-
Full Terms & Conditions of access and use can be found
athttp://www.tandfonline.com/action/journalInformation?journalCode=batc20
Critical Reviews in Analytical Chemistry
ISSN: 1040-8347 (Print) 1547-6510 (Online) Journal homepage:
http://www.tandfonline.com/loi/batc20
Disaccharides Determination: A Review ofAnalytical Methods
Marta Pokrzywnicka & Robert Koncki
To cite this article: Marta Pokrzywnicka & Robert Koncki
(2018) Disaccharides Determination:A Review of Analytical Methods,
Critical Reviews in Analytical Chemistry, 48:3, 186-213,
DOI:10.1080/10408347.2017.1391683
To link to this article:
https://doi.org/10.1080/10408347.2017.1391683
Accepted author version posted online: 20Oct 2017.Published
online: 11 Dec 2017.
Submit your article to this journal
Article views: 134
View related articles
Citing articles: 1 View citing articles
http://www.tandfonline.com/action/journalInformation?journalCode=batc20http://www.tandfonline.com/loi/batc20http://www.tandfonline.com/action/showCitFormats?doi=10.1080/10408347.2017.1391683https://doi.org/10.1080/10408347.2017.1391683http://www.tandfonline.com/action/authorSubmission?journalCode=batc20&show=instructionshttp://www.tandfonline.com/action/authorSubmission?journalCode=batc20&show=instructionshttp://www.tandfonline.com/doi/mlt/10.1080/10408347.2017.1391683http://www.tandfonline.com/doi/mlt/10.1080/10408347.2017.1391683http://www.tandfonline.com/doi/citedby/10.1080/10408347.2017.1391683#tabModulehttp://www.tandfonline.com/doi/citedby/10.1080/10408347.2017.1391683#tabModule
-
Disaccharides Determination: A Review of Analytical Methods
Marta Pokrzywnicka and Robert Koncki
Department of Chemistry, University of Warsaw, Warsaw,
Poland
ABSTRACTDisaccharides are determined mainly for dietetic
purposes, hence the most analyses are carried out forfood and drink
samples. Its content can also be used to profile groceries in order
to identify the origin andquality of the products. They also can be
an indicator of the rate of metabolism as well as for the control
ofsome technological and biotechnological processes. Unfortunately
most of technological analysis areperformed with nonselective
polarimetry methods. Sugars due to specific physicochemical
properties ofcompounds are difficult to determine with classical
analytical techniques. The most commondisaccharides are composed of
several types of monomers connected by a different configuration of
theglycosidic bond, therefore, there are subject of the same
characteristic reactions. This often enforces theneed for
pre-separation of sample components. Therefore, nowadays the most
popular analyticalmethodologies for disaccharides determination are
based on chromatographic and electrophoretictechniques. An
alternative is enzymes application that allow both selective
recognition of target analyteand its conversion to easy detected
product, allowing detection by relatively simple
conventionalanalytical methods. Another approach is the use of
advanced chemometric methodologies for computingof data obtained
from some spectroscopic techniques. This article is a review of the
recent analyticalliterature devoted to non-selective and selective
methods for disaccharide determination in real samples.
KEYWORDSChromatography;Disaccharides;Electrophoresis;
Enzymaticmethods; Spectroscopy
1. Introduction
From a huge group of sugars, disaccharides are the most popu-lar
analytes especially in food industry and agriculture. Amongthem the
most popular are sucrose, maltose and lactose(Fig. 1), which were
the main analytes reported in 85% of publi-cations cited in this
review. It is no surprise that the most oftenanalysed disaccharide
is sucrose (68%). In the 19th century thetable sugar became
strategic resource with special dotation forproduction and quality
control improvement. For many yearsit was recognized as the best
source of energy, nowadays it is adietician’s nightmare. WHO
considers it as one of the majoritysource of obesity and dental
caries and strongly recommendreduction of intake.[1] Therefore
sucrose is most often deter-mined in food products, especially soft
drinks[2–14] andsweets.[13–17] Because of its plant origin, sucrose
is also deter-mined in fruits and vegetables[17–26] (with
particular emphasison sugar cane[27] and sugar beet[28–31]) and
other plant mate-rial,[32–38] where it is an indicator of regular
plant growth.[36]
There are also some examples of sucrose assays in
urine[39–41]
and blood plasma[40–43] with special application to
blood-brainbarrier permeability investigations.[43]
Similar concern as target analyte share lactose (37%) andmaltose
(32%). Lactose, characteristic for mammals milk, isdetermined
exactly in milk[44–50] as well as in cheeses,[51]
yogurts[52] and different dairy products.[53] Considering
thatabout 50–75% of human population (data from different
sources[51,54]) suffer from lactose intolerance, not
surprisingnumber of publications reported new methods for lactose
assay.Finally, maltose occurs mostly in cereals grains. Except
cerealproduct[55,56] and starch hydrolates[57,58] it is often
determinedin alcoholic drinks[59] especially in beer[60–63] where
togetherwith glucose it is a marker of progress of fermentation
processas well as a marker of quality of final product. Of course
notonly these three disaccharides are targets of analytical
interest.There are also reported several methods for determination
oftrehalose,[34,64–68] lactulose,[69–73] isomaltose,[56,74]
cellobi-ose,[68,75–78] xylobiose,[75,79] mellibiose[78,80] and
more.
Selective methods of disaccharides determination involvemany
analytical challenges. Most of known disaccharides are iso-mers,
with the same molecular formula, molecular weight andalmost the
same functional group. Only small structural changessuch as the
inversion of groups at a single chiral carbon atom or achange in
the position of the carbonyl group, decide about differ-ent
sweetness, solubility, and chemical reactivity. This diversity
isclear even for monosaccharides, for example when simple
mono-saccharides like glucose, galactose and fructose are compared.
Incase of disaccharides it becomesmore complicated.
Disaccharidesare compounds with acetal bond between anomeric carbon
ofone monosaccharide molecule and any hydroxyl group of
secondmonosaccharide, so more isomer variations appear. In Figure
2the structures of ten different disaccharides are presented. Each
ofthem is built of two glucose units only and differ by
anomeric
CONTACT Marta Pokrzywnicka [email protected]
Department of Chemistry, University of Warsaw, Pasteura 1, Warsaw,
02-093, Poland.Color versions of one or more of the figures in the
article can be found online at www.tandfonline.com/batc.© 2018
Taylor & Francis Group, LLC
CRITICAL REVIEWS IN ANALYTICAL CHEMISTRY2018, VOL. 48, NO. 3,
186–213https://doi.org/10.1080/10408347.2017.1391683
https://crossmark.crossref.org/dialog/?doi=10.1080/10408347.2017.1391683&domain=pdf&date_stamp=2018-03-05mailto:[email protected]://www.tandfonline.com/batchttps://doi.org/10.1080/10408347.2017.1391683
-
form and order of bounded carbon atom in second molecule.Eight
of presented compounds are reducing sugars and theyundergo the same
characteristic assays based on Fehling’s,[81]
Smogy-Nelson[82] or Summer[83] methods. Other differences
aretaste (for example different sweetness rate or bitter in case of
gen-tiobiose[84]) and susceptibility to digestives enzymes. What
isobvious, because of different configuration of stereogenic
carbonseach of them have different specific rotation angle. These
proper-ties could be a base for selective determination method,
unfortu-nately effect of the light beam refraction is additive, and
thereforesuch approach is useless in case of quantitative analysis
of sugarmixtures. Nerveless, polarimetry together with other
non-selec-tive techniques such as refractometry and hydrometry are
cur-rently used in industrial analysis.[85]
2. Non-selective analytical methods
Similarity in disaccharides structures and properties causesmost
of analytical methods non-selective. Therefore in manycases
information about disaccharide content, especially in
food and beverages, is limited to total sugar content –
theparameter which presents the total concentration
disaccharidesand related monosaccharides in sample. This parameter
pro-vides satisfactory information from nutritional point of
view.In case of these assays there is no necessary to apply any
selec-tive techniques. For such purposes, several methods based
onnon-selective chemical reactions or physical properties of
sugarsolutions have been developed.
