Effect of Sucrose Addition on the Sugar and Sorbitol Composition
of Frozen Sweet Cherries and Their Derived Concentrates
CHRISTOPHER
J. CORNWELL,
RONALD
ABSTRACTGlucose, fructose, sucrose, and sorbitol contents of Van and Black
Republican varieties of sweet cherries were determined by high
performance liquid chromatography (HPLC) and enzymic analytical
procedures. Samples analyzed included frozen fruit, fruit packed as
3.6 parts fruit plus 1 part added sucrose,and cherry juice concentrates
derived from the sugar-packed fruit. Cherry fruit showed invert
patterns of glucose and fructose and contained trace amounts of
sucrose, and 2.6-3.9g sorbitol/lOOg. Sucrose was either not detected
or found in trace amounts in samples to which sucrose had been
added. There is evidence that sucrose hydrolysis was caused by
presence of invertase. The percent sorbitol content could be used to
detect addition of sucrose. Results for sugar and sorbitol content as
determined by HPLC or enzymic methods were very similar.
INTRODUCTION
A FREQUENT
COMMERCIAL
practice is to pack sweet
cherry fruit in 55 gallon drums with added sucrose and hold
in frozen storage for subsequent use in yogurts, ice creams,
sauces, bakery items, etc. The free-run juice produced when
the barrels are thawed is rich in color and flavor and yields
a high-quality syrup when concentrated. The major objective
of this study was to determine whether addition of sucrose
to cherry fruit could be detected in the frozen samples and
derived juice concentrates through analyses of their fructose,
glucose, sucrose, and sorbitol contents. Wrolstad and
Shallenberger (198 1) recently published a table listing the
free sugar and sorbitol content of sweet cherry fruit as
reported in the literature
by various workers. Sweet cherries
have essentially an invert pattern, containing nearly equal
quantities of glucose and fructose and either no or trace
levels of sucrose. Their sorbitol content is among the highest
of the various commercial fruits. Richmond et al. (198 1)
analyzed the sugar and sorbitol content of several fruits by
high-performance liquid chromatography (HPLC) and
reported a similar pattern for sweet cherry fruit. Another
objective of this study was to compare the effectiveness of
enzymic analytical procedures with HPLC for glucose,
fructose, sucrose, and sorbitol measurements. HPLC instrumentation requires considerable investment while enzymic
analysesutilize equipment which is more commonly available
in industrial laboratories.
EXPERIMENTAL
Sample
Frozen samples were obtained from The Dalles Cherry Growers,
Inc., The Dalles, OR 97058, and stored at -15°C until use. Samples
included individually quick frozen unpitted sweet cherry fruit (Van
and Black Republican varieties, 1978 season) and two lots each of
the same fruit packed as 3.6 parts fruit and one part added sucrose.
Also provided were samples of cherry juice concentrate which
had been manufactured from frozen Black Republican fruit packed
with added sucrose.The fruit which had been frozen in 55 gal drums
was thawed at ambient temperature and then heated to 60°C.
Authors
Cornwell,
Wrolstad, and Reyes are with the Dept. Of Food
Science & Technology,
Oregon State Univ., Corvallis,
OR 97331.
Author
Reyes is on sabbatical
leave from Universidade
Estadual De
CampinadF.
E.A.A., Campinas, S.P. Brasil.
E. WROLSTAD,
and FELIX
G. R. REYES
The concentrate was made from the freerun juice. Three lots
of concentrate were from the 1978 season, one lot was from 1977
and an additional lot was from the 1976 season. All concentrates
were 5 1” Brix.
Extraction and isolation of sugarsand sorbitol for HPLC analysis
Fruit samples were homogenized in a Waring Blendor. Forty ml
95% ethanol were added to lO.Og of homogenate and the mixture
was sonicated for 2 min through direct immersion of an ultrasonic
probe (Bronwill Biosonik III Ultrasonicator; dial setting of 70).
