57(
25):
209­
211.

Charcoanct­
Earding,
f
.,
C.
E.
Dalgticrh,
and
A.
ileuberger.
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The
relation
bctwea
riboflavin
and
tryptophan
rctabolirr,
rtudied
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521.

D
i
C
i
a
c
w
,
A.
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Citrur
errmtial
oil;.
XXIX.
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hr­
n,
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17(
f1):
460,
462.

Drauert,
?.,
urd
A,
von
Rapp.
.l%
6.
The'
componcotr
of
rurtr
(grape
j
u
i
c
e
r
)
and
wincr.
VIf.
C
u
Cbra8tOgr8phiC
invtrtigatioa
of
a
r
a
a
t
i
c
r
u
b
r
t
u
~c
e
r
i
n
vi­*
and
tbeir
bi@
gencrir.
Vitir.
5:
3S1­
376
.

Drawert,
?.,
and
A.
voo
Uapp.
I%&
Car
chraatograghic
malyri8
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of
plant
araar.
I.
The
enrichment,
reparation,
and
identification
of
v
o
l
a
t
i
l
a
arm.
rubatancer
i
n
grape
mrtr
aid
vines.
Chromato­
g
r
q
i
d
&
1:
446­
457.

Ekman,.
B.,
and
J.
P.
Strabcck.
1%
9.
The
effect
of
some
rplitproductr
of
2,
.3'­
acotoluene
on
t
h
e
u
r
i
n
a
r
y
b
l
a
d
d
e
r
i
n
t
h
e
rat
and
t
h
e
i
r
excretion
at
wriour
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Acta
Pathol.
Xicrobial.
Seand.
26:
467­
471.

Sracntial
O
i
l
.Arroeiatioo
of
U.
S.
A.,
fnc.
1970.
MIA
rpteificatioor
and
rtandardr.
#cu
York,
Reu
York.

m,
1974.
S
c
i
e
n
t
i
f
i
c
l
i
t
e
r
a
t
u
r
e
r
e
v
i
e
v
of
aliphatic
pr,
imary
aIcoholr,
aldehyder,
aeidr
and
id
:era
i
n
f
l
a
v
o
r
usage.
Vol.
1­
VLI,
Publiahcd
by
0.
S.
Food
md
Drug
Admamiatration,
Yarhington,
D.
C.

7s
15
16
17.

18.

19.

20
21
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22
23
24.

25.

26
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270
28
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29
FEU
G
U
S
Subrtance8,
1965­
18.
Recent
progrerr
in
the
coaridtratioa
of
flavoring
ingredients
under
the
food
additives
onendment.
CRAS
rubrcancer.
A
series
of
9
articles
publirhed
i
a
Food
technol.
19(
2,
part
2):
151,
1965;
2
f
O
):?S
,
197C;
2
O
(5
i
:3
5
,
1972;
27(
1):
64,
1973,
27(
11):
50,
1973;
26(
9):
76,
1974;
29(
8):
70,
1975;
31(
1):
65,
1977;
32(
2):
60,
1976.

Fieldr,
E.
K.
1971.
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anthranilates.
U.
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3,625,389.

Food
m
d
Drug
Adminirtration.
1977.
Swmrry
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toxicity
&ti.
Unpublirhcd
Repoct.

Food
a
d
Drug
Research
Labotrtoticr,
Ine.
1963.
Hetabolic
fate
of
methyl
o­
methyl
anthraailatc.
No.
84919.
Unpublirhed
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Food
Citemicala
Coder,
2nd
ed.
1972.
C­
ittee
on
specifications.
Food
Chemicals
Codex
of
the
Camittee
of
Food
Protection,
Uation.
1
Research
Counc'il;
National
Academy
of
Sciences,
Yashington,
DC.
1039
p
p
o
Poreiaa
C­
pound
Hctrbolia
ia
Mmmrlr.
Vol.
I.
1970.
Vol.
11.
1972.
Vol.
111.
197s.
The
Chemical
Society.
Burlington
Roure.
London.

Puria,
t.
B.,
and
W.
Btl~
mer,
cd.­
197s.
Fcnaroli'r
handbook
of
fl.
vot
ingrtdieats.
2nd
ed.
Vol.
1­
11.
The
Chemical
Rubber
Company,
Cleveland,
Ohio.

Cabel,
L.
P.,
and
U,
R.
J.
Sirpron.
1972.
Dialkyl
­
cliaubrtitutcd
4
­
~~
hydroxyalkyl)
amino)­
quinazoline
nitrates.
U.
S.
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3,637,699.

Gaunt,
I,
l!.
.Sharratt,
P.
Crarro,
and
X..
Uri8br.
1970.
Acute
and
rbort­
term
toxicity
of
methyl­
U­
mcthpl
anthranilate
ia
rats.
rood
Cornet.
Toxieol.
8(
4):
359­
368.

Civaudan
Index,
1961.
Specifications
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ryntheticr
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d
isolate8
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431
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Hagan,
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0.
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W.
I.
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ll..
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Food
C08met.
toxicol.
5:
141­
157.
.

euet,
Eo
1968.
The
of
citrus
fruit
juicer.
Fruits.
U(
g):
493­
471.

76
**
X.

31.

32.

33,

**
34.

35.

36.

3?.

**
38.

**
39.

**
bo.

4'
41.

**
12.

**
03.

**
44.

b
S
e
**
66.
Ienner,
P.
X.,
E.
C.
88g.
n~
J
*
X.
Taylor,
E.
Lo
Cook,
and
0.
C.
Pitthugh.
1964.
rood
flavourings
aad
compouadr
of
r
e
l
a
t
e
d
s
t
r
u
c
t
u
~e
.
X.
.\
cute
oral
toxicity.
Food
Coract.
Toxicol.
2:
327­
343.

Kahn,
J.
H.
(Rcvicv).
1969.
Compound8
identified
i
n
whiskey,
vine,
i
n
d
beer:
a
tabulation.
3.
Assoc.
Off.
Agric.
Chca.
52(
6):
1166­
1178.

Kicchncr,
J
.
G.,
und
3.
M.
Hiller.
1953.
Volatile
oil
conatituentt
of
grapefruit
juice.
J.
Agric.
Food
Chem.
1(
7):
512­
518.

Kugler,
E.#
and
3.
Kovatr.
1963.
Information
w
Handarin
peel
o
i
l
(Citrus
rcciculata
Blanc0
or
Citrus
nobilir
var.
delicioza
Suinglc
"Xandarin").
Ilclv.
Chim.
Acta
46:
1480­
1513.

Longland,
R.
C.,
W.
P.
Shilling,
and
S.
D.
Gangolli.
1977.
The
hydro­,
lysis
of
­flavouring
esters
by
a
r
t
i
f
i
c
i
a
l
g
a
s
t
r
o
i
n
t
e
s
t
i
n
a
l
j
u
i
c
e
r
md
rat
risrue
preparation..
Toxicology.
8:
197­
204.

Warck
Index
­
An
encyclopedia
of
chemicals
a
d
drugs.
9th
edition.
1916,
Herck
aad.
Company,
Inc.
Rahvay,
R.
3.
1313
pp.
i
National
Ac8dcap
of
Scieacrs.
1970.
Evrlwtioc!
of
food
eheucalr.
Wa8hiagtoa,
D.
C.

National
Pomdrry
XIP.
14th
ad.
1975.
Aaerican
Phomaceutical
Asrociation.
Yashiagtou,
D.
C.
..

Opdykc,
D.
L.
J.
19fSa.
Xonographs
oa
fragrance
raw
rateriala.
a­
Eutyl
aathrMilrte.
lood
Coatcat.
Toxicol.
13:
727­
728,

Opdykc,
De
L.
J.
197Sb.
Mouographr
on
fragrance
raw
mat,
erialr.
Cinn.
ay1
anthraailate.
lo&
Comet.
Toxicol.
13:
731.

Opdyke,
b.
L.
3..
1975~.
Honogrrpha
oa
fragrance
mu
materials.
Dimethyl
aarhrmil8te.
Food
Comet.
Toxicol.
133791.

r
Opdyke,
0.
L.
J.
1976..
~oaographs
00
fragrance
rau
materiala.
C
i
a
l
y
l
aathrmihte.
Pood
Comet.
Toxicol.
14(
5):
459.

Opdyka,
D.
L.
J.
l976b.
l4onographa
021
fragrance
raw
materials.
Ethyl
mthr.
nilate.
rood
Comet.
Toxicol.
14:
7f9.

Opdyke,
D.
L.
J.
1976~.
Monographs
on'frrgrancc
raw
materials.
Phenyl­
ethyl
urthtaaitrta.
Food
Comet
*
Toxicol.
:&
:831.

Oser,
B.
L.,
S.
Carson,
and
X.
Osct.
1365.
Toxicological
teet8
06
flavouring
aatfer8.
Food
Cosmet.
Toxicol.
3:
563­
369.

'Oset,
B.
L.,
a
d
It.
L.
8.11.
1977,
Criteria
employed­
by
tbe
.
.

Expert
P,
aael
of
P.
t.
KA*
for
the
GRAS
evaluatioa
of
flavorizq
,rubstancta.
h
o
d
Comet.
Toxicol.
15:
457­
466.

pelli­,
b.,
lt.
Longland,
n.
Dufltp,
and
S.
D.
C;
n~
o;
1.5,­
*A
.
study
of
intestinn1
abrorptios
of
four
flavouring
caters
in
the
guinea
pig.
ToxicologJl,
(s
u
h
i
t
t
t
d
f
o
r
p
u
b
l
i
c
a
t
i
o
n
).

77
­.

*.
**
48.
Registry
of
Toxic
Effects
of
Chemical
SubstJnces.
1977.
U.
S.
Dept.
of
Heaith,
Educ8tion
and
Welfare.
Public
Health
Service
Center
f
o
r
.­

Disease
C
o
n
t
r
o
l
,
N
a
t
i
o
n
a
l
I
n
s
t
i
t
u
t
e
for
Occupatimal
Safety
a
d
Health,
Rackville,
Maryland.
1296
pp.

49.
Roger,
N.
F.
1961.
The
r
e
c
o
v
e
r
y
o
f
m
e
t
h
y
l
a
n
t
h
r
t
n
i
l
a
t
e
i
n
concord
grape
1
essence.
Food
Technol,
13(
6):
309­
314.
­­
.

..
SECI'ION
V
I
.

DATA
GUIDE
The
following
guide
provides
a
slfmrmy
each
substance
i
n
t
h
i
s
S
c
i
e
n
t
i
f
i
c
­
Literat.­
of
various
data
included
for
Review.

The
principal
name
and
FEMA
number
of
the
substance
is
foliowed
by
its
synonyms.
?he
Code
of
Federal
Regulations
(CFR)
refereace
is
given
for
those
sub­

stances
publibhed
by
the
Food
and
Drug
Administration.

?%
e
range
of
average
usual
and
average
maximum
usage
levels
of
the
sub­

stance
is
shown
next,
as
u
e
i
l
as
annual
volumc
i
n
pounds.
These
data
are
taken
from
tho
1970­
71
NAS
and
FEW
surveys.
Tfie
pur
cupitq
daily
intakes
are
calculated
as
described
i
n
t
h
e
introduction
t
o
Table
IV­
2
froa
the
annul
MlUIDU.

I
f
the
substance
has
been
found
as
a
natural
component
o
f
food,
this
is
noted
a
f
t
e
r
the
=age
data,
w
i
t
h
the
number
of'different
foods
in,
which
the
substance
is
found
indicated
i
n
parenthesis.
Finally,
a
sununary
of
the
biological
data
available'for
the
substance
is
listed.
.
The
references
am
t
o
the
sunme
of
the­
principal
author
and
the
year
of
tho
publication
for
each
study.
lhe
bibliography
can
be
consulted
.for
the
complete
journal
referbncc,.
This
guide
has
been
generated
by
computer
and
is
therefore
printed
in
capital
letters,
with
the
brief
descriptions
of
the
biological
data
being
limited
t
o
60
characters.
uhbro
Greek
l
e
t
t
e
r
i
should
occur,
these
8­
printed
as
t
h
e
i
r
English
equivalent
followed
by
ui
ampersand;
..
i.
0.
u
AIL
mc!
A
.DL.
,

J
.

VGLUHE
(L
E
I
73300­

RAT
CRAL
HAGAN
6
7
FOA
UR
77
r
(continued
on
next
page)

<I
I
N
V
I
T
R
G
GRUNDSCHOBER
77
VL'LUCIE
4431
1784­

NArURAC
GCCUHREhCt
Ih
FbkO
(
2)

..
REFEWENCE
CPOYKE
16
.
..
.

USE
LEVEL
(P
P
N
,
MAX
1.00
f
O
2.
so
USUAL'
4­
30
TO
2.00
S
E
M
I
2020
dUlYL
AklHRAhlLATE
[J]

BUlYL
2­
A*
IhC&
EhZCATE
dlllYC
O­
AMl
hCdEN2l;
ATE
CFR
172­
5lS
FEHA#
2181
REFERENCE
OPDYKE
.
75
i"
*
7
I
..
8
tULOG
ICAL
.OA
rA
NONE
38.00
33.33
REFERCNCE
NO&€
CFR
1?
2­
51S
A&­
TERP
INYL
ANTHRAkIl4Tt
[8
]
i
FEPAr
3048
~

TERPINYL
2­
AFINUSEkLUATf
7EAPINYL
O­
Af41Ni26EhLOAlEE­
TERPINYL
ANLNThRAhILlTE
"SO
CALLtO'
P­
MEhIHA­
l­
EN­
8­
YL
2­
AHIhCBthZCAIE
P­
MEN7H­
I­
Eh­
8­
YL
AYTHRANlLAIE
8
lOLOCICA1
DATA
kONE
REEERENCE
Noh€
.­
C
F
R
172.515
PER
C
A
P
I
T
A
IFtTAKE
(W
/O
A
Y
)
0.00*
25
OPOY
KE
7s
1
B
I
U
L
O
G
I
C
A
L
D
A
l
A
hUNL
,
R€
fERENC€
hONE
..

f
..