2.1. Chemical methods
For estimation of total sugar content often phenol-sulphuricacid
assay[86] is applied. This method is useful for determina-tion of
reducing and non-reducing sugars in complex samples,also in the
presence of salts and proteins residues. The assay isbased on
measure of colour of aromatic complex absorbance at490 nm. The
method was developed in the middle of XX cen-tury but is still
applied because of its simplicity and availabilityof reagents. Also
3,4-dimethylphenol forming colour adductsexhibiting absorption
maximum at 510 nm wavelength can be
Figure 1. Structures of three common disaccharides: sucrose,
maltose and lactose.
Figure 2. Examples of disaccharides composed of two glucose
units.
CRITICAL REVIEWS IN ANALYTICAL CHEMISTRY 187
-
applied for total sugar determination.[87] However because
ofdifferent absorptivity of individual sugars, this method can
besuccessfully applied only under specific conditions.
Inmany cases almost all sugars presented in sample have
reduc-ing properties caused mainly by monosaccharides and
reducingdisaccharides. There is a lot of methods for reducing
sugars deter-mination and some of them are presented in Table 1.
Predomi-nantly these are modifications of classic Fehling’s
methoddeveloped in the middle of XIX century[81] based on the
reductionof cupric ion to cuprous oxide. The main differences are
in thetitrant and titrand. That could be dissimilarities in
reaction condi-tions like in Loof–Schorl[88] and Ofner’s[89]
methods or exchangethe role of sample and titrant as in case of
Lane and Eynonsmethod,[88] where Fehlings solution with known
cupric concentra-tion are titrated with sugar solution. Copper
based reaction can bealso applied for spectrophotometric detection.
Cuprous ions canreact with neocuproine to produce a yellow complex
that stronglyabsorb the light of 457 nm wavelength. It is obvious
that the deter-mination methods based on reducing properties of
sugars arestrongly influenced by other reductans (almost the same
methodsare applied for estimation of total antioxidant capacity
assays)
Methods based on ferricyanide reduction also have somevariation.
Except iodometric Mohr method[92] or simple photo-metric
measurements of decay of ferricyanide absorption at480 nm
wavelength[92,93] also formation of Prussian Blue, after
addition of ferric nitrate can be applied allowing observation
ofabsorption increase at 690 nm wavelength.[35] Under flow
anal-ysis condition, thanks to following measurements of
hexacya-noferrate obtained after reduction by sugars from
non-hydrolysed and hydrolysed sample, quite selective
determina-tion of sucrose was possible in beet juices and
syrups.[31]
Also benzoic acid derivatives containing nitrogen as
3,5-dinitro-salycilic acid (DNS)[83,91] and p-hydroxybenzoic acid
hydrazide(PAHBAH)[94–96] can be reduced by sugars and form
colouredproducts that absorb in visible range at 540 and 410 nm
wave-length, respectively. An interesting case ismethylamine. Its
reactionwith reducing sugars leading to formation product, which
colourdepends on sugar structure.With monosaccharides it forms
yellowproduct that absorb at 400 nm, whereas with oligosaccharides
(dis-accharides and trisaccharides) violet- carmine adduct is
formedthat absorbs light of 540 nm wavelength.[97] This way the
discrimi-nation and determination of chosen saccharide groups is
possibleas well as estimation of total reducing disaccharide
content only ifoligosaccharides composed of higher number of
monosaccharidesunits are not present.
Some reactions of reducing sugars allow to distinguish aldo-ses
and ketoses. For example Seliwanoff’s test is
characteristicreaction of ketohexoses, which in acidic solutions
are convertedinto hydroxymethylfurfural forms that react with
resorcinol togive red complexes with strong, characteristic
absorption peaksat 398 and 480 nm wavelength. This qualitative test
has beensuccessfully applied for quantitative photometric
determinationof lactulose in pharmaceutical samples.[98]
Hydroxymethylfur-fural can react also with cysteine
hydrochloride–tryptophanreagent producing pink chromophore with
maximum absorp-tion at 518 nm. This reaction also has been applied
for determi-nation of lactulose in pharmaceutical products.[99]
So-called Raybin test based on reaction of sugar with
5-diaz-ouracil gives positive results (bluish precipitate) for
compoundscontaining 1,2-glycosidic bond such as sucrose, raffinose
or sta-chyose, while palantinose and melezitose also containing
thisbond type yielded a reddish-brown precipitate. This test
hasbeen adapted for photometric determination of sucrosetogether
with rafinose in honeys samples.[100]
Reducing sugars in alkaline solution tautomerize to form
eno-diols. These compounds can react with zirconyl chloride to
formfluorescence derivatives. Tautomerisation to enodiols is
muchfaster for ketoses, so under adequate conditions (lower
reactiontemperature and shorter time) selective fructose
determination canbe achieved. Similar fluorescent complexes with
zirconyle chlorideare formed by fructose-based disaccharides, so
this method is alsouseful for fluorometric determination of
sucrose.[7]
Finally, in case of reducing sugars determination it is worthto
mention about qualitative osazone test.[101] At high tempera-tures
reducing sugars react with phenyl hydrazyne to form yel-low
crystals called osazones. For each reducing sugar crystalswith
different structure, precipitation time and melting pointare
formed. Comparison of those parameters is useful for sac-charides
differentiation.
2.2. Physical methods
Physical techniques developed for investigations of liquidslike
polarimetry, hydrometry and refractometry also belong to
Table 1. Methods for reducing sugars determination. Table 1.
Examples of non-selective chemical methods for determination of
reducing sugars.
Method Reaction Detection Source
Lane and Eynon’smethod*
sugars reduce cupric ionsto cuprous oxide
end point titrationindicated bymethylene blue
[88]
Ofner’s method* sugars reduce cupric ionsto cuprous oxideexcess
of cupric ionsare reduced bypotassium iodide
iodine titrated by sodiumthiosulphate withstarch as
indicator
[89]
Loof -Schorlmethod*
sugars reduce cupric ionsto cuprous oxideexcess of cupric
ionsare reduced bypotassium iodide
iodine titrated by sodiumthiosulphate withstarch as
indicator
[88]
Knight and Allenmethod*
sugars reduce cupric ionsto cuprous oxide
residual cupric ions aretitrated with EDTAwith murexide
asindicator
[88]
Munson andWalkermethod
sugars reduce cupric ionsto cuprous oxide
cuprous oxide are driedand weight
[89]
Smogyi- Nelsonmethod
sugars reduce cupric ionsto cuprous oxidecuprous oxide
reducedmolybdic acid tomolybdenum blue
photometricdetermination ofmolybdenum blue at500 nm.
[82,86]
Sumner’s method sugars reduce 3,5-dinitrosalycilic acid
to3-amino-5-nitrosalicylic acid
photometricdetermination of 3-amino-5-nitrosalicylicacid at 540
nm.
[83,91]
Hagedorn-Jensenmethod
sugars reduceferricyanide toferrocyanide
iodometricallydetermination offerricyanide by theMohr method
[92]
photometricdetermination offerricyanide at480 nm.
�official methods recommended by ICUMSA.