Juice concentrate samples were diluted (25.Og + 75.0 ml distilled
water), and a 10.0-galiquot combinedwith 40 ml ethanol.Samples
were held at 2°C for 1 hr to facilitate precipitation of high molecular
weight components, and then filtered by suction through a pad of
log of hydrated polyvinylpyrrolidone (PVPP) on Whatman No. 1
filter paper; the PVPP had been washed by the procedure of Loomis,
1974. The PVPP pad and sample residue were rinsed with 25 ml
80% ethanol three times; the filtrate and washings were combined
and concentratedto about 5 ml on a rotary evaporator(water bath
= 3?C, pressure = 23 torr) to ensure ethanol removal from samples.
The samples were passed through two 2 x 7 cm columns in series,
the first column containing 6 ml Biorad AG 50 W-X4 cation exchange
resin, (200-400 mesh, Na+ form) and the second containing 6 ml
Biorad AC 1 X-8 anion exchange resin (200-400 mesh, F- form).
The columns were washed with deionized distilled water and the
eluantscollectedin 50 ml volumetric flaskscontaining5 ml of 10%
mannitol as an internal standard and 1.0 ml 0.5% CaCl2’2H20.
Samples were filtered through 0.45 I.rrn Millipore filters prior to
injection.
HPLC analysis of extracts
The HPLC system was composed of a Varian 5000 liquid chromatograph equipped with a column heater and refractive index
detector, a 7.8 X 300 mm Aminex HPX-87 column (Biorad Labora-
tories), and a Hewlett Packard3380 A integrator.Column temperature was maintainedat 85°C. The mobile phasewasdeionized,0.45
urn Millipore filtered water containing 0.1% CaCl,*2H?O: the flow
rate was 1 ml/min. Injection volume was 20 ~1. Refractive index
range was 0.5 x 1O-5 RI units.
Relative detector response (RDR) factors for sucrose, glucose,
fructose, and sorbitol were determined to be 1.01, 1.03, 1.04, and
1.07, respectively, using the method described by Akhavan et al.
(1980). Percent recoveries (R) for standards were determined by
submitting reagent sugar solutions in 80% ethanol through the
isolation and separation procedures; percent recoveries for sucrose,
glucose, fructose, and sorbitol were 98.7, 98.3, 95.5, and 98.3,
respectively. Content of individual sugars was calculated by the
following formula:
g sugar/lOOg sample = (As)(Wtis)(DF)/(Ais)(R)(RDR)
where As = peak area of sugar; Ais = peak area of internal standard;
W& = weight of internal standard; DF = dilution factor; R = percent
b
.
recovery;and RDR = relativedetectorresponse.
Enzymic analysis of sugarsand sorbitol
Sugarand sorbitol contents were determinedenzymically with
Glucose/Fructose, D-Sorbitol,
and Sucrose/Glucose kits from
BoehringerMannheim Biochemicals,IndianapoIis,IN, accordingto
the manufacturer’s instructions. Samples (2-Sg) were homogenized
in a Waring Blendor and diluted with 2L distilled water; a 50-ml
aliquot was centrifuged in a Sorvall RCZB centrifuge at 12,061 x
g for 10 min and a lo-ml aliquot of the supernatant diluted 25-fold.
After combining 2-ml aliquots with reagents absorbance readings at
340 nm were determined on a Perkin Elmer model 550 spectro-
photometerand the concentrationcalculated.
-Continued
Volume
47 (1981)--JOURNAL
OF FOOD
on next page
SCIENCE-281
CHERRY
I
i
I
I
SUGAR
COMPOSITION.
..
Determination of invertase activity in Black Republican fruit
Presence of invertase activity in the Black Republican frozen
fruit sample was determined from glucose analysis of the following
samples: (1) lO.Og cherry fruit; (2) lO.Og cherry fruit plus 5.Og
sucrose; (3) 5.Og sucrose. All samples were made up to 100 ml with
pH 4.5 acetate buffer and incubated at 50°C for 25 min. Glucose
determinations were made using the enzymic procedure.