..
.:
.
I
f/"#
Lp
I
CFR
172a5LS
VOLUME
118)
PER
CAPITA
I
N
f
A
K
€
tl4G/
DA7)
.

2036.
0.02036
NATURAL
OCCURRENCE
IFC
FGOU
[S)

BIOLOGICAL
O
A
T
A
R
A
T
MAL
LO1
5
0
)
2­
25­
3.38
GfUG
REF
ER€
NCE
GAUNT
70
,'

(continued
on
next
page)

93
i
..
I
4
.."
""_
"i
..
.
.
.
"
.­
.
­
..
.
.

USE
L€
VEL
IPPMJ
:
MAX
7.00
TO
12.00
USUAL
2.67
TO
6.00
6IOLOGICAl
O
A
I
A
NGN
E
..
95
fiEfERENC€­

NOM
.
..
1.
Food
Flavoring
and
Compounds
of
Related
Structure.
1.
Acute
Oral
2.
Acute
Oral
Toxicity
and
Repellency
of
933
Chemicals
to
House
and
Toxicity.

Deer
Mice.

Factors
Affecting
Absorption
from
Hamster
Cheek
Pouch.
4.
The
Acute
Oral
Toxicity,
Repellency,
and
Hazard
Potential
of
998
Chemicals
to
One
or
More
Species
of
Wild
and
Domestic
Birds.
5.
ComparisonofFish
Toxicity
Screening
Data
and
QSAR
Predictions
for
48
Aniline
Derivatives.
,
3.
Studies
of
Drug
Absorption
from
Oral
Cavity:
Physico­
chemical
'
I
I
Research
Section
Food
Flavourings
and
Compounds
of
Related
Structure
I.
Acute
Oral
Toxicity
p.
M.
J,
ENNER,
E.
C.
HAGAN,
JEAN
M.
TAYLOR,
E.
L.
COOK
and
0.
G.
FITZHUGH
Di,.
tsion
o/
Toxicological
Evaluation$
Food
and
Drug
Administration,
Vnifed
States
kpartment
of
Health.
Education,
and
Wel/
m,
Washington
25,
D.
C.,
U.
S.
A.
(Received
11
MUF
1964)

Al&
act"
Oral
dosages
of
107
synthetic
and
naturai
flavourings
and
structurally­
related
corn
~~u
n
d
s
were
administered
by
intubation
to
the
mouse,
rat
or
guinea­
pig.
A
n
i
d
were
ob­
served
usually
lor
2
weeks
during
which
time
the
development
of
toxic
si­
was
fdlowed
and
tim
of
death
r~~
rded.
The
acute
oral
LD,,
of
each
compound
was
determined.

MRODUCTlON
Substances
w
d
as
food
flavourings
have
rcceived
little
attention
from
the
toxicological
vjewoint.
Becaw
of
their
extensive
ux
as
food
additives,
the
Food
and
Drug
Administra­
tion
has
been
investigating
their
toxicity.
The
initial
step
in
our
toxicity
studies
was
the
determination
of
the
acute
oral
effects.
This
paper
presents
data
on
acute
toxicity
for
a
large
number
of
flavouring
matters.
Similar
data
are
reported
for
additionai
compounds,
not
ncccssarily
flavourings,
but
included
as
a
means
of
correlating
structure
with
toxicity.
These
relationships
b
v
e
been
discussed
by
Taylor,
Jenner
&
Jones
(1964)
and
Hagan,
JeMer,
Jones
&
Fitzhugh­
Toxicology:
Long,
Brouwer
&
Webb­
Pathology
(1964).
Flavour
additives
include
compounds
with
a
wide
variety
of
chemical
structures,
and
mixtures
of
variable
composition
derived
from
plants
and
other
natural
sources.
Some
of
the
substances
are
synthetic,
others
are
isolares
or
extracts
of
natural
products.
Since
the
purpose
of
the*
studies
was
to
evaluate
the
toxicity
of
thex
materials
in
relation
to
their
UK
as
food
additives,
a
commercially
available
material
was
used.
No
attempt
was
made
to
secure
chemically
pure
compounds.

METHODS
Groups
of
10
young
adult
Osborne­
Mendel
rats
evenly
divided
by
sa
were
fasted
for
approximately
18
hr
prior
to
treatment.
Groups
of
guinea­
pigs
consisting
of
both
males
and
females
were
fasted
for
the
same
period.
Mice
were
treated
on
full
stomachs.
Animals
had
access
to
water
at
all
times,
and
the
food
was
replaced
in
cages
as
soon
as
animals
reccivcd
their
respective
doses.
A
I
1
doses
were
given
by
intubation.
All
animals
were
maintained
under
clox
observation
for
recording
toxic
signs
and
time
of
death.
Such
observation
was
continued
until
animals
appeared
normal
and
showed
weight
gain
The
usual
observation
period
was
2
weeks;
in
a
few
cases,
where
no
acute
toxic
signs
were­
seen,
the
animals
were
observed
for
only
one
i*
eek.
LD,
's
were
computed
by
the
method
of
titchfield
&
Wilcoxon
(1949).

A
327
343
Condiments
et
Complexes
de
Strucfurc
I'oisine.
I.
Tosiciti
Ai@
par
\'&
Buccale
~Cwni­
On
adrninistra
par
intubation
des
d
m
de
complexes
faits
&
107
condimenu
syn­­
thetiqua
et
naturels,
de
structure
chimique
voisinc.
i
da
souris,
des
rats
et
des
cobayes.
On
obxrva
habituellemnt
ks
anirnaua
pendanI2
scmaines,
durant
lerquelks
on
suivit
k
divelog
pcment
de
sigtus
toxiqua
et
on
nota
la
date
de
la
mort.
Pour
chaquc
compkAe
on
dClcrmina
la
dose
ordelimite
au­
deli
de
laquellc
commence
I'intoxication
aigul!.

Lebensmittelg~
hmac~
zusatze
und
Verbindungen
verwdter
Struckt\
mn
1.
Akute
Onltoxititat
Zu~
tnrnenf~~
smng"
107
synthetirhen
un
narorliche
Gtk­
hmackuudtte
und
nrukturvcr­
wandte
Vcrbingdungm
wvrden
durch.
Intubalion
an
M
a
u
~,
Ratten
und
hlecrschweinchen
vcraixeicht.
Die
Tiere
wurdcn
gewohnlich
2
Wochen
Ian8
unttr
Beobachtung
­Iten.
uahrend
wekher
Zcit
die
Entwicklung
toxixhcr
Sxmptome
vcrfolgl
und
die
&it
da
Tod­
exintritts
rcpistrien
wurde.
Die
alrute
orale
rn~
ltlcre
Wliche
Dosir
j
d
e
r
Verbindung
wurdc
feslgcs~
ellt.

B
J
x
..,
<
cy
I
326
FORTHCOMING
PAPERS,
CORRICESDA
I
the
same­
an
initial
description
of
the
substance
is
followed
by
txamplcsof
its
likely
use,
the
toxic
dose
andlor
maximum
allowable
concentration
(i
f
known)
and
likely
pathological
effects.
Descriptionsof
clinical
findings
in
acute­
andchronicpoisoning
arc
related
to
laboratory
and
X­
ray
examination,
and
an­
outline
given
of
emergency
treatment
and
prop
nosis.
The
author
has
not
hesitated
t
o
use
trade
names
uherc
the
composition
of
the
formula.
tion
is
no*
obvious.
This
has
added
to
the
usefulness
of
the
book,
which
is
concise
and
cleal,
yet
sufficiently
detailed
possibly
lo
instruct
those
who
may
considerthcmselvcswell
ac­
quainted
with
industrial
poisons.

FORTHCOMNG
PAPERS
I
t
is
hoped
to
publish
the
following
papers
in
the
nextissue
of
Food
und
Co.~
ntetir.
t
ToxicologJJ:

Food
flavourings
and
compounds
of
related
structure.
I.
Acute
oral
toxicity.
By
P.
M.
Jenner,
E.
C.
Hapn,
Jean
hl.
Taylor,
E.
L.
Cook
and
0.
G.
Fiuhugh.

The
effect
on
rats
of
longyterm
exposure
of
Guinea
Green
B
and
Benzyl
Violet
4B.
By
W.
A.
Manncll,
H.
C.
Grice
and
lsabelle
Dupuis.
Recherchcs
dephysiologie
cellulairc
sur
la
losicitti
de
I'alcool
tthlique.
Par
R.
Low,
et
"1
:

I'olunte
I
(I963)
.
­

­I
G.
Griffaton.

Etudes
sur
I'activiti
azoreductasique
des
surnageants
d'hornogenat
de
foie
de
rat.
Par
ph.
Manchon,
S.
Gradnauer
et
R.
Low.

CORRIGENDA
x
,
p.
229,
linc
13:
for
E
214
p­
Hydrox_
vberuc?
ic
acid,
and
its
sodium
salt
IMJ
E
214
Lth!
l
4
ester
oip­
hydroxybenzoic
acid
and
its
sodium
salt.

p.
57,
lines
28­
35
inclusive
u.
hich
constitute
'Srction(
4)
Resinous
and
Pol>
neric
coaliny.
'
,'

I­
were
included
in
error
and
should
be
deleled.
1
i
Volunte
2
(1964)

.

.
­,
'
I
­1
......
..
,.
::.
L
...........
.....
­
..
,
~bmd.
Fiveindividualbioassayrepeliency
or
toxicity
variables
were
estimated
or
determined
for
deer
mice
(Peromyscus
mnniculafus)
and
house
mice
(MUS
musculus)
under
laboratory
conditions.
ALDS
(Approximate
Lethal
Doses)
or
LD"
s
of
2~
chcfnids
to
deer
mice
are
presented,
as
are
f&
nduction
(FR)
values
(3day
feeding
test
as
a
2
.~
treatment
rate)
for
whitewheatseeds
(Tri­
ticum
aesfivum)
for
6%
chemicals
and
Douglas
fu
x&
(Pseudofsuga
menziesii)
for
81
chemicals.
A
GmilN
repellency
evaluation
(REP)
using
a
Sday
test
with
white
wheat
seeds
at
a
2.0%~
treatment
rate
was
conducted
with
house
mice
and
the
results
For
347
chemicals
are
presented.
These
toxicity
and
re­
pcltency
data
should
be
useful
to
those
desiring
to
predict
the
potential
for
acme
toxicity
in
wild
mam­
mals
following
exposure
to
a
wide
variety
of
chem­
M
S
.

A
calculation
of
the
daily
chemical
dose
ingested
in
mgkglday
during
the
wheat
test
on
deer
mice
and
its
resultant
effects
on
mortality
are
also
presented
for
most
of
the
696
chemicals.
This
calculated
value,
when
used
along
with
the
ALD
or
LD5,.
should
permit
a
rough
estimate
o€
the
potential
sub­
acute
toxicity
of
any
tested
chemical,
on
wild
m
a
­
mals
for
which
both
types
of
data
are
availablt.

A
series
of
publications
summarizing
the
results
of
opproximateiy
25
years
of
chemical
research
con­
ducted
by
the
Denver
Wildlife
Research
Center
(UWRC),
on
wild
or
domestic
birds
and
mammals
has
been
initiated.
The
first
publication
presented
wild
avian
toxicity
or
repellency
results
for
998
chemicals
(Schafer
et
01.
1983).
This
paper
will
present
similar
data
for
933
chemicals
tested
on
wild
deer
mice
and
white
(house)
mice.
Our
purpose
is
to
make
available
these
generally
unpublished
test
results
so
that
they
tan
be
referenced
or
usedby
the
various
public,
private,
and
governmental
groups
that
may
require
this
information.

Methods
'
h
e
chemicals
included
in
&
tests
were
technical
of
analucii.
1
grade
pesticides
and
other
commercially
available
or
cxperi­
mental
chemicals.
Tbcy
were
purchased
from
various
commer­
cial
sources
or
conmbutcd
by
cooperating
chemical
cmnpanics.
For
presentation
purpoxs.
they
have
k
e
n
prrangcd
by
Chemical
Abstracts
Registry
Number
(CASRN).
and
PIC
idenlified
by
an
accepted
trade.
coined.
product
or
other
chemical
name
that
is
penedl?
ncrr
included
in
thc
8th
or
Rh
Collcctiw
In&\
of
the
Chemical
Absuacts
Senice.
'
Wild­
trapped
house
and
deer
mice
or
domestically
bred
house
mice
were
used
in
dl
test
procedures
which
are
dcscnid
in
deldil
by
Kvem
(1954)
and
Kverno.
a
01.
(1965).
Five
bioassay
tests
were
conducted.
resulting
in
six
basic
data
sets
as
f0llow.
t:
..

Repellency
k
c
rcpeileacy
tests
were
conducted,
two
on
dew
mice
and
one
on
house
mice.
?bc
initid
test
used
five
individually
caged
deer
mice.
Each
was
offered
2S
white
wheat
seeds
treated
with
2.0%
(wtlwt)
oftbe
candidate
chemical
daily
for
3
days.
followed
by
4
days
d
observation
for
gross
subacute
effects.
An
alternate
less
preferred
food
(laboratory
d
e
n
t
pellets)
and
aater
were
.

Because
of
the
length
and
complexity
of
Chemical
Abstracts
nomenclature.
tbe
names
used
to
identify
chemicals
in
Table
1
were
extracted
from
several
sources.
Primary
consideration
uras
given
to
the
common
name,
but
shorkned
chemical
names.
code
numbers,
or
registered
trademarks
were
also
used
but
may
not
be
specifically
identified.

..