188 M. POKRZYWNICKA AND R. KONCKI
-
non-selective methods of disaccharides determination. How-ever,
there are not applied to total sugar determination
becausecontributions of each solute (sugar) in total liquid
density,refraction index and light beam polarization are individual
andnot simply additive as in case of chemical stoichiometric
reac-tions reported above. Those techniques are widely applied
insamples where influence for refractive index is expected onlyfrom
single disaccharide.[9,15] However, due to measurementsimplicity
and non-expensive equipment, from over 100 years,these methods are
still widely used for routine analysis in sugarindustry. There are
also so called Saccharimeters the speciallydesigned polarimeters
and refractometers with scale recalcu-lated from angle of rotation
or refractive index to sucrose con-centration. Analysis is very
simple because proper values aretabulated.
Obviously, density, rotation angle and refractive index dependon
temperature. Additionally, polarization and refractive indexdepend
also on wavelength and therefore the measurements haveto be
performed under precisely specified conditions: temperatureof 20�C
(293K), and wavelength of the D line of sodium(589.3 nm) as
standard and symbolized by nD. It should be empha-sized that
accuracy of such measurements is based on assumptionof substance
purity. Although the result of analysis is true only ifsingle sugar
is present in sample, for fast routine assays suchmethod is
sufficient. These techniques could be applied to analysisof complex
samples after sugars separation. However, in the analyt-ical
practice only refractometry has found wider application asdetection
technique in liquid chromatography. Recently[15], thetechniques
based on refractive index difference were applied fordetermination
of sucrose content. In case of candy floss analysisretro-reflected
beam interference based on refractive index detec-tion has been
applied For cola drinks analysis instead of conven-tional
refractometer measurements were carried in photometricflow cell.
They were based on detection of light deflected by Schlie-ren
effect formed due to refractive index gradient.[9] The result
ofmeasurements were presented using Brix degrees.
Brix (�Bx) and related scales (Balling �Bg, Plato �P) arewidely
applied for sugar liquids characterization. One Brix,Balling and
Plato degree is percentage by weight of sucrose inpure water
solution. Difference between these three indices isin reference
temperature and closeness of measurements values(3,5 and 6 decimal
places for �Bx, �Bg, and �P, respectively).Nowadays the most
popular and the most used is the Brixindex. Plato index is
sometimes applied in brewing industry.Generally these units are
closely connected with concentrationof sucrose in pure water but
sometimes the industry uses theseunits somewhat loosely to refer to
any sweet solids in a prod-uct.[102] In case of sugars other than
sucrose Brix is called the“apparent Brix” and is always a relative
value. The Brix indexcan be determinated both by hydrometry and
refractometry.Specially designed devices have scale both in
[kg/m3]/ [g/cm3]or nD and Brix. However, in some cases values
obtained byhydrometer and refractometers can differ each other,
especiallyin case of “apparent Brix” measurements and samples
withcomplex matrix. For example analysis of orange juice
samplesrequired special correction because of sample
acidity.[103]
An interesting issue is industrial analysis of sucrose.
Nowadaysall commonly accepted methods for table sugar assay are
codifiedby ICUMSA (International Commission for Uniform Methods
of
Sugar Analysis). ICUMSA Methods Book.[104] contains
descriptionof official methods recommended for quality control in
sugarindustry, both for sucrose and impurities (lead, arsenic,
copper,iron, etc.) concentrations determination as well as
tabulated valuesof specific rotation, refractive index and density
of sucrose solu-tions. This book contains also regulations
concerning some selec-tive determinations based on chromatographic
and enzymaticmethods, reported in the next paragraphs of this
review.
3. Selective analytical methods
Taking into account rather high content of sugars in real
sam-ples and practically unlimited accessibility of these
samples(mainly food and agriculture products), very low detection
lim-its and wide determination ranges are not extremely
importantparameters of modern methods developed for analysis
ofsugar-containing products. A crucial analytical factor of
thesemethods is a selectivity allowing detection of particular
saccha-ride in the presence of complex matrix of sample
additionallycontaining various other very similar sugars. Currently
thereare three main trends for the development of methods for
selec-tive determination of disaccharides. The most popular
andeffective are various separation techniques like
chromatographyand electrophoresis. The second direction is the
development ofbioselective methods based on almost specific
recognition ofanalyte by enzymes and its conversion into easily
detectedproduct. A third relatively new direction, also dedicated
forsample analysis without analyte separation, is the application
ofadvanced spectroscopic methods (IR, Raman) combined withuse of
sophisticated chemometric tools.
3.1. Separation-based methods
Nowadays the most dynamically developed and widely reported
inthe analytical literature separative technique for
disaccharidesdetermination is liquid chromatography. Some recent
papersdescribing application of various chromatographic methods
fordisaccharide determination are cited in Table 2. High
PerformanceLiquid Chromatography (HPLC) is the official technique
for rou-tine sugars analysis recommended by AOAC International
(Associ-ation of Official Analytical Chemists).[86] HPLC gives
bothqualitative (identification of the carbohydrate) and, with peak
inte-gration, quantitative information. The analysis is rapid,
applicableto samples with a wide range of sugar concentrations,
precise andaccurate and do not required derivatization of
carbohydrates.
Liquid chromatography for disaccharide separations exploit
dif-ferences in polarity (HPLC normal and reversed- phase,
HILIC,HTLC) or electrical charge (IEC: cation, anion exchange and
ionexclusion) of target molecules. The most popular
chromatographicmode is HLPC with normal-phase configuration. In
this modecommonly applied stationary phase is silica gel with amino
groups,whereas acetonitrile–water (40–95% acetonitrile) is used as
amobile phase. The elution order is monosaccharides,
disaccharidesand finally higher oligosaccharides. The gradient
elution allows toavoid interferences caused by sugar alcohols
(mannitol can coelutewith maltose and lactose while inositol with
sucrose). In reversedphase HPLC the hydrophobic stationary phase is
silica gel withadded alkyl chains, for example C18 column.[42]
Thismode of sepa-ration has been used for separation of mono- di-
and trisaccharides
CRITICAL REVIEWS IN ANALYTICAL CHEMISTRY 189
-
Table2.
ApplicationofLiqu
idchromatograph
yford
isaccharides
determ
ination.