RESULTS
& DISCUSSION
FIG. 1 SHOWS typical HPLC chromatograms
of cherry
extract samples and Table 1 lists the free sugar and sorbitol
content of all samples aa determined by HPLC. The fruit
samples had invert sugar patterns, containing nearly equal
quantities of glucose and fructose. Sorbitol was found in
Van Cherries
+ Sucrose
Van Cherries
man
I-HPLC
chromatogram
of extracts
of Van cherries
and Van cherries
plus
added
sucrose.
Column
* Aminex
HPX-87; mobile phase = water containing
0.01%
CaC12; flow rate = 1 ml/min;
injection
volume = 20 pl. Typical retention
times:
Sucrose
(sue), 4.70 min;
glucose
(glul. 5.98 min; fructose
ffru),
8.08
min;
mannitol-internal
standard
(man), 10.65 min; sorbitol
(sorb), 13.53
min.
Fig.
fru
sorb
lLA
2
sue
1 -Free
sugars and sorbitol
Fructose
Sample
Van frkt
Mean
Black
Republican
gll 00s
% C.V.
% T.S.
s/l 00s
6.38
6.80
6.59
5.01
1.32
4.50
46.1
7.32
7.55
7.44
1.78
1.19
2.15
8.86
6.93
6.79
3.15
1.44
2.79
44.9
8.04
8.31
8.18
2.1 1
0.48
2.32
14.99
49.6
15.55
15.26
15.40
1.03
0.46
1.29
49.1
15.04
15.34
15.19
0.66
2.48
1.38
48.6
14.73
14.99
14.86
2.38
2.80
1.21
48.3
13.99
13.74
13.86
0.79
2.69
1.30
49.8
21.22
21.41
21.31
2.59
1.86
0.81
48.9
23.22
23.16
23.19
2.24
0.99
0.17
48.8
22.80
22.12
22.46
1.80
2.03
2.14
49.4
19.64
19.03
19.34
3.56
0.68
2.22
48.7
21.61
21.14
21.37
0.46
1.47
1.54
fruit
Mean
Van Fruit
(Lot 1)
Mean
+ Sucrose
14.93
0.93
0.27
0.50
Van Fruit
(Lot 2)
Mean
t Sucrose
14.64
14.55
14.75
0.27
2.09
1 .Ol
Black Republican
fruit
+ Sucrose (Lot 1)
Mean
14.03
14.55
14.05
2.85
4.64
7.11
Black Republican
fruit
+ sucrose (Lot 2)
Mean
12.95
12.97
12.96
1.16
1.54
0.10
Syrup from
Republican
Mean
Black
(Lot 11 (19781
21.02
21.02
21.02
3.28
1.09
0
Syrup from
Republican
Mean
Black
(Lot 2) (1978)
22.26
2
2.92
0.95
0.49
Syrup from
Republican
Mean
Black
(Lot 31 119781
21.89
20.98
2(.44
3.85
2.95
2.98
Syrup from
Republican
Mean
Black
(Lot 4) (1978)
19.38
1m
19.03
2.89
0.58
2.57
Syrup from
Republican
Mean
Black
(Lot 5) 11977)
20.57
20.25
20.41
1.31
1.63
1.12
a % C.V. = pa;cent
b fructose ratlo.
282-Volume
14.87
coefficient
47
Of variance;
(1981)-JOURNAL
%T.S.
of cherry
samples
determined
Glucose
= percent
OF
total
FOOD
% C.V.
wgars;
total
SCIENCE
I
I
I
1
0
5
IO
I5
min
by HPLP
sucrose
% T.S.
52.1
54.1
51.2
50.6
51.4
51.7
50.3
51.1
51.2
50.2
51.0
g/l 009
% C.V.
% T.S.+S
14.29
2.81
2.57
2.59
3.45
3.89
1.16
15.3
15.13
3.94
3.88
3.91
1.01
5.41
1.02
20.5
30.09
1.92
1.99
1.96
7.81
9.64
0.05
6.5
30.02
2.26
2.10
2.18
6.64
8.57
5.04
6.8
28.91
2.85
3.02
2.93
9.82
6.29
4.09
9.2
26.82
3.08
2.76
2.92
2.59
10.14
7.88
9.8
42.33
3.23
2.91
3.07
10.52
10.31
7.49
6.8
45.38
3.18
3.30
3.24
10.69
4.84
2.47
6.7
43.90
3.22
3.28
3.25
7.45
6.40
1.23
6.9
38.50
3.52
u
3.45
9.37
1.77
2.89
8.2
0.2
41.87
3.68
3.54
3.81
5.16
12.99
2.77
7.9
%T.S. + 6 = perc**t
tota,
g/l oog
% C.V.