.............
................
......
...........
..
..
....
..
..
.........
2..
;
t
12
Alloxan
lremorine
U
re
t
h
u
K
Bay
37341
DL­
Penicillamine
pilocupint
hydrochloride
3,&
Diaminopyridim
Baycr
37342
Fcnthion
Tetraethylammonium
chloride
Tributyltin
oxide
Tiiutyltin
acetote
Dicthylstifkstcrd
.
'

Bis(
phcnoxyarsiny1)
oxide
2­
Hydroxyquinoline
2­
Bentoxm\
d
Strychnine
sulfate
Dimethoate
CNilroknzoic
acid
Sodium
fluoroaceute
c­
1
Phcnylacetylurea
3­
Mcthylphcnyluru
I
­Phenylurca
Physostigmine
sulfate
Tctracyciinc
bydrochloride
~Aminosllicylic
acid
Bcnzoic
acid
Phena+
ine
hydrochloride
CydoHexamide
CM~
zhylkntenesuKoaMlidc
Endrin
MestTIlrd
N,~­
Dipkn~­
l.
Cphcnylcnediamine
p­
Nitrophcnylazoresorcind
Ethyl
mcrcrptra
Isopropyl
mercaptan
~~
H­
BuC~
I
AnOXanti8
Auytripbenyltin
Triphenyhiu
hydroxide
Diiurylcin
dilaunte
Dimcthyhydanloin
Bis(
tripheay1tin)
sulfide
.
hiutyltia
maleate
Menaura
Thiosemicarbazide
2­
MelhacryIic
acid
Dapsone
Tetramethylenedisdfotetmmine
coumaphoz
I­
Ethyb­
k
3,5­
Dunelhylpynzole
Dichlow~
thyl
b
i,.+.
c
­.
:.>
x
N­
Bu!
y~~
su~
o~~
a.
sulfa"
anijidine
".,­>,

i,.":.
p
.,
i.
:.
TigIic
acid
+:_.
Musk
xyld
Warfarin
/'

­
­1"­
50715
+1m
51730
+70.0
517%
+
1150
52608
+S
I
.
52664
+
1230
54711
­
54966
+
1125
55378
t
347
s5389
461.5.
S6348
+
I250
.%
359
+Mo
56W
437.5
56531
+I
4
6
56572
­
58366
+
50.0
59314
­
59494
­
60413
­
60515
+375
61289
­

62748
­263
63252
+6Qo
63989
+I
2
5
0
63990
+
1225
6410%
+I238
64471
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654%
+1113
65850
+
1250
66057
+336
"9
­125
67516
­
70553
­.

72004
+a
72208
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72333
+950
14317
­
74395
­
75081
+I238
62231
­

647s.
+925
75332
+
1238
75661
+I
2
5
0
76244
+
1013
76631
­68.8
76879
+
75.0
77587
+427
77714
i
l
2
5
0
77805
+663
78046
+250
78579
+613
791%
+317
79414
­1238.
8oO80­
80126
­113
80228
+121
80591
+flu)
8
i
m
+
1213
81812
+1233
..
.
.
.
.
.
,
,

'..
..
.
.
0.00
8.00
1.60
94.4
20.7
­
10.0
72.3­
83.0
95.0­
97.0
84.0
97
.O
39.5­
98.8
w.
0­
98.0
0.00
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­
­
­
70.0
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s2.
o
0.00
2.00.
1.00
.

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q1.0095)
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417.0­
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80.0
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8.00
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t
3
3
.
.
...
:
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.
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,
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82439
+
1
125
82431
+
1225
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­
83078
.+
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a3u1
+u
3
M662
­
am
+
1238
855%
­
s55983
+
1125
86533
+850
174'18
+313
87514
+
1243
87M1
­
1240
87887
+I
1
2
5
88Mo
+425
88120
+lo90
88142
?1225
60299
+
1038
89009
+I163
89021
+I
1
3
8
892s
+I
2
5
0
893%
+
1150
89394
+12M
89623
­
@I690
isso
90017
+I
2
3
0
91021
+8U
91634
+I176
92524
+450
92535
+
I225
9jm
­
92842
­
93049
+825
93107
+925
93185
+
1213
93469
"

93754
+463
94091­
94188
­
94520
+l
l
S
9
4
8
6
0
.­

95034
+975
95647
+900
95692
+315
95716
­
95761
+
75.0
957w
+.
843
9588s
­
96128
+I240
96242
+625
,

96311
+I250
964s7
+I
2
5
0
96480
+I245
96504
+350
96537
+20.0
96968
+1175'
96991
+I250
925991
­
10.0
2.00
­
56.0
"5­
98.4
­
I
­00
­
10.0
32.0
75­
0
0.5"
10.6
0.
m­
2.40
10.0
9.0
12.8
2.00
17.0­
50.0
7.00
9.00
0.00
8.
W
0.00
­
56.0
34
.O
64.0
.
,
0.00
5.87.

2.00
­
­
­
u.
0
,
.
.
26.0
3.00
­
63­
0
­.

­
5.00
­
22­
0
28.0
70.0
94
­0
31
­0
­

­
0.800
0.00
0­
00
0.330
72.0
38.4
50.0
.
.

6.00
0.00
­
.

Yellow
sulfon
chloride
Genite
+Chloro­
2,
Cdimethoxrrnilinc
2­
Amino­
S­
azotolume
Anantoin
&nzenesulfonyl,
chloride
4­
Bromobenzenesulfonyl
chloride
&Chlorobenzenesulfonyl
cbbridc
>Nitroaniline
Citrarinic
acid
SNitro­
2­
mclhylanili~
ZCNitro­
2~
methoxyaniline
.
.
1.3­
Dinitrobenzene
Methylparaben
Ethyl
4­
nitrobtntoate
Moslenc
4'­
Aminoacetopheno~
&Nitroaniline
Isocinchomcronic
acid
KyanopyridiM
Benzaldehyde
3€
yanopyridiw
3­
F+
yridylcubinol
2­
Cyanopyridine
Hexamethylenetetramine
h
c
n
e
+hind'­
nitrodiphenyI
sulfide
2­
Ethylpyridin1
~Isopropylaminophenyllmine
4­
Bcnzencazodiphcnylamine
4.4'­
Diaminophen).
lmethanc
U'­
Diaminqphen)
l
ethc:
:­
Benzylpyridine
h'.
N"
Di­
srr­
butyl­
p­
phca)­
fcnediunine
Phinyluretbam
2.5­
Dimethox)
aniline
kra­
Nitrostyrene
­
Hydroquinone
monobenzyl
ether
Azobenzcne
Benzyl
succinate
DiethyE3ox~
utarate
lj­
Diethylthioulta
F'mpyl
butyrpte
em01
CChIoroaniline
4Methplaniline
l­
Methyl4piperidinol
CChlorobenzenelhiol
.
'

Acrolein
l­
propyl
mercaptan
Tetraethylpymphosphate
>Mereaptopropionic
acid
Pyrazoxon
3­
Methylthiophenol
2.4­
Dimethylpyridine
2.6Dimethylpyridine
Thiophenol
w
;none
.
.
..

Registry
Numhcr
14,
(CAS)
(mg'kddayl
97M5
+338
97165
+1152
97507
+m
97563
+
1250
975%
+
1038
98099
+m
98588
+888
98602
+I113
99092+
37s
99116
+
1200
99358
+788
:
99592
+
IOU)
9%
u)­
99763
+I
2
5
0
99774
+IW
99854
+I238
99923
­82s
100016
+1250
100265
+I
2
5
0
..
1
w
1
+I
2
3
8
100527
+
1250
I00549
+I225
1o05u)
+I250
100709
+
1188
io0710
+
I175
100970
+I
2
5
0
101053
+
I
175
101597
­
101724
­
141757
+
838
101779
.­
1100
IO1804
­
10'5
101816
­
1W
101w2
­
101995
+
1138
102567
­
+638
102965
+I23
103162
.­
103333
+SO
io3435
­
lwm
L
105555
­213
105668
­
106445
+
1238
106478
­

106514
+
10%
106525
+
1138
106547
+lo25
107028
+1m
107039
+
1250
107493
­
113
107960
+950
108349
­12.5
108407
+I
2
3
108474
+
1238
108485
+1250
108&
+I238
105490
.
+
IO3
Deer
m
o
u
e
­
­
100
40.0
­
100
30.0
70.0
0.00­
20.0
1
­
i
2o.
o.
Q
40.0
40.0
\

10.0
­
­
..........
..
:.,
.........
..
........
.......
.....­...
.
_:_.
r...
i.
109002
+1225
109046
+I250
109057
.+
1250
109091
+I240
109433
.­

109466
+B25
109579
+I00
109795
+
I163
109808
+I
5
0
lM20
+I
1
2
5
110203.
+
1238
1
10383
+
i
I
5
0
1I"
+lo88
110601
­1225
110667
+I
t
s
0
110%
61
+
1250
110872
+12M
111864
­
111886
+I
2
3
8
112129+
950
112312
+IO50
112527
+
1238
IIUU)
+138
112889
+1206
113928
­
114261+
760
115695
+
1
0
0
115786
+
150
115902
­50.0
1159913
­50.0
116063
­55.2
11G38
­625
I
.i
i
5
2
­
1250
117782
+625
118741
+I
2
5
0
118752
+.
W.
i
1
~2
3
+
1237
118934
+I227
119324
­
319380
+I
8
8
119539
+lo25
11975s
­
119813
­
120354
f
1188
120887
­
120934
+ll75
121508
+I
2
2
5
121664
­
121711
'+
I237
121755
+490
122145
+I25
1221%
+463
122667
+
1213
122883
­
123693
+
913
123751
.
+914
120729
+
1001
124129
­
2
.00
0.00
0.00
0.00
­
34.0
92.0
7.00
88.0
10.0
I
.00
1.00
13.0
2.00
0.00
0.00
0.00
I
.00
­

24
.O
16.0
1
.00
9.
w
3.47
­
29.8­
47.9
92.0
88.0
96.0
96.0
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95.2
cn.
0
0.00
50.0
0.00
?LO
.
1.07
...
1.87
85.0
18.0
­
­

­,

5.00
19.5­
46.0
6.00
.
240
1.07
­

­

60.8
90.0­
95­
0
63.0
3.00
­
27.0
26.9
­
116
Name
Hydroxyurea
Endothd
1,4­
Naphthquinone
Quinine
hydrochloride
2­
Chioraonthraquinone
CBenrcne~
I­
naphlhylamim
I­
Naphthyiacetonitriiee
Captan
Folpct
.
2­
Methoxy­
4­
nitro­~
methy~
iline
phenyl­
2naphthylamine
CHcxylresorcind
2.
Aminobcnrolhia~
le
2­
Methylthiopknd
.
2­
Aminobcntcnethid
2,
CDiaminophenol
dihydrochloride
Thiram
zinun
Bis(
4aminophenyi)
sulfide
.
Maraluride
cchlorobcnryl
cyanide
DeXOa
Dicrotophos
2.6Diaminopyridine
2­
Hydroxypyridine
Nabam
Lakc
8cid
Decylammonium
chloride
Decyl
mercaptan
Chlordecone
Musk
tibctene
p­
Saphlholbcnzein
,

Tribuf
yltin
chloride
2­
Mercaptobenzoic
acid
8­
Hydroxyquinoline
N.
N"
Diphenykthylenedirmine
e
Methyl
anthranilate
2­~
er~
pt~
bcnZothhZok
o
c
l
a
m
e
t
h
y
l
p
y
r
o
~p
~o
~

Zinc
mercaploknzothiatdt
Methscopolamiae
bromide
2­
AminottbaneW
hydrochloride
ltobenzan
Phosphamidon
Thiodemeton
Zytron
Acreoline
hydrobromide
Lead
acetate
chloral
hydrate
2­
Chlorovinyl
diethyl
phosphale
Mexacarbatc
S­
ChlorosaIicylic
acid
.

Tirichloronate
Chminonicotinamide
3.5­
Disopropyl
phenyl
carbamate.
Tnbieth~~
y~
bOsphine
tuKde
.