Chromatograph
icseparatio
n
Detectio
ncolumn
elution
Determined
disaccharid
esOtherdeterm
ined
substance
Sample
Reference
Normal-Phase
HPLC
ELSD
Zorbax
RX-SIL(m
odified
byethylene
diam
ine)
isocratic
acetonitrile-water
(72%
:28%
)with
0.03%
ethylene
diam
ineand0.05%
ammoniumhydroxide
sucrose,maltose,lactose
glucose,fructose,raffinose
Drin
ks(app
lejuicepineapple
juiceorange
juicegrapewine
liquor)
[10]
ELSD
carbohydratecolumn
Isocratic
acetonitrile–w
ater
(70%
:30%
)sucrose,maltose,lactose
glucose,fructose,galactose
raffinose
fruits,vegetables,grains,seeds
andleaves
[32]
ELSD
Phenom
enex
Luna
5uNH2100A
isocratic
acetonitrile-water
(82.5%
:17.5%
)sucrose
glucose,fructose,sorbitol
fruits(peach,app
lewatermelon,
cherry)
[23]
ELSD
SpherisorbNH2
gradient
from
81%acetonitrile
/19%
waterto
75%
acetonitrile
/25%
waterover
40min
maltose
glucose,fructose
beer
[62]
ELSD
PrevailCarbohydrateES
isocratic
acetonitrile-water
(80%
:20%
)lactose,lactulose
glucose,fructose,galactose
synthetic
samples
[105]
YMCPack
Polyam
ine
Zorbax
Carbohydrate
Analysis
UnisonUK-Am
inoHT
ELSD
andC-CA
DNH2-Krom
asil
isocratic
acetonitrile–w
ater
(70%
:30%
)sucrose,maltose,lactose
glucose,fructose
maltotriose
sauces,syrup
s,jellies,glazes,
hone
anddairy
products
[13]
CAD
ShodexAsahipak
NH2P-50E4
gradient
from
90%acetonitrile
/10%
waterto
77%
acetonitrile
/23%
waterover
22minandkept
constant
until40
min
sucrose,maltose
glucose,fructose,erythritol,
xylitol,sorbitol,mannitol,
maltitol
drinks
(juices,nectarsand
syrups)
[106]
RID
Tracer
carbohydratescolumn
isocratic
acetonitrile–w
ater
(75%
:25%
)sucrose,lactose,lactulose
glucose,fructose,galactose
milk-based
form
ulae
[69]
RID
SupelcosilLC-NH2
isocratic
acetonitrile–w
ater
(80%
:20%
)trehalose
—Selaginella
lepidophyllaplant
[67]
RID
Zorbax
Carbohydate
aminopropyl
isocratic
acetonitrile–w
ater
(82%
:18%
)sucrose,maltose,lactose
glucose,fructose,m
annose,
sorbito
l,xylitol
wine,juices,honey,dairy
products,biscuits
[14]
RID
UltraAminoColumn
isocratic
acetonitrile–w
ater
(75%
:25%
)lactose,lactulose
—HeatTreated
Milk
[44]
PinacleIIAm
ino
isocratic
acetonitrile–w
ater
(75%
:25%
)2PinacleIIAm
ino(in
series)
isocratic
acetonitrile–w
ater
(75%
:25%
)RID
Zorbax
Carbohydate
aminopropyl
isocratic
acetonitrile–w
ater
(82%
:18%
)sucrose,maltose,lactose
glucose,fructose,m
annose,
sorbito
l,xylitol
wine,juices,honey,dairy
products,biscuits
[14]
RID
UltraAminoColumn
isocratic
acetonitrile–w
ater
(75%
:25%
)lactose,lactulose
—heattreatedmilk
[44]
PinacleIIAm
ino
isocratic
acetonitrile–w
ater
(75%
:25%
)2PinacleIIAm
ino(in
series)
isocratic
acetonitrile–w
ater
(75%
:25%
)RID
PrevailCarbohydrateES
isocratic
acetonitrile–w
ater
(75%
:25%
)lactose,lactulose
—conservedmilk
[45]
RID
Agilent
Zorbax
Carbohydrate
isocratic
acetonitrile–w
ater
(80%
:20%
)sucrose
glucose,fructose,sorbitol,
organicacids
grapefruitpu
lps
[24]
RID
SpherisorbAm
ino
isocratic
acetonitrile–w
ater
(70%
:30%
)lactulose
–lactose-free
milk
[107]
190 M. POKRZYWNICKA AND R. KONCKI
-
ESIM
S/MS
PrevailCarbohydrateES
isocratic
acetonitrile–
ammoniumform
ate
(70%
:30%
)
lactose,lactulose
ESI/M
SAcqu
ityBEHam
ide
gradient
from
95%ofacetonitrile/5
%0.1%
ammoniainwaterisocratic
for
2min70%ofacetonitrile/3
0%0.1%
ammoniainwaterfor2
minand60%of
acetonitrile/4
0%0.1%
ammoniain
waterfor2
min
sucrose
glucose,fructose,kestose,
nystose
datesfruits
[22] MS/MS
Acqu
ityBEHam
ide
isocratic
acetonitrile–0.1M
ammoniumhydroxidein
water(80%
:20%
)
sucrose
—ratp
lasm
a,blood,andbrain
homogenate
[43]
MS/MS
Acqu
ityBEHam
ide
gradient
from
75%of
0.1%
ammoniumhydroxidein
acetonitrile/2
5%0.1%
ammoniumhydroxidein
waterto
67.5%of0.1%
ammoniumhydroxidein
acetonitrile/3
2.5%
0.1%
ammoniumhydroxidein
waterfor1
0min,to62.5%of
0.1%
ammoniumhydroxide
inacetonitrile/37.5%0.1%
ammoniumhydroxidein
waterfor2
minand75%of
0.1%
ammoniumhydroxide
inacetonitrile/25%0.1%
ammoniumhydroxidein
waterfor0
.1minthen
isocratic
for6
min
isom
altose
pannose,isom
altotriose
milk
powder
[74]
MS/MS
ZIC-
HILIC
gradient
from
75%acetonitrile
/25%
5mMofam
monium
acetateinwater
to40%
acetonitrile
/60%
5mMof
ammoniumacetateinwater
in10
min
sucrose,lactulose
raffinose,m
annitol
urine
[39]
AscentisSi
gradient
from
80%acetonitrile
/20%
5mMofam
monium
acetateinwater
to65%
acetonitrile
/35%
5mMof
ammoniumacetatein
waterin6min
SupelcocilLC-NH2
gradient
from
75%acetonitrile
0.05%form
icacid/25%
H2O
0.05%form
icacidto
40%
acetonitrile
0.05%form
icacid/60%
H2O
0.05%
form
icacidin6min
(Continuedon
nextpage)
CRITICAL REVIEWS IN ANALYTICAL CHEMISTRY 191
-
Table2.
(Continued)
Chromatograph
icseparatio
n
Detectio
ncolumn
elution
Determined
disaccharid
esOtherdeterm
ined
substance
Sample
Reference
chem
iluminescence
Krom
asilNH2
isocratic
acetonitrile–w
ater
(70%
:30%
)lactose
glucose,fructose,xylose,
arabinose
grapefruitsextracts
[108]
Reversed
-Phase
HPLC
DAD
andRID
Aminex
HPX-87H
isocratic
3mMsulfuric
acid
lactose
glucose,fructose,organicacids
goat’smilk
yogu
rts
[52]
MS
Alltech
C18
gradient
from
90%0.1%
acetic
acidinwater/1
0%acetonitrile
with
0.1%
acetic
acidto
100%
0.1%
aceticacid
inwater
over8min,isocratic
for3
min,to0.1%
aceticacid
inwater/10%acetonitrile
with
0.1%
aceticover3min
andisocraticallyat90%0.1%
aceticacidinwater/10%
acetonitrile
with
0.1%
acetic
for3
min,for
atotalrun
time
of17
min
sucrose
—equine
serum
[42]
Hydroph
ilicInteractionCh
romatog
raph
y[HILIC]
QTO
FMS
XamideColumn
gradient
5%,10%
or20%of
100mMam
moniumform
ate
(pH3.2)in:0%acetonitrile/
100%
water
to80%
acetonitrile/2
0%waterfor
60min
lowmolecularweigh
theparindisaccharid
eheparin
—[109]
ESI/TOFM
S/MS
Zorbax
NH2am
inopropylsilica
gradient
from
88%8mM
ammoniumform
atein
acetonitrile/1
2%8mM
ammoniumform
ateinwater
isocratic
for1
0minto
80%
8mMam
moniumform
atein
acetonitrile/2
0%8mM
ammoniumform
ateinwater
for8
min,to75%8mM
ammoniumform
atein
acetonitrile/2
5%8mM
ammoniumform
ateinwater
for4
min,to70%8mM
ammoniumform
atein
acetonitrile/3
0%8mM
