A Research Note

HPLC Determination of Fructose, Glucose,

and Sucrose in Potatoes
A. M. WILSON, T. M. WORK, A. A. BUSHWAY, and R. J. BUSHWAY

- ABSTRACT .The column system consisted of a 3.2 m m i.d. x 4.0 cm precolumn

A high-performance liquid chromatographic (HPLC) method has ,I packed with cl8 Porasil B (Waters Associates) and a 3.9 m m i.d. x
been developed to (etermine fructose, glucose, and sucrose con- 30, dm column packed with bBondapak/carbohydrate packing.
tent of potatoes. Tl~e HPLC system consisted of a pBondapak/ The mobile phase was acetonitrile-water (75 :25). Operating condt
carbohydrate columr , a solvent system of acetonitrile-water (75: tions. were flowrate: 1.8 ml/min, detector sensitivity: 8X, and chart
25), a flowrate of 1.1: ml/min, and a refractive index detector. An- speed: 0.4 in/min.
alysis,including samfle preparation,was completein 30 min. With
the exception of big! concentrationsof sucrose(8 mg/g or above), Standard preparation
the method recoverer! 93% or more.of all sugars.The coefficients of A standard containing the three sugars (fructose, glucose, and
variation for the prcpcedureranged from 1.39-13.31s. TLC indi- sucrose) was prepared by placing 250 mg of each sugar in a 25
cated that there wer,: no interfering compounds eluting with any ml volumetric flask and dissolving them in water. A 10 ml aliquot
of the three sugars. was removed and mixed with 10 ml of methanol followed by
centrifugation at 43,000 x G for 5 min (Sorvall RC2B, DuPont In-
struments, Newtown, Conn). After passing the supernatant through
INTRODUCTION a C18 Sep Pak (Waters Associates), the first milliliter was discarded
and the next 2.5 ml was collected. A standard curve was prepared
INDIVIDUAL DE::‘ERMINATION of glucose, fructose, and each day by injecting 2, 6, 10, 14, and 18 pl of the standardsolu-
sucrose, the major sugars in potatoes, is becoming essential tion. Quantification of each of the sugarswas accomplished by com-
as more studies a:.e being ‘conducted on genetic and bio- paring peak height of the samples to peak height of the standard
chemical mechanhms of carbohydrate formation and de- since peak height was directly proportional to concentration of the
gradation in potato tubers. There is also an interest in standard throughout the concentration range used.
predicting the storage potential of tubers based on their
sucrose content a: harvest (Sowokinos, 1978) and in an- Sample preparation
alyzing sugar cant :nt of potatoes stored at various condi- Cultivars analyzed were Kknnebec, Katahdin, and Russet Burbank, I
tions. all stored at 3.3 and 7.2”C, and Monona and Norchip, which were
Previous method:; include calorimetric procedures which stored at 12.8”C. A center cut from each of six tubers of uniform
do not distinguish between the monosaccharides, glucose size was removed, peeled, and diced. A 200g random sample was
and fructose, direl:tly. Furuholmen et al. (1964), using a
modification of Ti ng’s method (1956), determined glucose
concentration as the difference between total reducing
sugars and fructose. Della Monica et al. (1974), using
Furuholmen’s procedure and a glucose oxidase method,
quantified fructo: e by subtracting glucose determined
enzymatically fro n total reducing sygars. Sucrose was
determined as the increase in glucose content after acid
hydrolysis of sucrose.
Chromatographic procedures include gas chromatography
which differentiatlss between the three major sugars but I
requires a time-consuming derivatization step (Kimura et
al., 1969; Shaw, 1969; Varns and Shaw, 1973). HPLC b
methods determinc: the individual sugars and do not require
derivatization. Currently there are HPLC methods available
to analyze sugars in boiled sweets and jelly (Hunt et al.,
1977), milk and ice cream (Warthesen and Kramer, 1979),
milk chocolate (Hurst and Martin, 1977), and confection-
ary products (Tintbe and Keeney, 1977). This paper de-
scribes and HPLC procedure for analyzing fructose, glucose
and sucrose in pot; .toes.

MiiTERIALS& METHODi
Liquid chromatograpl~y
A Waters Associates liquid chromatograph, model ALC/GPC 244
(Milford, MA), equippedwith a 6000A pump, a U6K injector, and
a differential refractometer, was used. The detector signal was re-
I
r
-

1
J L
1 r I

corded on a HoustolI Instruments (Austin, TX) dual-pen recorder. 012345678

T IME(min)
All authors are affiliated with the Food Science Dept., Univ. of Fig. l-Chromatogram of fructose, glucose and sucrose present in
Maine, Orono. M E 0’ 469. Russet Burbank tubers stored at 3.3oC. Peaks: (al fructose, fbJ
glucose, (cl sucrose.
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300-JOURNAL (IF FOOD SCIENCE-Volume 46 (1981)

 HPLC ANAL YSIS OF POTATO SUGARS.. .

Table l-Reproducibility study of the HPLC method for sugars in three cultivars at two storage temperature?