2,4DiN?
rothymd
N­
2404
I
I7
333299
+
22.0
333437
­
125
367215
+
I038
379522
+
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Tesi
abbreviations
used
as
follou­
s
(see
text
for
detailed
description):
LDfr­.+
mount
of
chemical
ingei!
cd
during
the
FR
test
uhich
illled
or
did
not
kil!
more
than
.W
of
the
test
mice
ALD­.+
pproximate
Lethil
Dc*
r
tacute
orslr:
mJiaics
an
lD>@
FR­
Food
Reduction
RSI
using
2.
m­
treated
wheat
Keds
FRdf­
Same
as
R1
except
that
Douglas
fir
seeds
were
used
REP"
Percentage
of
mice
refusing
Io
eat
more
lban
­XE
of
?.@%­
treated
wheat
available
ud
lib.
Momlit).
and
the
amber
of
wheu
seeds
con­
sumed
daily
were
recorded.
fbe
I
d
number
of
treated
seeds
consumed
by
all
mice
over
the
3
d
a
y
test
period
were
subtracted
from
tbc
total
number
miiablc.
The
difference
was
convened
into
&e
percentage
of
reeds
refused,
and
the
value
w
s
desig­
oated
as
the
FR
(Food
Reduction).
For
highly
repellent
or
toxic
chemicals.
the
FR
test
war
fol­
loued
by
another
test
using
treated
Douglas
fu
seeds.
These
seeds
more
closely
represented
rht
durable
and
inedible
seed
coats
encountered
by
wild
rodents
that
cause
reforestation
seeding
problems.
Test
procedures
were
similar
to
those
used
for
the
€3
lest.
except
that
the
test
chemical
concentration
was
I.@%.
Values
resulting
from
this
test
were
designated
as
FRdf.
The
third
repeliency
lest
method
u&
d
house
mice
and
25
white
wheat
seeds
treated
with
2.
W.
ol
the
candidate
chemic+
The
wheat
seeds
were
offered
IO
10
individually
caged
mire
for
a
5­
day
period
along
with
the
same
alternate
food
used
in
the
FR
test.
?he
lest
results.
designated
as
REP,
are
summarized
by
the
p­
centake
of
mice
refusing
to
eat
an
avenge
of
13
or
more
treated
seeds
per
day
during
the
,May
test
period.
Toxicity
T\
Wo
acute
d
toxicily
tests
were
.Is0
conducted
on
deer
mice.
The
fmt.
tbe
ALD
(Approximatt
Lethal
Dose).
represented
a
range­
finding
modification
of&
Dcichman
and
LcBlanc
(1
5
~3
1
method
using
approximatcly
6
rnimals
and
a
pndualed
dosap
scale.
Using
this
single
animal
per
level
methd,
each
succeeding
treatment
­as
505
higher
Ilw
UIC
preceding
level
and
continued
until
mondity
occurred.
All
chemicals
were
administered
by
gavage
using
wafer.
corn
oil.
or
1.0%
carbopol
as
cam'ers.
fd,
towed
by
3days
d
observath
for
monality.
The
second
iox­
icily
test
(L&
I
was
conduacd
on
I
more
limitedbasis
III
a
similar
manner.
except
that
2
to
4
animals­
were
used
per
pee
metrically
spaced
dosage
kvcl.
Tbe
statistic&
method
used
IO
estimate
the
acule
o
d
wr>,
was
that
of
Thompson
(19481
and
lhompson
and
Weil(
19S2).
Thcse
tests
gemrally
requ~
rd
from
6
to
20
animals
per
expcrimtnr.
The
final
.set
of
toxicity
data
uas
derived
from
the
FR
value.
rbe
known
average
weight
of
mdr
vidual
wheat
seeds
(SO
mgl
and
the
known
average.
u.
clpht
of
each
individual
deer
mouse
(20
gm).
Tbis
calculated
value.
~h
c
g
c
s
u
l
~

me
results
of
the
tests
conducted
with
933
&mi­
cats
are
presented
in
Table
l
.
Of
the
chemicals
Icsted.
ALD's
(or
LDs'S)
on
deer
mice
were
avail­
r
~c
for
230
chemicals,
FR's
(and
usually
LDfr's)
far
696
chemicals
and
REP
values
on
house
mice
for
347
chemicals.
No
statistical
correlations
were
attempted
between
FR
and
ALD
or
LDM
values
or
REP
and
ALD
values,
because
of
the
multi­
day
na­
of
the
FR
and
REP
studiesand
the
approxi­
mation
assumed
by
the
ALD.
We
feel
that
the
ALP
or
LDfr
and
the
FR
and/
or
REP
values
could
be
used
to
generate
potential
hazard
indexes
for
acuie
andlor
subacute
exposure
of
wild
mammals
to
chemicals
in
the
environment.
Such
an
index,
or
even
a
visual
interpretation
of
the
data
presented.
should
provide
ihe
reader­
with
an
lpproximate
idea
of
the
potential
for
ingestion
and
~ubsequent
mortality
in
wild
mammalians
exposed
to
the
listed
chemicals.
Such
an
index,
if
generated
and
defined
as
the
one
proposed
by
Schafer
et
a[
4
1983).
may
serve
as
an
additional
predictive
tool
in
determining
potential
mammalian
mortality
lowing
envirunmental
exposure
to
chemically
con­
taminated
or
treated
fdod.
I
4
2
9
Artno@.
lrd~
mrnfr.
The
authors
wish
lothrnk
!he
following
in­
dividuals
fop
their
assistance
on
cdkctimg.
typing
and
editing
the
data
contained
in
this
publication:
M
J
I
~
Eschen.
Patrick
Fundcrburg.
David
Hayes.
Glenn
Hood.
Brenda
Lcchup,
Ce
cila
Nelson.
Florence
Poe.
and
Peter
slv.
rie.
RK
compilation
8d
prcwration
of
the
data
included
in
this
publication
was
sup
­ported.
in
pan.
through
Interagency
Agreement
14­
164009­
8l­
957,
whichwas
funded
by
the
U.
S.
Environmental
Protection
Agency.
Ollicc
of
Toxic
Substances,
Washington,
X.

References
Deichmann
W.
LeBlanc
TJ
(1943)
Determination
of
the
approx­
imate
lethal
dose
with
about
six
mimais.
J
Ind
Hygiene
Tox­

KGem
NB
(19541
Development
of
better
.seed
protectants.
J
Forestry
52~
826­
827
Kvemo
NB.
Hood
GA.
Dodge
WE(
1W)
Development
ofchem­
i
u
l
s
to
control
forest
wildlife
damage.
hac
SOc
Amer
For­

Schafer
EW
Jr.
Bowles
WA
Jr.
Hurfbut
JIlW13)
The
acute
oral
toxicity.
repellency
and
huvd
potcnlirl
of
998
chemicals
to
one
or
more
species
of
wild
and
domestic
birds.
Arch
En­
viron
Contam
Toxicol
12355­
382
Thompson
WR
(1948)
Ux
of
moving
rverages
and
interpolation
to
estimate
median
effective
dosc.
Bacterial
Rev.
11:
I
15­
145
Thompson
WR.
W
i
l
CS
(1952)
On
the
constmction
of
tables
for
moving
average
interpolation.
Biometrics
831
­54­
icol
Tj:
llS­
417
tSten
65~
222­
226
......
..
..

...
.................
..
..

P
STUDIES
ON
DRUG
ABSORPTION
FROM
ORAL
CAVITY:
PHYSICO­
CHEMICAL
FACTORS
AFFECTING
ABSORPTION
FROM
HAMSTER
CHEEK
POUCH
y~?]
KUROSAKI,
NORKO
AYA
(nee
YAMASHITA).
YUhllKO
OKADA,
TAU]
NAKAYAMA
AND
mol
o
i
TOSHIKIRO
KUIURA"
Ptctle
(Received
August
27,1985)

Keywords
"8bsorption;
oral
mucosa;
hamster
cheek
pouch;
pharmacokinetics;
physico­
chemical
factor;
salicylic
acid
INTRODUCTION
In
this
paper,
we
describe
a
new
experimental
The
absorption
of
drugs
from
the
oral
cavity
method
for
studying
absorption
processes
across
has
been
investigated
by
the
buccal
absorption
the
keratinized
oral
mucosa
in
vivo
usinga
ham­
test
of
Beckett
and
Triggs"
in
man.
In
these
stud­
ster
cheek
pouch,
which
consists
of
keratinized
ies.
it
has
been
shown
that
the
ability
of
a
com­
stratified
squamous.
epithelium,
and
discuss
pound
to
cross
the
oral
epithelia
is
highly
depen­
some
characteristics
of
drug
absorption
through
the
lipophilicity
of
its
unionized
form.*"
1
The
oral
mucosa
has
been
classified
into
three
types
MATERIALS
AND
METHODS
according
to
function;
i.
e.,
masticatory,
lining
Chemicals
­All
chemicals
usedwere
rea­
and
specializedmucosa..
Also
rhere
are
regional
gent
grade
commercial
products.
.

variations
i
n
the
epithelial
thickness
and
the
Procedure
ofAbsdrption
Experiments­
Male
degree
of
keratinization.
5)
Although
the
buccal
golden
hamsters
(80­
110
g
body
weight)
were
absorption
test
in
man
can
easily
provide
data
anesthetized
with
urethane
(1.5
g/
kg,
i.
p.
)
and
concerning
the
absorption
rates
of
drugs
admin­
were
fastened
to
a
platform
at
an
angle
of
&ut
istrred
in
the
oral
cavity,
the
test
does
nor
show
5
5
3
The
cheek
pouch
was
cleaned
by
multiple
~h
r
regional
differences
in
the
absorprion
rates6)
rinses
with
saline
and
the
iumen
was
wiped
with
o
r
the
rate­
limiting
step(
s)
(permeation
barrier)
cotton
balls
to
remove
excess
water.
A
vinyl
of
the
absorption
process.
5.6)
Studies
on
mucosal
tubing
(0.
d.
1.2
nun,
i.
d
0.8
mm,
Dural
Plastics,
permeability
to
drugs
in
virro,
using
diffusion
Australia)
was
inserted
into
the
cheek
pouch
and
cetls,
provide
valuable
information
concern&
the
was
fsed
with
Aron
Alpha
(Toagosei
Chemicals,
relationship
between
the
rare
of
transfer
and
the
japan)
to
the
corner
of
mouth
in
40
mm
depth
physico­
chemical
properties
of
substances
such
(Fig.
1);
Drugs
were
dissolved
in
appropriate
as
Lipid
solubility,
molecular
size,
chemical
com­
isotonic
buffer
solution
(citric
acid­
Na,
HPO,
at
position
and
abiliy
to
form
hydrogen
b
~n
d
s
,~­~l
'
p
H
2.0
to
5.0
and
NaH,
P0,­
Na2HP0,
ar
pH
but
no
information
concerning
the
transfer
pro­
4.0
a
d
pH
7.0).
One
ml
ofthe
drug
solution
was
C
~S
S
frbm
the
oral
tissue
to
rhe,
sy,
stemic
circula­
admin­
isrered
into
the
cheek
pouch.
After
stand­
ing
for
a
definite
period,
the
luminal
contents
.
,
dent
upon
both
the
degreeof
its
ionization
and
a
keratinized
region
of
or­
al
mucosa
.*
TO
whom
correspondence
should
be
addressed
288
were
withdrawn
and
the
cheek
pouch
was
washed
with
the
same
buffer
solution.
The
wash­
ings
were
combined
with
the
luminal
contents
and
made
up
to
20
ml
by
addition
of
saline.
The
amount
of
the
drug
remaining
was
determined.
The
disappearance
from
the
lumen
was
defined
as
the
apparent
absorption.
Determination
of
Salicylic
Acid
Remainiwin
Tissue
of
Cheek
Pouch
­in
some
absorption
ex­
periments
for
salicylic­
acid,
the
amount
of
the
drug
remaining
in
the
tissue
of
cheek
pouch
was
vinyl
tubing
(0.
d
1.2
mm,
i.
d.
0.8
mm)

FIG.
1.
SchematicRepresentation
of
Absorption
fiperimenl
f
o
r
Hamster
Cheek
Pouch
Y.
Kurosaki,
el
ai.

determined.
After
washing
out
the
residual
salicylic
acid,
the
cheek
pouch
was
irnmedately
cut
off,
A
half
N
KaOH
solution
was
added
to
the
tissue
to
make
the
total
amount
20.0
g.
The­
tissue
in
alkaline
solution
was
then
placed
in
a
boiling
water
bath
for
30
min.
After
cooling
the
resultant
solution,
some
impurities
were
extracf­
ed
with
chloroform.
The
remaining
aqueous
phase
was
.made
acidic
with
concentrated
HCl
and
salicylic
acid
was
extracted
with
chloroform.
Then,
salicylic
acid.
in
the
organic
phase,
'was
re­
extracted
with
0.1
hT
NaOH
solution
and
the
drug
in'the
final
aqueous
layer
was
determined
by
a
high
pressure
liquid
chromatography
(HPLC).
For
the
calibration
curve,
appropriate
amounts
of
salicylic
acid
were
added
to
the
tissue
of
cheek
pouch
and
the
same
procedures
were
carried
out.
Analytical
Methods
"p
Aminobenzoic
acid
and
sulfisoxazole
were
diazotized
following
a
regular
procedure,
coupled
with
2­
diethyl­
aminoethyi­
1
­naphthylamine
and
extracted
with
isoamylalcohol
after
addition
of
1
g
of
NaCl.
The
optical
density
of
the
organic
layer
was
determined
at
560
and
555
nrn,
respectiv$
ty.
HPLC
was
used
to
determine
the
other
com­
pounds
remaining
in
both
lumen
and
tissue
of
the
cheek
pouch.
The
chromatograph
was
5703
(Gasukuro
Kogyo,
Japan)
equipped
with
a
ultra­
violet
(UV)
detecror
(5025,
Gasukuro
Kogp).

TABLE
1.
HPLC
Condirions.
for
the
Anabsis
qf
Test
Compounds
Compound
Mobile
phase"
'
Wave­
length
(nm)

.
Salicylic
acid
45
:
55':
21
5
Phenol
50
:
50bi
220
Benzoic
4
0
:6
0
b
t
227
Acetylsalicylic
acid
50
:
50
"
220
Propionylsalicylic
acid
50
:
50
b'
220
Butyrylsaliqlic
acid
40
:
60°
'
220
m­
Hydroxybenzoic
acid
50
:
50h'
300
p
Hydroxybenzoic
acid
50
:
50b'
253
0­
Toluic
acid
30
:
70
'
I
220
Anthranilic
acid
70:
30:
3''
248
Acetanilide
40
:
606'
237
Acetaminophen
40
:
60
bl
240
Phenacetin
#:&
Ib)
243
Methylparaben
35
:
65
b1
254
Ethylparaben
30
:
70
'1
254
Propylparaben
­
.
25:
75b'
254
a)
Valuesareexpressed
as
volumetovolumecomposition
of
two
orthreecomponents.
.
b)
0.025'~
phosphoric
acid
:
methanol.
c)
H2
0
:
acetonitrile
:
aceric
acid.
..
D
T.
C
P'
f
f
sa
P'
Pi
Ot
m
u'

ca
Tht.
column
used
was
a
reversed­
phase
Unisil
Q
c:
1
8
(4
.O
x
3(
K)
mm,
Gasukuro
Kogyo).
Mobile
+ses
and
the
wave­
lengths
for
determination
summarized
in
Table
I
.
An
aliquot
of
the
simple
soluti'on
was
filtered
through
a
0.45
p
m
pore­
size
triacetylcellulose
membrane
(Fuji
Photo
Film,
Japan)
and
an
appropriate
volume
(,(­
the
filtrate
was
injected
into
the
liquid
chro­
matograph.
The
concentration
of
the
compound
u';
ls
calculated
from
the
peak
height
using
the
c;
ilibration
curve.
Plasma
Concentration
of
Salicylic
Acid
­Under
urethane
anesthesia,
the
carotic
artery
was
cannulated
with
polyethylene
rubing
(0.
d.
(1.8
mm,
i.
d.
0.5
mm,
Dural
Plastics)
and
then
heparin
(500
unitlkg)
was
administered
intrave­
nously.
In
a
n
.
i
v
.
administration
study,
salicylic
acid
was
dissolved
in
saline
to
make
20
mM
and
was
injected
into
a
femoral
vein
(1
mVkg=
20
pmol/
kg).
In
the
study
of
intra­
cheek­
pouch
ad­
ministration,
salicylic
acid
dissolved
in
isotonic
buffer
solution
at
the
concentration
of
10
mht
was
administered
into
the
cheek
pouch
in
a
simi­
lar
manner
as
described
in
Procedure
of
Absorp­
rion
Experiments
(10
ml~
kg=
100
pmoVkg1:
in
ht.
h
studies,
blood
samples
(0.2
m
l
)
were
col­
lected
periodically
from
the
cannula
for
4
to
6
h
after
the
administration
and
the
plasma
was
separated
immediately
by
centrifugation.
Afrer
addition
of
acetonitrile,
the
plasma
was
cenrri­
fuged
and
the
supernatant
?as
filtered
through
a
0.45
pm
pore­
size
filter
(h'ihon
Millipore
.