ammoniumform
ateinwater
for5
min,and
isocratic
for
2min
sucrose,mellibiose
glucose,fructose,galactose,
raffinose,m
anninotriose,
stachyose,verbascose
crud
eandprocessedRadix
Rehm
anniae
[80]
ELSD
Acclaim
Trinity
P2isocratic
acetonitrile–100
mM
ammoniumform
iatebu
ffer
pH3.65
(80%
:20)
lactulose
—milk
[72]
192 M. POKRZYWNICKA AND R. KONCKI
-
HighTemperature
Liqu
idCh
romatog
raph
y[HTLC]
ELSD
Hypercarb
isocratic
water
sucrose,maltose,lactose
glucose,fructose,galactose
milk,orang
eandmandarin
efruits
[110]
Ion-Exchange
Chromatog
raph
y[HPA
EC](anionexchange)
PAD
CarboPac
PA1
gradient
from
16mMsodium
hydroxideto
250mMsodium
hydroxidefor6
0min
maltose,lactose,trehalose,cellobiose
glucose,galactose,rafinose,
ribose,rham
nose,arabinose,
ethanolsorbitol,glycerol,
arabito
l,erythrito
l
yeastculturesandferm
entatio
nbroths
[76]
CarboPac
MA1
isocratic
480mMsodium
hydroxide
PAD
CarboPac
PA10
isocratic
5mMsodium
hydroxide
sucrose
glucose,fructose
wastewaterfrom
thebeverage
indu
stry
[66]
PAD
CarboPac
PA10
isocratic
50mMpotassium
hydroxide
sucrose,lactose
glucose,fructose
chocolate
[16]
PAD
CarboPac
PA10
gradient
87.5mMsodium
hydroxideisocratic
for
10min,to500mMsodium
hydroxidefor0
.1min,
isocratic
for7
min
maltose,isomaltose
glucose,rib
ose
bloodserum
[111]
PAD
CarboPac
PA20
isocratic
8mMsodium
hydroxide
lactose
glucose,galactose
naturally
“lactosefree”hard
cheese
[51]
PAD
CarboPac
PA20
isocratic
water
sucrose
glucose,fructose,galactose,
arabinose,mannose,
rham
nose,m
annitol
greencoffeebean
[33]
PAD
AminoPac
PA10
gradient
from
17.5mMsodium
hydroxideto
25mMsodium
hydroxidefor2
min,to
37.5mMsodium
hydroxide
for3
min,to90
mMsodium
hydroxidefor6
min,2
min
isocratic,to100mMsodium
hydroxidefor2
min,to
175mMsodium
hydroxide
for5
min,to175mMsodium
hydroxideand3mMsodium
acetatefor0
.1min,to
175mMsodium
hydroxide
and7mMsodium
acetatefor
8min,to175mMsodium
hydroxideand10
mM
sodium
acetatefor5
min,to
175mMsodium
hydroxide
and20
mMsodium
acetate
for7
min,to75
mMsodium
hydroxideand60
mM
sodium
acetatefor5
min,
10minisocratic
maltose,isomaltose
glucose,fructose,arabinose,
maltotriose,isomaltotriose,
panose,m
altotertose,
aminoacids
ricewines
[59]
PAD
CarboPac
PA20
isocratic
1mMsodium
hydroxide
sucrose
glucose,fructose,arabinose,
mannose,xylose
aqueousextractsand
hydrolysates
ofbiom
ass
[112]
(Continuedon
nextpage)
CRITICAL REVIEWS IN ANALYTICAL CHEMISTRY 193
-
Table2.
(Continued)
Chromatograph
icseparatio
n
Detectio
ncolumn
elution
Determined
disaccharid
esOtherdeterm
ined
substance
Sample
Reference
PAD
CarboPac
PA200
gradient
0.1M
sodium
hydroxide
isocratic
for9
min,to0.1M
sodium
hydroxideand0.04M
sodium
acetatefor1
7min,to
0.1M
sodium
hydroxideand
0.25Msodium
acetatefor
0.1min,isocraticfor1
4min
cellobiose,xylobiose
linearxylo-oligosaccharides
and
cello-o
ligosaccharides
lignocellulosicprocessing
products
[75]
PAD
Ham
ilton
RCX-30
gradient
from
50mMsodium
hydroxide25
mMsodium
acetateto
50mMsodium
hydroxide100mMsodium
acetatefor1
5min,isocratic
for1
0min
maltose,isomaltose
glucose,malto-oligosaccharides
wheatflour
[55]
PAD
CarboPac
PA10
gradient
2mMsodium
hydroxideisocratic
for5
min,
to5mMsodium
hydroxide
for3
min,isocraticfor2
min,
to20
mMsodium
hydroxide
for2
min,isocraticfor3
min,
to30
mMsodium
hydroxide
for1
min,isocraticfor2
min,
to40
mMsodium
hydroxide
for3
min,isocraticfor1
min,
to45
mMsodium
hydroxide
for3
min,isocraticfor2
min
sucrose,lactose,trehalose
glucose,fructose,arabinose,
ribose,xylose
rawsugar
[65]
ESI/M
SCarboPac
PA20
gradient
3mMpotassium
hydroxideisocratic
for
30min,to80
mMpotassium
hydroxidefor5
min
sucrose,lactose,trehalose
glucose,fructose,m
annitol,
glucosylglycerol
intracellularextractsof
cyanobacteria
[113]
ELSD
CarboPac
PA1
isocratic
32mMpotassium
hydroxide
sucrose
glucose,fructose,arabinose,
xylose
drinks
(cola,orange
juice,
watermelon
juice)
[12]
Ion-Exchange
Chromatog
raph
y[HPC
EC](catio
nexchange)
ESI/M
SIOA-1000
9mm
isocratic
20mMform
icacidand
10mMtrichloroacetic
acid
sucrose,lactulose,sucralose
rham
nose,erythritol
urineandbloodplasma
[40]
RID
Carbosep
Corgel87H3
isocratic
5mMsulfuric
acid
sucrose
glucose,fructose,sorbitol
apple’sleafandfruitp
eel
[25]
RID
RNMCarbohydrateNaC
isocratic
water
xylobiose
xylo-oligosaccharides
enzymaticallyhydrolysed
pulp
[79]
RID
SugarP
akI(Ca
2C)
isocratic
water
lactose,lactulose
—heattreatedmilk
[44]
RID
RezexRSO-OligosaccharideIE
(AgC
mode)
isocratic
water
maltose
glucose,fructose
beer
[61]
IEXC
a2C
isocratic
water
Ion-ExclusionCh
romatog
raph
y
RID
Bio-RadAm
inex
HPX
87H
Isocratic
0.005n
phosph
oricacid
sucrose
glucose,fructose,organicacids
fruitjuices
[114]
Abbreviatio
ns:CAD
-Charged
AerosolD
etectio
n;C-CA
D-C
orona-Ch
argedAe
rosolD
etector;DAD
-Diode
ArrayDetector;ESI-Electrospray
Ionizatio
n;ELSD
-EvaporativeLigh
tScatteringDetectio
n;MS-MassSpectrom
etry;M
S/MS-Tan-
demMassSpectrom
etry;PAD
-PulsedAm
perometric
Detectio
n;QTO
FMS-Quadrup
olTime-Of-Flight
MassSpectrom
etry;RID-R
efractiveIndexDetector;TO
FMS-Time-Of-Flight
MassSpectrom
etry.
194 M. POKRZYWNICKA AND R. KONCKI
-
by groups. A main disadvantage of this technique concerns
mono-saccharides separation (short retention time results in
elution as asingle unresolved peak). There could be also problem in
the pres-ence of anomers that can results in peak doubling
and/orbroadening.
For HPLC systems a variety of detectors can be
coupled:Refractive Index Detector (RID),[14,24,44,45,67,69]
EvaporativeLight Scattering Detectors (ELSD),[10,23,32,62] various
MassSpectrometry techniques (tandem MS,[39,43,74] ESI
MS[22,115])Charged Aerosol Detectors (CAD)[13,106] or even
chemilumi-nescence.[108] A simple and economic RID seems to be the
bestsolution for determination of separated sugars although it
isless sensitive then other types of detectors. Unfortunately dueto
its strongly dependence on solvent, type RID cannot beapplied in
case of gradient elution. On the other hand, ELSDand MS required
solvent evaporation.