% T.S.
1.13
0.27
0.26
0.26
3.70
3.64
3.84
1..8
1.20
0.16
0.15
0.18
18.7
13.3
6.25
1.03
0.04
0.08
0.06
1.05
1.07
1.01
1.04
50
0.2
b
&
b
0
0.3
0
0
0
0
0
0
0
1.05
0
1.02
0.15
0.11
0.13
b
b
23.1
b
1.05
0.18
0
0.09
suga,s = g1ucoH) + f,“ctcse
1 .l
b
b
0.16
0
+ s.“c,osa;
b
Sorbitol
Total Sugars
gl1Wg
Glu/Fru
1.03
n
--ii
Time,
Table
sorb
.-C
0.3
*ugars + so,b,to,;
g,u,tru
= glucose:
I
large amounts while only small quantities of sucrose were
found. This compositional pattern is consistent with the
sugar and sorbitol content for sweet cherries as recently
compiled from the literature by Wrolstad and Shallenberger
(1981). The fructose, glucose, sucrose and sorbitol content,
the glucose:fructose ratio, and the total sugar content of
the Van and Black Republican samples are within one
standard deviation of the mean values reported in the literature compilation. There is a greater difference between the
sorbitol contents of Van and Black Republican varieties
(15.3 and 20.5% of total sugars plus sorbitol, respectively)
than there is between fructose, glucose, and sucrosecontents.
The most striking information in Table 1 is that either
no or a very low level of sucrose was detected in the samples
to which sucrose had been added. Clearly, sucrosehydrolysis
is occurring during processing and/or storage. Glinskaya and
Shkurina (1969) have previously reported invertase activity
in cherry fruit. The results presented in Table 2 demonstrate
the presence of invertase activity in the Black Republican
fruit sample where 58% inversion of sucrose occurred after
25 min incubation. It is highly likely that invertase activity
prior to freezing, during storage, and/or during thawing
accounts for there being little or no sucrose in the various
samples.
The glucose:fructose ratio in Van and Black Republican
fruit, 1.13 and 1.20, respectively, is very close to the mean
value for sweet cherries, 1.17 (standard deviation = 0.357),
compiled from the literature (Wrolstad and Shallenberger,
1981). Sucrose hydrolysis in the samples containing added
sucrose lowered the glucose:fructose ratio to values ranging
from 1.01-l .07, which is still within the range reported in
the literature. The glucose:fructose ratio has very limited
value in detecting added sucrose in these samples as the
fruit itself has close to an invert pattern.
Cherry fruit is very high in sorbitol, but there can be
considerable quantitative variation with variety and season.
The sorbitol content of Van and Black Republican cherries
in this study was 15.3 and 20.5% of total sugarsplus sorbitol, respectively. Neubeller and Stosser (1977) studied the
sugar and variation of 20 varieties of sweet cherries over 3 yr
and reported the sorbitol content varied from 15.5 - 24.0%
Table 24ucrose
Inversion
Fruit
Sample
by Black
Sucrose
Republication
Fruit
addition
Measure
(cl)
(cl)
1
10
2
3
10
0
5
5
0.76
2.44
0.16
0
% inversion
sample
=
glucose
2 glucose
yield
-
from
(sample
100%
Table d-Free
sugars and sorbitol
1 + sample
hydrolysis
XlOO=$$$f=
of cherry
samples determined
Fructose
% T.S.
analysesa
Sucrose
1 .oo
0.78
% T.S.
GWFru
g/lOOg
%C.V.
%T.S.
Total
Sugars
Sorbitol
g/lOOg
%C.V.
2.65
3.61
-2.52
2.64
6.46
%TS+S
g/l OOg
% C.V.