Kvgyo,
Japan).
Salicylic
acid
concentrations
in
plasma
were
determined
by
HPLC
equipped
yic
h
a­
fluorescence
detector
RF­
540
(Shimadzu;
Japan)
at
300
anh
430
nm
for
elcitation
and
emission.
respectively.
Pharmacokineric
Studies
­Plasma
concen­
t
r
a
t
i
o
n
­t
i
m
e
d
a
t
a
f
r
o
m
i.
r.
a
n
d
i
n
t
r
a
­
cheek­
pouch
administration
studies
were
simul­
taneously
fitted
to
a
two­
compartment
model
(F
I
~
6)
using
a.
nonlinear
least­
squares
program,
hlULT1.
'2'
In'this
model,
the
parameters
of
Iag
times
and
absorption
rate
constants
for
two
intra­
cheek­
pouch
administration
studies
were
independent
of
each
other.
Determination
of
Lipophilic
hde­
x.
log
(k
;I,
at
p
H
3.0
­The
lipophilic
index,
fog
(k
b),
of
each
compound
at
pH
3.0
was
derermined
by
reversed­
phase
HPLC
according
to
the
method
d
Yamana
er
~l
.
13)
The
apparatus
for
HPLC
HJS
the
same
as
described
in
Analytical
Methods
and
all
compounds
were
detected
at
254
nm.
The
mobile
phase
of
methanol­
buffer
solution
(pH
3.0)
made
of
12.6
mM
citric
acid,
6.4
mM
Na,
HPO,
and
10
mht
NH,
CI
was
run
at
a
flow
rate
of
1
.O
rnL'min.
Ammonium
salt
was
added
to
block
the
acrive
siianol
sites
of
the
column.
Formamide
was
used
as
an
unretained
substance.
When
log
(A").
which
w.
as
defined
as
follows;

where
t
R
and
t
o
are
rhe
retention
times
of
a
retained
peak
and
of
an
unretained
peak,
respec­
tively,
was
plotted
against
methanol
concentra­
tion
(v/
v
%),
reasonable
linear
relationships
for
ali
compounds
tested
(r
<
­0.999)
were
ob­
tained.
The
lipophilic
index,
log
(kb),
was
defined
as
a
log
(k')
value
extrapolated
to
0%
methanol.

RESULTS
Absorptionfrotn
the
Hamster
Cheek
Pouch
pH
3.0
from
the
hamster
cheek
pouch
was
exam­
ined.
Table
I1
summarizes
the
results
of
the
ab­
sorption
experiment.
The
percentage
of
absorp­
tion
of
compounds
from
the
lumen
of
the
cheek
pouch
i
n
1
h
varied
over
a
wide
range.
In
the
case
of
alkyl
acid
esters
of
salicylic
acid,
the
ab­
sorption
increased
as
the
alkyl
chain­
lengths
in­
creased,
i.
e.,
acetyl­
<
propionyl­
<
butyryka­
licylic
acid.
Similarly
among
parabens,
the
ab­
sorption
increased
in
the
order
of
methyl­
<
ethyl­
<
propylparaben.
These
results
are
in
.

agreement
with
the
buccal
absorption
data
of
~~
'­
n­
alkylarnin~
s.~
'
The
absorption
of
both
m­
hydroxy­
and
p
­
hydroxybenzoic
acid,
the
struc­
tural
isomers
oi
salicylic
acid,
were
about
one­
tenth
of
salicylic
acid.
Likewise,
for
aminoben­
zoic
acids.
the
0­
isomer
(anthranilic
acid)
was
.

more
absorbable
than
the
p
­
isomer.
Elkcr
of
Llrrnirlcl
Concentration
on
zhe
Absorption
q
f
Saliqslic
.4
cid
The
effect
of
luminal
concentration
of
salicylic
acid
on
its
absorption
from
the
cheek
pouch
was
examined
in
the
concentration
range
of
1
.O
to
10.0
m5f.
The
percentages
of
the
absorp­
tion
in
I
h
ar
pH
3.0
were
48.6,498
and
48.2
at
1
.0,
5.0
and
10.0
mhi
respectively
(Fig.
21,
show­
ing
the
linearity
o
i
salicylic
acid
absorption
in
the
concentration­
range
examined.
Time
Course
ofSali&
c
Acid
Absorption
To
clarify
the
process
of
the
absorption
of
a
drug
after
inrrz­
cheek­
pouch
administration.
10gW)
=
1.
i
N
t
R
­t<))/
tol
(1)

The
absorption
of
18
aromatic
compounds
at
.
'i
t
t
TABLE
11.
Ahsorpi<
w
(?
f
M(
lCkd
Conlpouttdsfrom
rhr
Hamster
Check
Pwch
arpH
3.0
t
ion
P
i
a
Molecular
Conc.
c/
c
absorbed
in
tere
Compound
weight
(mM)
1
ha'
the
Phenol
'
94.1
1
.o
34.62
2.6
(4)
pro1
Benzoic
1.0
37.62
1.5
(31
t
imc
Salicylic
1.38.1
1
.o
Acetylsalicylic
1
­0
11.1
f
1.6(
3)
seer
Propionykalicylic
acid
194.2
1
.0
25.7
2
1.9
(4)
mol
Buryrylsalicylic
1
.Q
40.8
5
5.5
(4)
sali
m­
Hydroxybenzoic
acid
138.1
5
.o
6.3
2
0.8
(4)
a
i
t
t
p
­
Hydroxyhcnzoic
acid
138.1
5
.O
Oi
5
u­
Toluic
acid
Anthranilic
acid
:
­
137.1
p­
Aminobenzoic
acid
137.1
1
.o
.
squ
Sldfisoxazole
267.3
kln
Acetanilide
135.2
1
.o
7.7
2
19
(4)
111.