An alternative HPLC mode for separating polar compounds
ishydrophilic interaction liquid chromatography (HILIC). In
thiscase a polar-hydrophilic stationary phase, characteristic for
normalphase mode, is coupled with also polar water-containing,
mobilephase characteristic for reversed phase.[116] But unlike in
reversed-phase chromatography, gradient elution HILIC begins with a
low-polarity organic solvent and elutes polar analytes by
increasing thepolar aqueous content.[42,80] Such inversion of
elution order allowsshortening of analysis when the target analyte
is a single particularsaccharide. Another variation of HPLC is high
temperature liquidchromatography (HTLC).[110] In HTLC thanks to
raising separa-tion temperature to about 100�C there is a
possibility to apply purewater as eluent without elongate time of
analysis and degradationin resolution.
Carbohydrate separation can be achieved also by High
Per-formance Thin Layer Chromatography (HPTLC).[117] Silica gel60
F254 plates with dropped of 1 mL of sample were developedat room
temperature with a mobile phase of acetonitrile: water(8.5:1.5,
v/v). Before determination sample was derivatives byaniline
diphenylamine o-phosphoric acid. This method wasapplied for
determination of maltose and total sugars in insweet potato
(Ipomoea batatas L.).
Disaccharides separation based on their electrical charge is
alsopossible, because carbohydrates are very weak acids (pKa
values>12).[118] In strongly alkaline solutions some
carbohydrate hydroxylgroups are ionized allowing sugars separation
on anion-exchangecolumns. The employed mobile phases are simple and
inexpensivesodium hydroxide[33,51,65,66,76,111,112] or potassium
hydrox-ide[16,113,114] solutions, with or without addition of
acetatesalt.[55,59,75] Also water may be used,[33] but then
postcolumn addi-tion of a electrolyte solution is required for
obtain adequate detec-tion conditions. This separation mode is very
often connected withPulsed Amperometric Detection (PAD).
Amperometric detectionhave a lot of advantages. Under specific pH
and voltage conditionsonly carbohydrates will undergo the redox
reaction. Coupling thismethod with chromatographic separation
additionally increasesselectivity of detection. The development of
so-called triple-pulsedamperometric detector,[118] solve the
problem of electrode poison-ing by accumulation of oxidized
products on its surface andallowed electrochemical detection for
carbohydrates. The entirecleaning process takes milliseconds and is
ongoing throughout therun. Because electrode reaction relies on
oxidation of carbohydratehydroxyl and aldehyde groups, this
detector is suitable for both
reducing and nonreducing carbohydrates. Also example of
applica-tion of ELSD detector coupled with cation exchange
separation isdescribed.[114] This type of detector requires
evaporation of eluentbefore detection step and therefore the
non-volatile potassium saltsin the basic eluent has to be removed
by a suppressor.
In cation exchange liquid chromatography stationary phasesare
often resin loaded with one of a variety of metal counterions
Ca2C,[44,61] NaC[79] or AgC[61], which react selectively withthe
weakly negatively charged hydroxyl groups of sugar mole-cules. The
selectivity of this process is controlled with theappropriate
choice of resin type and of the metallic speciesbonded to it, as
well as by the temperature of column (columnsnormally are operated
at elevated temperatures to increase itsefficiency). The mobile
phase is typically water. The mechanismof separation is based on
the strength of the bonding betweencis-glycols of sugar molecules
with the cation loaded on the col-umn. The elution order is related
to the number and strengthof cis-glycol complexes formed and takes
place in the order ofdecreasing molecular weight.[86]
Ion exclusion chromatography (IEC) also found applicationin
disaccharide determination.[114] In this techniques at ionexchange
resin the ionic substances are rejected while non-ionic or
partially ionized substances are retained and separatedby partition
between the liquid inside the resin particles andthe liquid outside
the particles. In effect the ionic substancespass quickly through
the column. Non-ionic or partially ion-ized substances are held up
and eluted more slowly.
Modern liquid chromatography techniques require
specializedlaboratory equipment and are often connectedwith high
consump-tion of eluent – mostly expensive chemically ultra-pure
solvents.Moreover, the most often applicable normal-phase HPLC
meth-ods, where organic solvents are applied, stay in contradiction
togreen chemistry assumptions. From this point of view Gas
Chro-matography (GC) seems to be better solution for
disaccharidesdetermination. Without consumption of expensive
ultra-purereagents it allows to determine much more analytes in the
courseof single run.[119] In the Table 3 some papers from last 20
yearsdevoted to GC determination of disaccharides are collected.
Inalmost all cases disaccharides have been determined together
withmonosaccharides and sugar alcohols.
Sugars are non-volatile and thermally unstable compounds.Their
determination with GC have to be preceded by the deriv-atization
process, often complicated and time consuming. Thisderivatization
required before separation step is the main prac-tical drawback
strongly limiting the development of GC fordisaccharide analytics.
For GC sugars determination methyl,acetate, trifluoroacetate and
trimethylsilyl derivatives can beapplied.[132] Nowadays the most
popular are trimethylsilyls(TMS) and trimethylsilyl oximes (TMSO).
Trimethylsilylationof carbohydrates is a simple reaction and
because non-volatilereagents or by-products are involved, the
complete reactionmixture can be injected directly into the gas
chromatograph,avoiding clean-up stages. Trimethylsilyl derivatives
of carbohy-drates acquire different chemical properties, molecule
increasevolatility and thermal stability and decrease
polarity.[133] It isworth to mention that TMSO derivatised
disaccharides are iso-meric molecules with monosaccharide ring
structure and anopen chain with the oxime group. The only
difference is in theposition of the substituents so they can have
similar mass
CRITICAL REVIEWS IN ANALYTICAL CHEMISTRY 195
-
Table3.
ApplicationofGas
Chromatograph
yford
isaccharides
determ
ination.
Chromatograph
icseparatio
n
Detectio
ncolumn
temperature
Determined
disaccharid
esOtherdeterm
ined
substance
Sample
Reference
FID
TR-1capillarycolumn(60m
£0.32
mmi.d.;0.25
mmfilm
of100%
methylpolysiloxane)
80� C
for2
minto
240�Cat5
� C/m
inleftfor5
minto
280�Cat20
� C/m
inleftfor1
5min
sucrose,maltose,lactose,m
elibiose
glucose,fructose,xylose,
mannose,galactose,sorbitol,
myo-inositol
tobacco
[36]
FID
ZB-5(30m
£0.25
mmi.d.;0.25mm
film
of5%
phenylmethylpolysiloxane)
100�Cto
180�Cat4
� C/m
inleftfor
2minto
215�Cat2�C/minto
325�C
at3�C/minleftfor1
0min
sucrose,maltose
trehalose,turanose,
cellobiose,palantinose,isom
altose
glucose,fructose,raffinose,
pann
ose
honeys
[120]
FID
SPB-1(30m£
0.25
mmi.d.;0.25mm
film
ofcrosslinkedmethylsilicone)
170�Cfor1
0minto
215�Cat15
� C/m
into
240�Cat1
� C/m
into
320�Cat
5�C/minleftfor2
0min
sucrose,maltose,a
,a-trehalose,
a,b-trehalose,turanose,cellobiose,
kojibiose,lam
inaribiose,nigerose,
maltulose,trehalulose,palantin
ose,
melibiose,gentio
biose,isom
altose
—honeys
[121]
Rtx-65
TG(25m£
0.25
mmi.d.;
0.1m
mfilm
ofcrossbond35%
dimethyl-65%
diph
enyl
polysiloxane)
200�Cfor2
0minto
270�Cat15
� C/
minto
290�Cat1�C/minto
300�C
at15
� C/m
inleftfor4
0min
FID
OV-101(25m£
0.25
mmi.d.)