Fructose Glucose Sucrose

Storage
Cultivar temp (“C) Mean (mg/g) SD Mean (mg/g) SD Mean (mg/g) SD

Kennebec 3.3 5.91 0.55 6.76 0.69 2.26 0.24

7.2 0.78 0.06 1.56 0.10 0.65 0.06
Katahdin 3.3 8.01 0.11 8.87 0.29 2.61 0.12
7.2 1.06 0.04 1.73 0.14 0.78 0.05
Russet Burbank 3.3 6.75 0.34 8.02 0.36 2.57 0.34
7.2 0.77 0.06 1.55 0.10 0.65 0.06

a Data are bpd on the analysis of five samples for Ka$,ahdin potatoes stored at 3.3’ and 7.Z°C and for Russet Burbank and Kennebec tubers
stored at 7.2 C. For the latter two varieties stored at 3.3 C, six samples were analyzed.

extracted in an Acme Juicerator (Lemoyne,Pa). A 10 ml aliquot of
the juice was preparedas was the 10 ml aliquot of the standard
described in the Standard Preparation section. (The methanol
was used to precipitate the protein in the samples.) The amount
of sample injected into the HPLC ranged from 6-20 ~1 depending
upon the concentrationof the sugarsin the sample.
Recoverystudies
The recovery of added fructose, glucose, and sucrose at two
levels (2 mg/g and 8 mg/g) was determined by dividing the juice of
200g of Kennebec potatoes into two equal portions with one
portion having no sugaradded while the other was spiked at the
appropriate level. These samples were prepared as describedin
the Sample Preparation section. The recoveries were calculated
based on the difference between the total amount determined in
the spiked samplesand the amount observedin the non-spiked
samples.
RESULTS& DISCUSSION
FIGURE 1 shows a typical separation of fructose, glucose,
and sucrose in potatoes. With a new column, a baseline
separation between the two monosaccharides was possible.
After 1 month of continuous use, the column lost activity
and the resolution between fructose and glucose decreased.
The column’s condition remained at this point for several
months even after periods of prolonged storage. To com-
pensate for this change, the water composition of the mo-
bile phase may be lowered and/or the flowrate decreased to
maintain the desired separation.
For fructose, glucose, and sucrose, percent recovery of the
added 2 mg/g was 101, 98, and 93%, respectively, while
recoveries were 105, 101, and 83% for the added 8 mg/g.
The 83% recovery for sucrose at the 8 mg/g level may be
due to the insolubility of the disaccharide at that high a
concentration in methanol.
Analysis of three varieties of potatoes, Kennebec, Katah-
din, and Russet Burbank, stored at 3.3 and 7.2’C, demon-
strated that the method is reproducible (Table 1). Coeffi-
cients of variation ranged from 1.39-13.31% with all but
three coefficients less than 10%.
To determine if interfering peaks might be eluting at the
retention times of any of the three sugars, peaks from the
HPLC were collected and spotted on TLC plates. One spot
for each collected peak was observed on the TLC plate
after it was sprayed with sulfuric acid (50% v/v) and the Rf
value of each spot corresponded to the correct sugar, thus
indicating that the chromatographic peaks represented only
one sugar (Stahl, 1965).
Other applications of this method include determination
of sugars in the blanch water from a processing operation.
As the method is sensitive to the 0.0 1% level, it is useful for
sugar determination in such situations. Also the method can
be used for determination of sucrose content alone, an ad-
vantage for some growers of chipping potatoes (Sowokinos,
1978). Increasing the water content in the solvent system
to 35% decreases the analysis time to about 15 min but
doesnot provide separation between the monosaccharides.
With the modification in the procedure, the mean sucrose
content of Norchip and Monona chipping potatoes was
determined to be 2.95 and 3.25 mg/g, respectively.
The results of this study show that this method is a rapid
and quantitative analytical technique, which is potentially
useful in both basic research and in industrial operations.
REFERENCES
Della Monica, ES., Calhoun. M.J., and McDowell, P.E. 1974. The
quantitative determination of glucose, fructose, and sucrose in
fruits and p&a&es. J. Food Sci. 39: 1062.
Furuholmen. A.M.. Winefordner. J.D.. KnaDD. F.W.. and Dennison.
R.A. 1964: The quantitative &l&s of g&ose and fructose ii
ootatoes. Aeric. Food Chem. 12: 109.
H&t. D.C.%ckson. P.A.. ~Mortlock.~ R.E.. and Kirk. R.S. 1977.
Qu&titative determination of sugars in foodstuffs by high-per-
formance liouid chromatoaraohv. Analvst 102: 917.
Hurst, W.J. aid Martin, R.x.-Jr: 1977.-Rapid high pressure liquid
chromatographic determination of carbohydrates in milk choco-
late oroducts. JAOAC. 60: 1180.
Kim&a, M., Tohma, M.. Okazawa, Y., and Murai, N. 1969. Quanti-
tative determination of mono- and oligosaccharides in potato
tubers by gas-liquid chromatography. J. Chromatog. 41: 110.
Shaw, R. 1969. Analysis of potato sugars by gas chromatography.
Am. Potato J. 46: 201.
Sowokinos, J.R. 1978. Relationship of harvest sucrose content
to processing maturity and storage life of potatoes. Am. Potato
J. 55: 333.
Stahl, E. 1965. “Thin-Layer Chromatography: A Laboratory Hand-
book.” Academic Press. New York.
Timbie, D.J. and Keen&. P.G. 1977. Mono-dissaccharide analysis
of confectionaw products by high pressure liquid chromatography
especially relating to precolumns and other suggestions for con-
tending with contaminants. J. Food Sci. 42: 1590.
Ting, S.V. 1965. Rapid calorimetric methods for simultaneous
determination of total reducing sugars and fructose in citrus
juices. Agric. Food Chem. 4: 263.
Varns, J.L. and Shaw, R. 1973. An internal standard for rapid analy-
sis of DOtat sugars by gas chromatography. Potato Res. 16: 183.
Warthesen, J.J. and Kramer. P.L. 1973. Analysis of sugars in milk
and ice cream by high pressure liquid chromatography. J. Food
Sci. 44: 626.
Ms received 512180: revised 7/9/80; accepted 7126180.

Volume 46 (1981)-JOURNAL OF FOOD SCIENCE-301