Acetaminophen
151.2
I
.o
0.9­
t
0.5
(4)
rim
Phenacetin
179.2
1
.o
16.0
­t
0.6
(3)
tru
Methylparaben
154.2
1
.0
30.1
f
2.5
(7)
anc
Ethylparaben
166.2
1
.0
44.8
k
0.8
(4)
cor
Propylparaben
180.2
0.9
64.6
+­
3
2
(4)
48.62
4.1
(4)
.
J
i
i
l
6.5
2
0.7
(4)
136.2
1
.o
50.
Gk
3.0
(4)
1
.o
14.3­
t
1.6
(4
)
a
8.6
k
1
.O
(4)
1
.o
6.0
2­
0.7
(4)
c
w
s
R
~~~/~~
are
eypri>
ssedas
the
mean
2
S.
E.
with
rhe
number
of
aperiments
in
parentheses.

salicyiic
acid
was
administered
and
the
arnounrs
of
salicylic
acid
remaining
in
both
the
lumen
and
the
tissue
of
the
cheek
pouch
were
period­
caliy
determined
ar
pH
3.0
and
at
pH
4
0.
Re­
sults
are
shown
in
Fig.
3.
The
semilogarithmic
plots
of
the
luminal
salicylic
acid
(open
circles)
indicated
that
the
drsapprarancr
from
the
lumen
could
he
described
by'a
biexponential
process.
f­
Iowewr,
the
rransier
to
rhr
systemic
circularion
(closed
circles)
was
shown
to
be
an
apparent
first­
order
prociss
at
both
pH
conditions.
AC­
cordin&,
it
is
rcasonddt
to
consider
that
the
­systemic
transfer
after
inrra­
cheek­
pouch
ad­
ministration
can
ht
approximared
by
the
first­
order
process
after
a
lag
time.
The
apparenr
transfer
rate
consranrs
t6
the
systemic
circulation
calculated
from
rhr
slope
`of
rhr
clcked
circles
and
the
lag
Times
estimated
by
extrapolation
were
0.49
`h­
1
and
7
min,
and.
0.11
h­
1
and
25
min
for
pH
3.0
and
pH
1.0,
respeaively.
Plasma
Concentration
ojSaIi@
ic
Acid
a$
er
Inrra­
Cheek­
Pouch
Administration
Plasma
concentrations
of
saliqlic
acid
after
intra­
cheek­
pouch
administration
(100
KmoVkg)
as
a
function
of
time
are
shown
in
Fig.
4.
The
plasma
level
wsus
time
curves
were
clearly
dependenr.
on'
the
pH
of
the
administered
solution.
When
salicylic
acid
was
administered
at
pH
3.0.
the
plasma
concentration
was
rapidly
,
­

increased
t
o
abour
100
nmol/
mi
and
then
de­
creased
thereafter.
On
the
other
hand,
ar
pH
4.0,
it
took
about
1.5
­
2
h
to
reach
the
maximal
con­
centration
of
about
25
nmol/
ml
and
this
concen­
tration
was
maintained
for
at
least
5
h..
Pharmacokinerics
of
Salicyiic
Acid
after
lnrra­
CheeA­
Pouch
Administration
To
confirm
the
plasma
elimination
kinetics
of
salicylic
acid
in
h
m
t
e
r
s
,
an
i.
r.
administra­

5
10
Iniria!
cnncrnaauon
(rnd
h
o
s
a
k
i
,
et
a/.

arion
kinetics
.
a+
ninisrra­

­q­
f
l
,)(l
wds
carried
out.
As
shown
i
n
Fig.
5
,
the
p
l
~~n
~a
concentrations
of
intravenously
adminis­
rcred
salicylic
acid
declined
biexponentialty.
As
tilt
ahsorption
from
the
cheek
pouch
can
be
ap­
ppximared
by
tbe
first­
ordgr
process
after
a
1%

1
imc
(Fig.
?),
the
two­
compartment
model
with
'1
tlrst­
order
absorption
process
shown
in
Fig.
(1
sctmrd
to
be
available
as
the
pharmacokinetic
model
for
intra­
cheek­
pouch
administration
of
salicylic
acid.
Plasma
concentration­
tim&
data
attrr
intra­
cheek­
pouch
and
i.
c
administration
~1
t
'
salicylic
acid
(Figs.
4
and
51
were
simultane­
ously
flttcd
to
the
model
using
a
nonlinear
least­
squares
program,
MULTI.
'2'
and
the
pharmaco­
kinetic
parameters
estimated
are
listed
in
Table
111.
The
absorption
rate
constant,
k
a,
and
the
lag
time,
5
,
estimated
here
agreed
with
the
apparent
,
transfer
rate
consrant
for
the
systemic
circulation
and
the
lag
time
obtained
from
Fig.
3
at
each
pH
condition,
respectively.
EJfcct
of
Lipophilici!
r
on
tlw
.4
bsorprion
from
thc
Hanlstcr
Chcek
Pouch
log
(li
7
values
together
wirh
log
values
for
18
compounds
at
pH
3.0
are
listed
in
Table
I
V
.
I
t
has
been
shown
that
the
absorption
from
the
oral
mucosa
is
related
to
the
lipophilicity
of
the
compound
in
horh
in
I
~w
~­~.~~~
and
in
correlation
(r=
O.?
64)
berween
log
t
k
i)
and
the
absorption
rare
at
pH
.i.
O.
Thr
theory
of
diffu­
sion
through
a
single­
layer
homogeneous
mem­
brane.
predicts
for
small
compunds
that
per­
meahility
is
inversely
proportional
to
rhe
square
root
of
the
molecular
weight
(,&
fr).
15)
When
this
concept
was
applied,
Fig.
7b
w'as
obtained
where
rhe'abscissa
was
altered
to
log
j
(A{,)
'".
A
better
correlation
(r
=
0.874)
was
observed
br­
tween
log
(k
b
I
(Mr)
''?
and
the
absorption
rate,
suggesting
that
not
only
the
lipophiliciry
but
also
the
molecular
size
may
affect
the
absorption
V/
lilrO
9­
1
1)
experiments.
Fig.
7a
shows
the
positive
FIG.
3.
Time
Course
of
Sali&
c
AcidAbsorption.
fion1
rhe
Hamster
Cheek
Pouch
Semilogarithmic
plors
qf
saliqlic
acid
remaining
in
lumen
and
in
tissue
of
the
cheek
pouch
(a
i
at
pH
3.0
and
(b)
at
pH
4.0:
0
,
saliq­
lic
acid
remaining
in
iuminalfluid:
0
,
sun1
of
saliqlic
acid
ren.
loining
in
lumen
and
in
tissue.
(el
Tusue
accumulation
of
sali~.
lic
acid:
C
,
pH
3­
0:
c)
,
p
H
4.0.
R
e
d
s
arc,
ex­
pressed
as
the
mean
4
S.
E.
ofaat
leas1
three
ex­
perimenrs.
Y
.
Kurosaki,
et
a/.
292
from
the
keratinized
oral
mucosa.
EIffecr
of
pH
011
thc
Absorpiion.
fron1
rhc
Hamsrer
Cheek
Pouch
To
determine
the
effect
of
pH
on
the
absorp­
tion
from
the
hamster­
cheek
pouch,
absorption
2oo
t
4
I
0
1
2
3
4
5
6
Time
(h)

FIG.
4.
PIasma
Concentration
of
Salit?­
lic
Acid
affer
Intra­
Cheek­
Pouch
Adminisrrariolr
100
p
nt0Vh­
g
of
saliqvlic
acid.
was
ateltzinistered
into
the
cheek
pouch:
C
,
pH
3.0;
e.
I..
'
'4.0.
Re­
1
sults
are
expressed
as
the
mean
i­
S.
E.
,f
a
t
least
three
'experiments.
Each
line
represenr.;.
ihe
curve
$nedwith
the
wo­
comparrmenr
modc.
shown
in
Fig.
6.
experiments
were
carried
out
for
salicylic
acid,
benzoic
acid
and
phenacetin.
Fig.
8
is
the
plots
of
absorption
as
a
function
of
pH.
The
pH
change
was
negligible
throughout
the
experiment.
The
absorption
of
both
salicylic
acid
and
benzoic
ancid
were
decreased
with
the
increase
of
the
pH
of
the
solution
and
almost
nil
at
pH
?.
o,
where
these
compounds
n:
ere
complettly
(
>99%
1
ionized
since
the
pk',
values
were
3.0
and
4.2,
respectively.
On
the
other
hand,
the
ah­
sorption
of
phenacetin,
which
is
not
ionized
in
thepH
range
examined,
was
not
affected
by
the
pH.
These
data
suggest
that
the
ionized
form
of
the
compound
was
poorly
absorbable
from
the
hamster
cheek
pouch
although
the
inflection
points
of
the
pH­
absorption
curves
were
slightly
shifted
to
the
basic
side
compared
with
the,
pK,
values
in
both
acids.

DISCUSSlON
The
morphology
of
the
oral
mucosa
varies
from
regi0.
n
to
region
depending
on
function.
It
is
likely
that
there
are
differences
in
permeability
among
structually
dfferent
regions
of
the
oral
mucosa,
such
as
keratinized
or
non­
keratinized.
6)
One
of
the
simplest
methods
hrercly
measuring
permeation
rate
through
the
oral
mucosa
in
vivo
is
buccal
absorption
test
of
Beckett
and
Triggs."
in
this
method,
the
uptake
of
a
compound
is
es­

FIG.
5.
Plasma
Concenrration
of
Salicylic
Acid
after
i.
v.
Administration
in
Hamster
­20
pmoVkg
of
salicylic
acid
was
injected
into
the
femoral
vein.
Resulrs
are
=pressed
as
the
mean
k
S.
E.
of
three
aprimenfs.
Line
represents
the
curw
fined
uith
the
two­
compartnlenr
model
show
in
Fig.
6.
­
FIG.
6.
Pharmacokinetic
Model
for
Salicylic
Acid
in
Hamsrer
D
or
D'
,
dose:
t
o
,
lag
time;
k,,
first­
order
ah­
sorption
rateconsrant;
k,,
.firsr­
orderexcretion
rate
consrant;
k
and
k
z,,
.first­
order
transfir
rate
constants
betneen
two
comparmlents:
1:.
volume
of
disrribution
of
censral
comparmwnc
V2,
volume
of
distribution
of
peripheral
comparr­
ntenf.
Saliqslicacidwasadministered
br
i.
r.
or
infra­
cheek­
pouch
administration.
Drug
Absorption.
fiom
Hamster
\
Cheek
Pouch
rrn,
ilrccf
from
t'hr
differences
in
amount
hetween
initial
and
the
final
solutions
in
the
mouth.
Hrtu.
ever,
this
method
cannot
provide
informa­
rlon
as
to
the
relative
permeability
of
differem
regions
in
the
oral
cavity,
because
the
area
where
the
absorption
may
have
taken
place
cannot
be
tclnfirmed.
In
addition,
since
the
measurement
­;
IS
carried
Out
in
human
volunteers,
the
test
conditions
should
be
restricted;
that
is,
the
;q:
cnrs
nhich
are
thowht
to
damage
the
biologi­
c
SI
membranes
cannot
be
used
for
studying
the
permeation
barriers.
Furthermore,
the
data
from
untrained
volunteers
showed
wide
deviation
in
this
In­
wlro
methods,
usually
using
diffusion
cells,
enable
anatomically
well­
defined
regions
of
mucosa
to
be
studied,
but
cannot
pro­
vide
information'concerning
the
transfer
from
oral
tissue
to
systemic
circulation.
In
this
study
we
used
a
hamster
check
pouch,
which
has
keratinized
suatified
squamous
epithelia")
simi­
lar
to
those
of
gingiva
aid
hard
palate
in
man
and
which
could
be
well
separated
from
the
Pharmacokinetic
parameter
Condkion
of
administration
pH
3.0
pH
4.0
0.103
to
(mid
8.4
24.0
k,
(h
­9
'
0.529
k,
(h
­I)
2.25
v,
Wkg)
0.125
V
,
(Vkg)
0.185
k
12
(h
1.05
k2i
(h
­')
0.7
1
Thc
parameters
were
estimated
by
Ihe
damping
Gauss­
New:
method.
Since
the
anabrical
limir
ofthe
plasma
salicylic
acid
was
less
than
I
nrnofml,
the
reliabiliw
Ofii
e
data
appeared
to
be
equivalent
in
each
paint.
Thus,
in
esrimarion,
(CJ­
2
was
adopted
as
the
weight
wht:
e
Ci
is
the
value
ofthe
i­
thpoint.
Aka&
e>
information,
A
K
,
in
this
estimation
was
­1
9.3.

TABLE
IV.
Lipophilic
Indexes
of
Model
Compounds
ar
pH
3.0
Phenol
­2.
Benzoic
acid
Sahcylic
acid
Acetylsalicylic
acid
Propionylsalicylic
acid
Butyrylsalicylic
acid
m­
Hydroxybenzoic
acid
p­
Hydroxybenzoic
acid
o­
Toluic
acid
.
Anthranilic
acid
Sullfisoxazole
$­
lit
Acid
p­
Aminobenzoic
acid
order
ab­
Aceranilide
i?
vcwtion
Acetaminophen
N.
D.
b'
­0.367
­0.1
13
0.1
16
0.344
1.06
­0.514
­0.212
0
.c
39
0.392
N.
D.
1.60
N.
D..
­0.513
­0J12
0.103
0.414
1.42
.p
.

N.
D.
­0.486
­0.1
74
0.1
03
0.412
1.30
­0.596
­0.253
0.(
92
0.450
N.
D.
1.84
­
037
1
"0.
W
0..:,
77
0.770
N.
D.
2.30
AT.
D.
­0.678
­()..;
a
­0.058
0.252
1.18
N.
D.
­O.
'U
­O.­
i23
­0.127
0.171
1.09
N.
D.
­0.405
"0.1
29
0.1
31
0.385
.
1.18
N.
D.
"0812
­0.439
­0.046
0.389
1.57
­0.320­
0.001
.
0.116
0.64
7
N.
D.
1.93
N.
D.
N.
D.
"0.682,
­0.388
­0.110
0.75
N.
D.
­0.340
­0.084
0.1
74
0.455
1.24
N.
D.
ND.
­0.824
­0.549
­0.253
OM,
N.
D.
1.69
N.
D.
1.70
N.
D.
2.1,7
a)
A~
obi~
ePhQse.
61
.%
t
determined.
Results
are
expressedas
the
mean
of
drrpIicafe
aprimena.
Phenaceti;
Methylparaben
Ethylparahen
Propylparaben
­0.441
­0.153
0.1
5
1
0.477
­0.522
­0217
0.102
0.430
­0.328
0.006
0.364
0.746
­0.1
18
0258
0.658
N.
D.
N.
D.
2.59
other
oral
mucosa
during
the
absorption
experi­
ment
to
investigate
(ii
the
process
that
a
com­
pound
administered
in
solution
disappeared
from
the
lumen
of
the
cheek
pouch,
(ii)
the
pro­
cess
that
the
compound
transferred
from
the
cheek
pouch
rissur
into
the
systemic
circularion
and
(iii)
the
factors
affecting
thost
processes.
It
has
been
demonstrated
at
various
sires
thar
the
absorption
r
a
m
of
compounds
a!­?
sorbed
by
a
passive
diffusion
mechanism
c
o
r
­1ated
well
with
their
IipophiIiciry.
Recently.
I
Jered
er.
at.
using
the
human
buccal
absorption
:est
showed
that
there
are
carrier­
mediated
tranyon
systems
for
nicotinic
acid.
nicorinamide,
lE
thiamineIg)
and
glutathione2"
in
the
human
oral
cayity.
In
the
keratinized
mucosa
including
h;
mter
cheek
pouch,
however.
the
carrier­
medm
ed
transport
system
may
nor
exst.
As
shown
in
''igs.
2
and
8.­
the
absorption
oi
s211cylic
acid
vas
increased
linearly
with
the
dosr
and
was
pji­
dependenz.
namely,
the
absorption
was
poor
x
thehigher
pH
regions
whert
This
compound
x
'as
compkte­
1y
ioniztd.
in
addxion,
there
was
a
[mirive
corrc­
larion
between
rhr
absorption
x
e
s
and
the
lipophilic
indexes.
which
correkxed
well
with
rht
partition
coefi1aenrs
in
1
­ocr;
nol­~
ater,*~
'
with
the