180�Cto
280�Cat2�C/minto
290�Cat
10� C
/minleftfor1
5min
sucrose,maltose,lactose,m
altulose
glucose,fructose,galactose
enteralformulations
[122]
FID
CP-SIL5CB(25m£
0.25
mmi.d.;
0.25mmfilm
ofmethylsilicone)
180�Cfor3
6minto
300�Cat10
� C/m
inleftfor5
0min
sucrose
glucose,fructose,m
annose,
galactose,mannitol,
bornesito
l,myo-in
osito
l
coffeeandcoffeesubstitutes
[123]
MS
FID
SPB-1(25m£
0.25
mmi.d.;0.25mm
film
ofcrosslinkedmethylsilicone)
200�Cfor2
0minto
270�Cat15
� C/m
inleftfor4
0min
sucrose
glucose,fructose,inositols
fruitjuices
[124]
MS
FID
fusedsilicacapillarycolumn(7.5m
£0.25
mm)
180�Cfor2
0minto
270�Cat20
� C/m
inleftfor3
0min
sucrose
glucose,fructose,m
yo-in
osito
lorange
juice
[125]
MS
fusedsilicacapillarycolumn(22m£
0.25
mm)
MS
HP-5msUltraInert(15
m£
0.25
mmi.
d.;0.25m
mfilm
of5%
-phenyl)-
methylpolysiloxane)
180�Cfor2
minto
320�Cat7�C/min
leftfor2
min
lactose
—milk,cheese,yogh
urt
[126]
MS
SPB-1(30m£
0.25
mmi.d.;0.25mm
film
ofcrosslinkedmethylsilicone)
270�C
sucrose,maltose,lactose,
a,a-trehalose,turanose,cellobiose,
kojibiose,lam
inaribiose,nigerose,
maltulose,palantin
ose,melibiose,
isom
altose,sophorose,epilactose,
lactulose,leucrose
3-O-b-D-
galactopyranosyl-D-arabinose,
galactobioses,mannobioses
——
[119]
MS
TBR-1(30m£
0.25
mmi.d.;0.25mm
film)
200�Cfor1
5minto
270�Cat15
� C/m
into
290�Cat1�C/minLeftfor3
0min
lactose,a,a-trehalose,cellobiose,
laminaribiose,gentio
biose,
soph
orose
catechin,epicatechin,ethyl-
glucoside,glyceryl
–glucosides
wines
[127]
MS
DP-5MS(30m£
0.25
mm;0.25m)
50� C
to130�Cat30
� C/m
into
300�Cat
10� C/m
insucrose,maltose,m
elibiose
hexoses,hexoltioles,pentose
andpentosealcohols
cerealandpseudo
cerealflour
[128]
MS
DB-5or
HP-5M
S(30m
£0.25
mm;
0.25mmfilm)
50� C
to250�Cover58
min
sucrose,maltose,trehalose,
mannitol,sorbito
lstabilizersform
icrobial
preparations
[129]
MS
Rtx-65TG
(25m
£0.25
mmi.d.;
0.1mm
film
ofCrossbond35%
dimethyl–65%diph
enyl
polysiloxane)
170�Cfor1
0minto
215�Cat15
� C/m
into
240�Cat1
� C/m
into
320�Cat
5�C/minleftfor2
0min
sucrose,maltose,a
,a-trehalose,
a,b-trehalose,turanose,cellobiose,
kojibiose,lam
inaribiose,nigerose,
maltulose,trehalulose,palantin
ose,
melibiose,gentio
biose,isom
altose
raffinose,1-kestose,6-kestose,
pann
ose,erlose,neokestose,
malezito
se,m
altotriose,
isom
altotriose
honeys
[130,131]
Abbreviatio
ns:M
S-MassSpectrom
etry;FID-FlameIonizatio
nDetector.
196 M. POKRZYWNICKA AND R. KONCKI
-
spectra. Moreover, their retention time could also be
similar.That is why coupling of gas chromatography with mass
spec-trometry could give foul analytical information about
disac-charides contents.[119] For GC disaccharide detection
FlameIonization Detectors (FID) and Mass Spectrometers (MS)
aremainly applied. MS offer advantages of both qualitative
andquantitative information. Depending on the used
detector,nitrogen (for FID) or helium (for MS) is applied as
carrier gas.
A relatively novel and attractive alternative for LC and
GCtechniques is Capillary electrophoresis (CE). CE is an
electri-cally driven separation technique with many advantages
suchas significant cost-effectiveness, high separation speed and
largenumber of theoretical plates. CE requires minimal amounts(only
microliters) of buffer, organic solvents and additives. Acommon
problem with CE is its low detection sensitivitycaused by extremely
low sample injection volume (nanoliters)and small inner diameter of
the capillary. On the other hand,CE can be easily coupled with a
variety of optical and electro-chemical detectors. CE systems have
successfully applied fordisaccharides separation and determination.
Some examples ofsuch analytical applications are collected in Table
4. However,none of them is officially recommended analytical
method.
Separation principles of disaccharides in EC systems is simi-lar
to this applied in anion exchange chromatography. Thehigh pH value
is required, that is why the most common back-ground electrolyte is
50–75 mM sodium hydroxide. Capillarylength, critical for adequate
separation resolution, range from10[11] to 120 cm.[63] Further
improvement of separation effec-tivity can be obtained by chemical
modifications of active sur-face of used capillaries.[77] There are
also described applicationof microchip in place of conventional
fused silica capillary.[134]
Disaccharide detection in capillary electrophoresis may
seemproblematic. In case of electrochemical detection, because
ofhigh voltage applied during separation, detector electrodesshould
be somehow separated from capillary. Detector electro-des are
placed about 50mm opposite to the capillary outlet.[135]
In case of photometric detection because of lack of
chromo-phores for carbohydrates, precolumn derivatization[136]
orother transformation[137] is required. There is also possibility
toindirect absorbance detection. In this case an ionic chromo-phore
is added to background electrolyte for example:
2,6-pyri-dinedicarboxylic acid, maleic acid and phthalic acid,[77]
sorbicacid,[138] 1-naphthylacetic acid[63] or
2,6-pyridinedicarboxylicacid.[139] The detector receives a constant
signal due to the pres-ence of these substances. The analyte
displaces some of theseions, and detector signal decreases during
the passage of ananalyte through the detector. Similarly
chemiluminescencedetection can be applied with presence of luminol
in back-ground electrolyte.[140]
3.2. Bioselective methods
Sugars as natural compounds are participate in many
biotransfor-mation processes catalysed by various enzymes. Several
biocatalyticpathways of such transformations are useful in the
analytical chem-istry of sugars. The enzymes and enzymatic pathways
involved inbiorecognition and biodetection for three main
disaccharides areshown in Figures 3–5. In almost all cases enzymes
involved in sac-charide metabolism are highly selective. Often they
exhibit both
substrate and reaction specifity. This specific biorecognition
of dis-accharides by respective enzymes can be applied in
analyticalchemistry for the development of highly selective methods
fordetermination of particular analyte without the need of
separationof sugars and other components of samplematrix.
As can be seen from Figures 3–5, in the course of
severalenzymatic disaccharides biotransformations various
specificco-products, mediators and intermediates are consumed or
cre-ated. This way enzymatic biorecognition could be easily
cou-pled with various kinds of simple chemical detectors
andsensors. A type of applied detector is closely related with
classof last enzyme in biotransformation path of target sugar.