48.3
6.62
0.46
51.9
45.9
7.82
0.78
0.85
54.2
51.7
1.08
1.18
1 .OJ
n.d.
n.d.
n.d.
0
0
0
27.9
13.92
1.97
51.7
1 .OJ
n.d.
0
26.9
3.05
0.76
10.2
21.5
0.28
50.4
1 .Ol
n.d.
0
42.7
3.70
1.57
8.0
6.13
6.61
13.48
0.46
48.3
14.42
13.00
1.89
48.3
21.22
0.58
49.7
coefficient
+ sorbitol):
by enzymic
Glucose
% C.V.
Van fruit
Black Republican
fruit
Van fruit + Sucrose
(Lot 11
Black Republican
fruit
%c.V.
= Percent
fructose
+ sucrose
sucrose
58%
gllOOg
+ Sucrose (Lot 1)
Syrup from Black
Republican
+ Sucrose
/Lot 1)
3 glucose)
of added
Sample
a
glucose
M
of total sugars plus sorbitol. Sucrose addition has a dilution
effect on sorbitol content; the percent sorbitol content in
all samples with added sucrose is reduced from the percent
sorbitol of the fruit samples by more than one standard
deviation of sorbitol variation as compiled from the literature (Wrolstad and Shallenberger, 1981). The low percent
sorbitol content of the samples with added sucrose (6.59.8%) is the best indication in this study that those samples
are not derived 100% from fruit. Since the fruit samples
with added sucrose were packed as 3.6 parts fruit and one
part sucrose, the percent sorbitol would be estimated to be
5.8 and 8.4% for the Van and Black Republican samples,
respectively. This is somewhat lower than was actually
measured in all four lots of fruit packed with sucrose.
Nonuniform distribution of sucrose in the drums and nonrepresentative sampling might account for this discrepancy.
The syrup concentrates were produced from the free-run
juice of the sugar packed fruit. The percent sorbitol levels
of the three lots produced from 1978 seasonBlack Republican fruit is lower than the 8.4% level of sorbitol estimated
to be the percent sorbitol content of the sugar packed fruit.
These results suggestthat there is not complete diffusion of
the sorbitol from the fruit to the juice and the free-run
juice from which the concentrate is made contains a somewhat higher proportion of sucrose than the sample as a
whole.
Duplicate runs were carried out for all samples analyzed
by HPLC and the value for each duplicate is shown in Table
1. The percent coefficient of variance (%C.V.) listed for
each replicate represents the variation among the three
HPLC injections of the sample extract. The %C.V. for
individual sample extracts was less than five percent, except
for some samples of low sucrose or sorbitol content. The
%C.V. listed for the mean value of each sample is the variation
between the two duplicates and would represent variation
due to sampling, extraction, and isolation. Variation
between duplicates is low, being less than five percent for
most samples.
Table 3 lists the free sugar and sorbitol content for five
of the samplesas determined by enzymic methods. Duplicate
runs were not made and the %C.V. represents the variation
between analytical determinations for the same extract.
This variation is low. Sucrose was not detected in any of
the five samples by the enzymic procedure. As the enzymic
assay for sucrose measuresthe increase in glucose concentration after sucrose hydrolysis, it will not be a sensitive
procedure for determining trace amounts of sucrose in
presence of large quantities of glucose. Results for fructose,
glucose, and sorbitol content as determined by the enzymic
(Table 3) and HPLC methods (Table 1) are very similar.
The correlation coefficient for fructose content as determined by the two methods for the same five samples is
0.994; for glucose the correlation coefficient is 0.997 and
for sorbitol it is 0.847.
-Continued
on page 290
of variance;
% T.S. = Percent
total sugars
Qlulfru
= glucose:fructose
ratio; n.d. = not
(glucose
detected.
+ fructose
Volume
+ sucrose);
% T.S.+S
= Percent
47 (1981kJOlJRNAL
12.7
14.4
total
sugars
O F FOOD
+ Sorbitol
17.2
20.0
8.3
(glucose
SCIENCE-283
+