18
aromatic
compounds
lasted
(Fig.?).
Consequently;
it
can
be
assumed
tiat
the
absorp­
tion
mechanism
from
the
keratinized
hamster
cheek
pouch
is
a
passive
diffusion.
There
are
only
a
few
studies
concerning
the
absorption
from
the
hamster
cheek
Whitford
e1
ai.
23'
reported
thar
the
absorptin
of
fluoride
through
this
keratinized
epithelium
occurred
mainly
by
diffusion
of
undissociated
acid,
HF.
and
this
agrees
ell
with
the
present
findings.
The
data
presenrtd
here
involve
two
prith­.
lems
to
be
solved.
One
is
a
pH­
shift
shown
in
.ed
by
a
d
well
~)r
~l
~~
.4bsor~
l10n./
r~
7m
Namsrcr
ChccA
Porrc­
h
~~
g
8
.
Sin,,
the
ph',
values
for
salicylic­
acid
and
t­
rcnzoic­
acid
are
.3,0
and
4.2.
respecrively.
the
pH­
proiiles
of
the
absorption
for
both
acids
U'erc'
shifted
to
basic
side
approximately
0.5
­
1.0
pH
unit
from
each
curve
of
unionized
fracrion.
The
shifts
of
pH­
absorption
curses
hSlve
already
been
reported
for
somr
drugs
in
the
psrrointesrinat
tract
including
the
oral
cayiry
2nd
the
contribution
of
physiolopical
factors
such
as
a
higher
acidic
pH,
if..
virtual
pH
at
the
membrane
s
u
~f
a
c
e
,~~.~~)
(he
buffering
surface
s!
grm,
16)
or
drug
binding
ro
the
mucosal
mem­
brane
surface26.
L)
i1
have
been
proposed.
However,
Anmo
et
al.
2F.)
using
a
recirculation
method
reported
that
the
absorption
of
salicylic
acid,
salicylamide
and
gentisic
acid
from
the
rat
oral
mucosa
agreed
well
with
each
dissociation
NVC
and
that
the
unionized
from
was
preferentially
absorbed
compared
with
the
ionized
form.
In
rhis
recirculario.
n
method.
the
recircularing
rate
of
t
h
i
d
r
u
g
solution
was
sufficient1)
y
rapid
and
rhe
solution
adjacent
to
the
absorption
surface
was
well
stirred.
Recently.
Tsuji
et
al.
29)
clearly
showrd
by
in
ritro
interphase
rransport
study
using
a
two­
phase
rolling
cell
that
the
aqueous
diffusion
layer
appeared
to
he
the
mosr
reasona­
ble
explanation
of
the
pH­
shifts
observed
in
penicillins
by
0.8­
2
pH
units
in
rar
stomach
and
small
intestine,
wirhour
application
of
physi­
ological
factors.
Although
the
experimentaf
proof
must
be
required,
it
is
difficulr
to
rule
out
rht­
efitst
of
such
an
aqutnus
diffusion
iayrr
on
thy
aSsorprion
irom
the
hamster
cheek
pouch
in
r
h
t
­
prritn:
expmmental
mechgd.
Thr
other
is
a
birspontnrial
loss
irom
rhr
lumen
of
the
cheek
pt)
uch
observed
ir!
the
rime­
course
study
of
salicylic
acid
absorption
{.
Fig.
')
Beckerr
and
Pickup3(
'!
a
h
s'ncwed
rht­
biexponenrial
loss
of
surne­
steroids
i
r
o
n
the
oral
cavity
in
man.
in
u.
hI:
h
the
d1s:
ribution
phasc
was
completed
u.
irhin
5
min.
The):
used
a
two­
comparrmmt
open
modei
which
meant
a
reversible
membrane
storage
t
e
explain
the
absorption.
Since
the
disrribution
phase
was
compiered
within
the
shvrrest
esprrimenral
period
of
15
min
in
the
present
scudy.
wc
coouId
not
consrrucr
an
ap­
propriate
modeling
under
this
condition.
Hon
­
eyer.
:he
mechacism
of
this
phenomenon
may
alsc
kt
responsibk
for
the
aqueous
diffusion
layer
a
well
as
rht
b
a
c
k
w
d
diffusion
from
the
fiscut
:o
:he
i
a
n
x
n
The
syremic
t
r
m
s
p
r
:
0:

5
".1
~
.,.­
7
~

.li
<'
'..<
S
C
I
~
a
t
t
t
r
intrn­
chtti­
pouch
zdminis­
,­
,.
295
tration
could
he
well
approximared
by
the
flrsr­
order
absorption
model
including
a
lag­
time
pro­
cess
and
the
absorption
rate
constant
esrimared
at
pH
3.0
was
about
5
times
larger
rhan
that
of
at
pH
4.0
(Table
H
i
).
Similar
results
were
ohrained
­j
from
the
rime
course
study
of
salicylic
acid
ah­­
!
sorption
(Fig.
1).
However,
since
the
pH
change
of
the
tissue
adjacent
to
the
blood
capillary
ap­
d
penred
to
be
negligible
(though
this
determina­
tion
has
not
been
accomplished
as
yet),
the
real
3
transfer
rate
constant
from
the
tissue
to
the
sys­
4
temic
circularion
is
assumed
to
br
independent
of
the
luminal
pH.
Taking
into
account
the
larger
tissue
accumulation
of
this
compound
at
pH
3.0
in
cornparkon
with
that
of
at
pH
4.0
(Fig.
SC).
the
larger
absorption
ratr
constanr
from
the
lumen
to
the
systemic
circulation
ob­
tained
at
pH
3.0
might
be
due
to
the
faster
trans­
fer
or
the
higher
partition
co
the
tissue
caused
by
rhe
lower
degree
of
ionization
at
pH
3.0
than
at
pH
4.0.
As
to
the
effect
of
molecular
weight
on
the
absorption
from
the
oral
cavity,
Siege)
cr
al.
reponed
that
the
permeability
coefficients
of
shon
chain­
length
alcohols
decreased
from
me­
thanol
to
propanol
and
then
increased
to
ocranol,
though
the
olive
oii­
to­
water
partition
coeffi­
cients
increased
simply
with
increasing
the,
chain­
length.
'"
'
They
concluded
from
the
in
vi^^^^^
and
the
in
vivo14)
experiments
that
the
absorption
parhway
of
oil­
soluhlt
compounds
is
a
transmembrane
route
whereas
water­
soluhle
molecules
with
a
molecular
volume
of
less
than
SO
crn'~
mmol
cross
primarily
through
rnrmbrant
pores
and
iarger
water­
soluble
molecules
pass
ex­
tracellulariy.
In
the
present
stud?.
a
mure
fayourable
correlation
could
be
obtajnrd
ht­

ween
the
ahsorption
rates
and
the
IipqMic
in­
dexes
when
the
molecular­
weight
factor
was
taken
inta
account.
This
is
one
d
t
h
t
reasons
uhy
the
absorption
from
the
krratinizcd
hamster
cheek
pouch
is
explained
by
cht­
passive
diffusion
mechanism.
A
new
experimental
method
using
a
hamster
cheek
pouch
proposed
in
this
paper
is
an
availa­
ble
method
for­
studying
both
the
absorption
and
the
i
d
l
~.~i
n
g
transfer
processes
to
rht­
systemic
circulation.
This
will
enable
us
to
investigate
the
nature
o
i
the
absorption
from
the
keratinized
oral
mucosa
in
various
severe
experimcncal
randitions
which
are
never
applied
to
human
voltlnrrers
in
the
buccz.
i
absorption
tt'sr.
Absorp­
tion
characterktia
through
the
keratinized
oral
branes:
The
physical
basis
of
ion
and
nonelectrolyrr
mucosa
may
be
clarified
by
this
hamster­
cheek­
pouch
method
in
rhe
near
future.
selr~
iviry,
Annu.
Rev.
Pbysiol..
31.581­
646
(19@~
t
161
'X1.
Schiirmann
and
P.
Turner.:
A
membrane
model
of
the
human
oral
mucosa
as
derived
from
buccal
a
b
q
,.

REFERENCES
A.
H.
Becketr
and
E
J.
Trigprs.
Buccal
absorption
of
basic
drugs
and
its
application
as
an
in
vivo
model
of
passive
drug
rransfer
through
lipid
membranes.
J.
Pharm.
Pharmacol.,
19,
Suppl.,
31S­
41S
(1967).
M.
H.
Bickel
and'H.
J.
Weder:
Buccal
absorption
and
other
properties
ofpharmacokinetic
imponance
of
imipramine
and
in
metaholites,
J.
Pharm.
Pharmucol..

A.
H.
Beckea
and
A.
C.
Moffat:
Correlation
of
partition
coefficients
in
n­
hepatane­
aqueous
systems
with
buccal
absorption­
data
for
a
series
of
amines
and
acids,
1.
Pharm.
Pharmarol..
21,
Suppl.,
144s­
150s
(1969).
A.
H.
Ekcken
and
A.
C.
Moffar:
Kinetics
of
buccal
ab­
sorption
of'some
carboxylic
acids
and
the
correlation
af
the
race
conscants
and
n­
heptane:
aqueous
phase'parti­
tion
coefficients,
J.
Pharm.
Pharmacol..
22,
15­
19
11970).
B.
K.
Berkovin,
G
.
R.
Hoiland
and.
B.
J.
Moxham
(e&.):
Oral
mucosa.
"A
Colour
Atlas
Bi
Textbook
of
Oral
Anatomy."
Wolfe
Medical
Publications
Lrd..
Hol­
land,
1978,
p.
136.
C
.
A.
Squjer
and
B.
K.
Hall:
The
permrabillry
of
skin
and
oral
mucosa
to
water
and
horseradish
peroxidase
as
related
ro
the
thickness
of
the
permeability
barrier.
J.
Invest.
Lkrrnarol..
84,176­
179
(1985).
M.
C.
Alfano.
J.
F.
Drummond
and
S.
A.
Miller:
LoCali­
zarion
of
rate­
limiting
barrier
to
penetrarion
.of
endo­
toxin
chrough
nonkeratinized
oral
mucosa
in
w
m
,
J.
Dent.
Res..
54.1143­­
1148(
19'5)
w'.
M.
Hill,
C.
A.
Squier
and
1.
E.
Lindrr:
A
hrsrolcgicd
method
for
rhe
v~
sualization
of
the
intercellular
per­
meability
barrier
in
mammalian
stratified
squamous
epithelia.
Hisrochrm.
J.,
14.64
1
­
(48
(I
9S2
i
.
1.
A.
Sirgel
and
K
.
T.
Izursu:
Ptrmeabdrry
oi
oral
mucosa
to
organic
compounds.
J.
Deni.
R
e
s
..
59.
21,160­
168
(1%
9).
tion
performance
and
physicochemical
properties
of
the
B­
blocking
drugs
atenolol
and
propranolol,
J.

Pharm.
Pharmarol.,
30,13­­
14:
(1778).
1')
F.
H.
W'hitr
and
K.
Gohart:
The
ulnascrucmral
mor.
phology
of
hamster
cheek
pouch
epithelium.
Arch
OrnlB/
ol..
26,563­
576
(1981).
18)
D.
F.
Evered,
F.
Sadoogh­
Abasian
and
P.
D.
Patel:
Ah.
sorption
of
nicotinic
acid
and
nicotinamide
acroS6
human
buccal
mucosa
in
vivo.
Life
Sci.,
27.

19)
Dl
F.
Erered
and
C.
Malletr:
Thiamine
absorprion
across
human
buccal
mucosa
in
vivo.
L
f
e
Sci.,
32.
1355­
1358
(1983).
2
0
)
M.
K.
Hunjan
and
D.
F.
Evered:
Absorption
ofgluw­
hione
from
the
ganro­
intestinal
cract,
Biochim.
BIP­

2i
1
M.
Tanaka,
N.
Yanagibashi,
H.
Fukuda
and
T.
Nagai:
Absorption
of
salicylic
acid
through
the
oral
muco\
Lz
membrane
of
hamster
cheek
pouch,
Chem.
Pharm.

22)
M.
lshida,
N.
Nambu
and
T.
Nagai:
Highly
viscousgel
ointmenr
conraining
carbopol
for
application
SO
thr
oral
mucosa,
Chern.
Phorm.
Bull..
31,4561
­4564
(1983.
2
3
)
G
.
M.
Oi'hidord
R.
S.
Callan
and
H.
S.
Wan&:
Fluoridt
absorption
rhrough
rhe
hamster
cheek
pouch:
A
pH­
dependent
event.
J.
Appl.
Toxicol..
1,303­
306
.
(1982).
'

,741
T.
Koizumi,
T.
Arira
and
K.
Kakemi:
Absorption
and
excretion
of
dtugs.
XX.
Same
pharmacokinetic
aspects
of
absorption
and
excretion
of
sulfonamides.
(2).
Ab­
.sorprion
from
rat
small
inrescint,
Chem.
Phorm.
BulL..
12,421
"427
<19&
4>.

3
1
D.
W'mne:
Shfi
of
pH­
absorption
cwves.
3.
Phorwce­
kine:.
Bmpharn;..
5,5.?­%
(l?
y­~.
26!
­l
W.
Bridges.
).
B.
Houscon.
hf
J.
Humpheq.
'X.
E
Lmdup.
D­
V.
Parkt.
J.
S.
Shilllngford
and
D.
G.
Fpsh­
all
Gasrrointcaxnal
absomtion
oicarknoxolone
in
rht
1449­
1651
(1980).

phJ73.
ACW
815,184­
188
(1985).

Bulf..
28.1056­
1061
(1980).

1601­
1605
11980).
IO)
1.
A.
Siegl.
Efien
oi
chemical
strucrae
r
m
n;
k­
iear.)­
lyre
penetration
of
oral
rnucosz.
1.
fnves;
lkrmnrr!..
76,
li7­­
140(
19S1).
11)
I.
A.
Sirgel,
K.
T.
lmnu
and
E.
Warson.
hlcchanwns
of
non­
elrcrrolyre
penetration
across
d
g
and
rabEtlc
or21
m
u
c
w
in
>,.
irru.
Arch.
OralBio1..
26,357­
361
(1981).
121
K.
Yamaoka,
Y
.
Tanigawara.
T.
Nakagaa­
a
and
1.
Uno:
A
pharmacokinetic
analysis
program
!ML'LTlj
for
microcompurrr,
1.
Phormacobio­
Dp..
4,

13)
T.
Yamana,
A
.
Tsuji,
E.
Mipamoto
and
0.
Kubo:
Novel
method
for
determination
of
parririon
coeffi­
cients
of
penicillins
and
cephalosporins
by
high­
pressure
liquid
chromatography.
J.
Phnrm.
Sri..
66,

14)
1.
A.
Siege]:
prrmeabiliry
oirht
rat
ora?
mucose
tc;
organic
soIutes
measured
in
viw.
,qr~%.
Or;
'
BI~,
'..
29.

15)
J.
M.
Diamond
and
E.
M.
%'rig$:
Bioio~~
ca_
l
mem­
879­
885
(1981?.

747­
749
(1977).

13­
16
(I9&­
4t.
rar
determined
in
vitro
and
in
siru:
Deriations
from
the
p!­
l­
partitdon
hypothesis.
J
.
Pharm.
Phorniawi..

2­
b
5.
Furusaaa,
K.
Okumura
and
H.
Sezaki:
Enhanced
r
n
l
­

pration
of
the
ionized
forms
of
aci&
c
drugs
from
water
into
chloroform
in
the
presence
of
phosphulipids.
1.
Pharm.
Pharmacol..
24,272­
276
(19721.
28)
1.
Anmo,
hi.
Washitake,
T.
Kurashigt,
Y.
Ozawa
and
K.
Kikuchi:
Srudies
on
rhc
absorpdon
through
the
oral.
mucous
membrane.
I.
Absorption
of
salicylic
acid
denratiws
from
the
rat
oral
mucous
membrane,
Yuku­
zaigoh.
28,113­
116
(1968:.
29)
A.
Tsuj"
E.
Miyamoto,
X.
Hashimoto
and
T.
Yamam.
GI
absxption
of
&laccam
anniiotio
II:
Deviation
from
pH­
partition
hypothesis
in
peniciliin
absorption
in
sin!
and
in
rim
lipoidal
barriers.
J.
Pharm.
Sci.,
67,
­

1705­
i'll
(1978).
A.
H.
btckerr
and
M
.
f
P
~c
k
u
~.
­4
mode1
for
a
r
r
o
d
rranspx
acros
biologics!
membrznes.
J.
€'hum.
Phar­
mco:..
27.1116­
Lli4
Clg­
jj.
28.11"
126
(1
9'6).
*'
Am&.
The
acute
oral
toxicity,
repellency,
and
m
d
potential
of
998
chemicals
to
one
or
more
of
species
of
wild
and
domestic
birds
was
deter­
mined
by
standardized
testing
procedures.
Red­
.;
irtg&
blackbirds
were
the
most
sensitive
of
the
bid
spcies
tested
on
a
large
number
of
chemicals,
on
index
based
on
redwing
toxicity
and
repel­
kMy
may
provide
an
appropriate
indication
of
the
probability
of
acute
avian
poisoning
episodes.
At­
ian
repellency
and
toxicity
were
not
positively
conejated
(Le.
toxicityvariedindependently
with
mpetlency).

In
a
program
designed
to
evaluate
chemicals
as
ptential
avian
toxicants.
stupefacients.
or
repei­
knts,
personnel
of
the
Wildlife
Research
Center
at
'Denver,
Colorado
have
tested
(since
1960)
­over
,mK,
chemicals
for
acute
oral
toxicity
to
one
or
more
species
of
wild
and
domestic­
birds.
The
pur­
p
c
of
this
paper
is
to
summarize
the
data
on
998
Lnown
chemicals,
draw
appropriate
generalizations
fiom
the
data,
and
make
recommendations
on
how
these
data
might
be
used
to
predict
acuteavian
pisoning
potentia?.

Methods
Thc
chemicals
included
technical
and
analytical
gtadc.
pcstici­
&I.
pharmaceutical.
and
other
commercial
or
experimental
compounds
that
were
either
purchased
or
solicited
from
caopcnting
firms.
For
presentation
purposes.
they
have
been
maneed
according
IO
Chemical
Abstracts
Registr).
Numbers
CCASI.
and
are
identified
by
an
accepted
trade.
coined.
produa
.........
.......................
......
...
or
other
chemiul
name
thu
is
a04
included
in
the
9th
Cdkaive
Index
of
Chemical
Abstms
Service.
'
Wild­
trapped
birds
we*
preconditioned
to
captivity
for
2
IO
6
weeks
and
were
usually
dosed
by
gavage
with
solutions
or
IW­
pensions
of
the
Ws!
chtmiul
in
propylene
glycol,
rccordiqL
IO
methods
d
e
s
c
n
i
d
by
Drcino
et
rrl.
WhX).
Schafer
(1972,.
rad
Schafer
e1
el.
(1%
7).
other
on!
dosing
methods
were
OCcIJiQII­
aIfy
used
(pellets.
whtin
upsuks)
but
are
not
noted
in
the
t.
Mer
(Schafer,
1972).
LD,
values
were
calculated
by
the
metbod
of
Thompson
(19411).
Thompson
and
Wcil(
1952).
and
Weil(
1952).
Repellency
tests
were
conducted
by
the
methods
of
Starr
rr
u/.
(1960
and
Schafer
and
Bmalon
(1971).
and
R;
s
(analogous
to
LD,
'rt
were
ulcuhred
either
bythe
method
of
tilchlield
and
Wikoxin
(1949)
or
Thompson
and
Weil(
l9S2).
A
mpeUency­
toxicity
indcx
(hazard
factor)
was
plculated
by
assuming
that
at
the
R,
kvel,
J
sixty­
live
g
male
redwing
watld
consume
50%
of
Ris
approximate
individual
maximum
food
a­
parit!
of
1
g
B>
m
a
h
p
'this
assumption.
i:
was
possibk
to
estim,
It
the
mg
Lg
of
a
chrrnlsal
that
could
concctuabl!
k
in­
.

gested
by
a,
redwing
tl
a
given
Rm
kvcl.
This
value.
when
divided
by
the
acute
oral
LD,
p~
ovides
an
index
for
indicating
bow
likely
it
would
be
for
acme
onl
poisoning
lo
occur
in
tbc
wild.
An
index
value
>
I
.W
indicates
weil­
accepted
toxic
agents
that
have
definire
potentid
for
causing
acute
poisoning
eplsodes.
an
iadcx
value
a0.25
61­
00
indmtes
these
compounds
witb
a
p
s
i
­
Me
gkxentiP1.
and
an
indu
nlw
~0.25
indicates
thoK
compounds
with
little
Of
no
.potentid
Io
cause
acute
avian
poisoning
cpiwdes.­
at
kast
io
redwiap.
Because
of
:be
hrpc
.mourn
of
data
accumuhted.
an
atrcmpt
was
made
to
determine
&e
signiicance
of
andlor
comltlion
bets­
een
the
tu0
of
Ihe
measured
parameters.
Statistical
com­
parisons
of
specks
xns*
iities
and
ranked
data
were
made
by
Friedmans
ranking
procedure
(Friedman
1937)
and
ANOVA
folloked
by
Duncans
Myiiipk
Range
Test.
Although
thc
clan­

parametric'
'Fricdrmns
pdedure
is
a
more
accurate
and
d
i
d
*
Because
of
the
kngrb
and
complexity
of
chemical
abstracts
nomenclature.
the
names
used
lo
identify
chemicals
in
fa&
2
.
were
extracted
from
several
sources.
Primary
consideration
was
given
lo
the
common
name.
but
shortened
chemical
names.
code
numbers.
or
registered
uademarks
were
also
used
but
may
not
be
specifically
identified
......
."
.......
".
Results
The
68
bird
species
tested,
along
with
their
cur­
.
rently
­acceptedscientificnames
and
a
four
letter
species
code
that
was
used
in
the
following
tabular
data,
are
detailed
in
Table
1.
Table
2
presents
a
tabular
listing
of
the
acute
oral
toxicity
(LD,
J
of
the
998
chemicals
to
one
or
more
of
three
avian
species
(redwing,
starling.
coturnix)
plus
:he
avian
repel­
lency
values
(R,)
and
the
toxicity­
repellency
index
for
redwings.
Redwing,
starling,
and
coturnix
data
were
analyzed
for
those
cases
where
LD,*
s
(other
than
c
or
values)
wereavailable
for
all
these
species
(n
=
73)
or
for
redwings
and
starlings
along
(n
=
130).
It
was
shown
that
redwings
were
sig­
nificantly
more
sensitive
than
starlings
(p
=
0.001).
and
that
starlings
and
coturnix
were
not
different
'
(p
=
0.05).
Thedifferenceintoxicological
sensi­
tivity
between
redwings
and
starlings
was
2.
Ix
and
the
difference
between
coturnix
and
redwings
was
1.4~.
This
agrees
with
previously
published
obser­

'
vations
of
therelativesensitivity
relationships
of
redwings
compared
to
other
wild
and
domestic
avian
species
(Schafer
1972;
Schafer
t
r
01.
1979).
~

Statistical
comparisons
of
the
correlation
be­
tween
redwing
LDds
and
R,
's
were
made
to
de­
termine
the
validity
of
observations
made
over
the
past
20
years
indicating
that
avian
repellent
activity
appears
IO
increase
with
increasing
acute
oral
tox­
icity.
Ofthe
998
chemicals
tested,
redwing
R,
's
and
LI),
's.
are
presented
for
836.
Qf
the
836,
Rw
and
LDs
values
for
501
chemicals
(60.0%)
were
both
greater
than
selected
minimum
activity
levels
(F.
0095
for
R,
and
100
m
g
Q
or
(90
mglkg)
for
LD&
84
(IO.
1%)
were
repellent
at
or
below
1
.WC
but
toxic
above
I
0
0
m
a
g
,
75
(8.9%)
were
toxic
at
or
below
100
m
a
g
but
repellent
above
1.00%.
41
(4.9%)
were
not
usable
and
135
(16.2%)
possessed
activity
in
the
range
(R,
1.00%
LDSO
sz
100
hglkg)
that
could
be
used
lo
examine
the
;elmti*
ship
between
these
two
facton.
However,
n
e
i
h
Parson
or
Spearmancorrelationcoefficients
(0.33
­
and
0.43,
respectively)
showredmy
positive
corn
.
lation
between
R,*
r
and
LD?,
's.
Thus,
the
dab
.

dicatethat
gross
acute
toxiaty,
as
defined
by
­_
LD,
is
not
positively
related
to
gross
as
defined
by
the
RY,.
at
least
over
the
small
nqe
examined.
The
repellencyftoxicity
index
or
acute
avipp
hazard
index
was
calculated
for
377
cbemicdr
I
where
one
or
both
R,
and
LD,
values
were
knom.
:.

Those
chemicals
for
which
the
LD,
and
R,
wen
only
known
to
exceed
some
value
could
not
be
u~
in
subsequent
calculations
since
no
meaningful
value
or
trend
could
be
determined
by
the
index.
01
the
223
chemicals
for
which
definite
index
v
d
u
a
could
be
calculated,
124
fell
into
the
>
1
.@
I
class,
47
­
intothe
20.25
d
1.00
class
and
52
intbe
~0
.3
­
class.
Examples
of
some
chemicals
in
tbe
>].
a
­
class
(hazardous)
are:
Mitomycin
C.
TEM,
thiotepa,
famphos,
parathion,
and
dimethoate.
Examplesof
chemicals
in
the
possibly
hazard­
'­.

class
(30.25
6
1
.OO)
are:
coumaphos,
aprocarb,
feb
sulfothion,
fenitrothion,
and
malathion.
Exampla
.;
._
of
chemicals
that
fall
into
the
probably
nom­
hazardous
class
(~
0.25)
are:
lidane,
sulphenone,
chlorpropham.
thiram,
and
chlorothion.
This
index
appears
to
have
great
potential
for
predicting
those
chemicalsthatmay
cause
acute
avian
poisoning
episodes
in
thefield.
It
is
the
first
time,
to
ow
knowledge,
that
an
attempt
has
been
made
to
J
equate
potential
hazards
to
an
index
that
combiner
the
toxicity
of
a
compound
with
a
behavioral
mea­
sure
that
predicts
how.
much
of
the
chemical
could
porentially
be
consumed
in
a
fieldsituation.
Thus,
1
fieldapplicationof
a
highlytoxicchemicalthat
is
'
­'
aversive
to
birds
could
have
the
same
or
less
likeli
hoodofinducing
acute
avianpoisoning
as
a
less
5
toxic
chemical
that
was
more
readily
accepted.
Table
3
presents
acute
oral
toxicity
data
of
82
.

chemicals
to
one
or
more
of
seven
additional
avian
species.
Table
4
presents
the
acute
oral
toxicity
and
repellency
data
of
90
chemicals
to
one
or
more
of
58
other
species
of
birds.
*

~a
b
k
­1.
Species
code.
common.
tad
scientific
names
of
birds
referred
10
in
this
paper
Species
code
Common
name
Scientific
name
d
e
s
American
kestrel
(F
u
h
spon­
crius)
bbv
Blue­
black
grassquit
(Vdariajucorinu)
bbmP
Black­
bt&
d
magpie
(Pica
picu
)

bhcb
Brown­
headed
cowbird
(Afoforbrus
urcr)
bjay
Blue
jay
(C.
vunocirru
rristaru)

brcb
Bronzed
cowbird
(fongm­
ius
urnrus)
bowl
Barn
owl
(T
~I
u
o
h
)
I
!
i
..

.
",
.
i­>­.­.
i7
..:.
.
.
­
,...
­,
..
..
.
.
.
,
,
.
.
.
.
.
..
.
.
.
.
"
.
Brown
thnshcr
Bat­
taikd
qackk
Brown­
throated
conure
Budgerigar
Common
bobwhite
Bluc­
winged
leal
Curve­
billed
ihnsher
Ameriun
crow
Cassins
finch
Canada
goose
Common
grackle
Plain
chichalam
Cooper's
hawk
Cownix
Rock
dove
or
common
pigeon
Norihern
raven
Cedar
wuwin5
Dickcissel
Eared
dove
Goldensrowned
spamow
Common
or
pound
dove
Golden
cagk
Cddea
spamow
Green
jay
House
finch
Homed
lnrk
HOUK
spurow
l
n
u
dove
LprL
bunting
Mallard
Mourning
dokc
Nonhem
bvrier
(Marsh
hawk)
Monk
parakeet
Sonhern
masked
weaver
Orurge­
fronted
conure
Common
pintail
Red
bishop
Ring­
billed
gull
Ruddy­
breasted
seedeater
Red­
cycd
cowbird
Ruddy
pound
dove
Ripg­
wkd
ph­
t
AmcTiunrobin
Red­
winged
blackbird
Scrub
p
y
scpty
dove
Shinycowbi
Sandhill
crane
European
starling
Swainson's
hawk
Tricolored
blackbird
Wild
turkey
California
qurj
.
Wage
weaver
Whitt­
crowncd
sparrow
White­
fronted
dove
Yellow­
beaded
blackbid
.YeUow­
billed
maspie
Red­
baed
qWlU
.
.

White­
wing4
dove
1
E.
W.
Schlct.
Jr.,
d.

8
\
!
Y
Ci
V
g
K
d
.
#I
..
.~.
...
..
i
­
Y
r
++
++

......
...........
i
:.
....
."~
.........
.­,._
:
.....................
..
.­
.......................................
M2
E.
W.
Schrlct,
kr.,

1
+++1
++

I
..

..
....
........
..
..
...........
..
.­
I
+++

­.
..
I
.
i
..
i
­*

.a
2
,
i
:
i
:
P
E.
W.
Schrfcr.
Jr..
et
d.
369
I
1
1
1
1
l
l
l
l
l
W"
".
."
­
..

37
I
Q
0
8
0
+
I
++
I
+.I
++
I
?++++
+
+I
++
++
+
+
+I
+++
++I
++
I
+
+
1
1
372
=
­
E
3
II
ci
Y
F
l
+
I
I
U
n
a
I
I
+++
++

3
s
0
I
I
I
+*

=s
I
I
f
*++
+
+
+
+.
+
++
++
+

1
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.
I
374
1
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375
4
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n
I
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A
i
Arknn*./
rd~
mrats.
fhc
ruthors
wish
lo
thank
the
following
i
d
­
viduak
for
astktunce
in
colkcting.
typin;.
and
editing
the
data
contained
in
this
puhlicarion;
Trudic
Abendroth.
Lore
Burbach.
Donald
Cunningham.
Patrick
Fundetbuq.
J
a
~t
Garcia.
Thomas
Hall,
hvid
Hayes.
Glenn
Hood.
Ccccli.
Nelson.
and
Barbara
Recktenwald.
The
preparation
md
compbtion
of
this
data
was
supported.
in
pan.
throughInteragencyAgreement
Ill­
16
Oooy­
81.957
whichwasfundedbythe
U.
S.
Environmental
he
tection
Agency.

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T.
1..
D.
J.
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m
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E.
W.
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Toxicity
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J.
Wildl.
Manage.
30,
249
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F.
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aduk
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C.
S.:
Tables
for
convenient
ulcuiation
of
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liw
dcwt
tLD5..
or
ED,
*
andinstructions
in
the::
use.
B~
omclncs
8.24Y
(19521.
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