Areview of detection methods coupled with enzymatic recogni-tion of
disaccharides reported in the literature is presented inTable 5.
Except amperometry, spectrophotometry, conductom-etry, fluorimetry
and chemiluminometry, several detectiontechniques based on
potentiometry[50,144] and ion selectivedevices[145–148] are also
possible. Light addressable potentiome-try[144] and differential
pHmetry[50] are based on acidificationby phosphorylation reaction
catalyzed by glucokinase (GK E.C.2.6.1.2) or hexokinase (HK E.C.
2.7.1.1). Ion-Sensitive FieldEffect Transistors (ISFET) are
sensitive for hydrogen ions gen-erated during reaction with
dehydrogenases (describedexamples concern glucose[147–149] and
galactose[148] dehydro-genases), or generated after electrolysis of
hydrogen perox-ide.[146] A bit more complicated is situation with
ElectrolyteIsolator Semiconductor (EIS). This device sensitive for
fluorideions required application of 4-fluoroaniline as HPR
mediator.For disaccharides determination also entalpimetric
measure-ments of temperature changes during single[27] or
multistepenzymatic reactions can be utilized.[150] Examples of
applica-tion of Fourier Transform Near Infrared
Spectroscopy(FTNIR)[151] or coulometry[152] have been also
reported.
Predominantly, the biosensing schemes for disaccharides arebased
on sequence of enzymatic conversions of target analyteinto
detectable final species (Figs. 3–5) These so-called cascadeenzyme
reactions, consisting of at least two even to
foursteps,[29,58,145,161] realized in the analytical practise are
listed inTable 6. Most of them are based on specific enzymatic
hydroly-sis of target disaccharide to respective monosaccharides
(thefirst step of enzymatic cascade) and then on their
enzymaticallycatalysed oxidation (the second step) allowing
detection usingconventional instrumental methods (Tab. 5). The
enzymeapplied in the first step defines which disaccharide will be
bio-recognized and determined, whereas further enzymes
convertintermediate products into final detectable species. For
exam-ple, amperometric detection of lactose could be performedusing
only two enzymes: b-galactosidase (b-Gal, E.C. 3.2.1.23),glucose
oxidase (GOx, E.C. 1.1.3.4).[48] However, in somecases[47,53]
horseradish peroxidase (HPR E.C. 1.11.1.7) improv-ing detection of
enzymatically generated hydrogen peroxide isalso implemented into
biosensing system. Anotherextraordinary applied enzyme is
mutarotase (Mut, E.C.5.1.3.3),[3,26,146,153,158,160,165] because
most of the hydrolasesdecompose disaccharides into a- glucose,
whereas the nextenzymes (glucose oxidase (GOx, E.C. 1.1.3.4) or
glucose dehy-drogenases (GDh, E.C. 1.1.1.47)) are specific for
b-D-glucose.However, this enzyme is not crucial in these biosensing
path-ways, because mutarotation can occurs spontaneously
CRITICAL REVIEWS IN ANALYTICAL CHEMISTRY 197
-
Table4.
ApplicationofCapillaryElectrophoresisford
isaccharides
determ
ination.
Detectio
ncapillarydimensions
background
electrolyte
voltage
Determined
disaccharid
esOther
determ
ined
substance
Sample
Reference
DAD
indirectdetectionof
maleicacidat210nm
61.5cm
effectiveleng
th138.2mMsodium
hydroxide,
40mMmaleicacid,5
mM1-
tetradecyl--3-
methylim
idazoliumchlorid
e
¡21.7kV
sucrose,lactose,
cellobiose,
xylose,fructose,glucose,
galactose,raffinose,
cellotriose,cellotetraose,
cellopentaose,cellohexaose
biom
ass
[77]
70cm
totallength
22.5mmID
DAD
directdetectionat278
and240nm
71.5cm
effectiveleng
th130mMsodium
hydroxide,36
mM
disodium
phosph
ate
C17kV
xylobiose
xylo-oligosaccharides
enzymaticallyhydrolysed
pulp
[79]
80cm
totallength
50mm
IDDAD
indirectdetectionof
sorbicacidat254nm
8.5cm
effectiveleng
th20
mMsorbicacid,40mMsodium
hydroxide,0.2mMcetyl
trimethylammonium
brom
ide
C25kV
sucrose
glucose,fructose
honey
[138]
60cm
totallength
50mm
IDDAD
270nm
absorbing
interm
ediateform
atted
byph
otooxidatio
n
60cm
totallength
98mMsodium
hydroxide,120mM
sodium
chlorid
e¡1
4kV
sucrose,lactose
glucose,fructose
post-explosion
residu
es,
smokedevice,cough
syrup,applejuice,red
wine
[137]
50mm
ID
DAD
directdetectionat
270nm
60cm
effectiveleng
th130mMsodium
hydroxide,36
mM
disodium
phosph
ate
C10kV
cellobiose,trehalose
fructose,fucose,galactose,
glucose,myo-in
osito
l,mannitol,mannose,
rham
nose,ribose,sorbito
l,xylose
PinotN
oirred
wines
[68]
50cm
totallength
50mm
ID
DAD
indirectdetectionof
1-naph
thylaceticacidat
222nm
120cm
totallength
1mMsolutio
nof1-naph
thylacetic
acidadjusted
topH
12.5with
sodium
hydroxide
C25kV
sucrose,maltose
glucose,fructose,m
altotriose
wort
[63]
75mm
ID
DAD
directdetectionat
280nm
afterp
recolumn
derivatizationwith
p-am
inobenzoicacid
57cm
totallength
20mMsodium
tetraborate
C20kV
maltose
glucose,malto-oligosaccharides
beers,orange
andplum
juices
[136]
75mm
ID
DAD
indirectdetectionof
2,6-pyrid
inedicarboxylic
acidat275nm
104cm
effectiveleng
th112.5
20mM2,6-pyrid
inedicarboxylic
acid,0,5mM
cetyltrimethylammonium
hydroxideadjusted
topH
12.1
with
1Msodium
hydroxide
¡25kV
sucrose,lactose
glucose,fructose
yogu
rt,orang
ejuice,sake
mash,pickledapricot
[139]
cmtotallength
50mm
ID
Chem
iluminescence
65cm
totallength
0.018gluminolin10
mL0.02M
sodium
hydroxidesolutio
nof
10%dimethylsulfoxide
C15kV
sucrose
fructose,rhamnose,cyclodextrin
–[140]
25mm
ID
Cond
uctometric
4cm
effectiveleng
th75
mMsodium
hydroxide
C5kV
sucrose
glucose,fructose,ribose
energy
drinks
[11]
10cm
totallength
10mm
IDAm
perometric
(special
design
edgraphene–
cobaltmicrosphere
hybridpasteelectrodes)
40cm
totallength
75mMsodium
hydroxide
C12kV
sucrose,lactose
glucose,fructose,m
annitol
Honey,m
ilk[141]
25mm
ID
Amperometric
(special
design
edgraphene–
copp
ercomposite
electrodes)
40cm
totallength
75mMsodium
hydroxide
C12kV
sucrose,lactose
glucose,fructose,m
annitol
honey,milk,peach,
banana
[135]
25mm
ID
Amperometric
(nano-NiO
modified
carbon
paste
electrode)
27cm
totallength
50mMsodium
hydroxide
C10kV
sucrose
glucose,fructose,m
annitol
honey
[142]
25mm
ID
Amperometric
40cm
totallength
75mMsodium
hydroxide
C12kV
sucrose
paeoniflorin,paeonoside,
glucose,andfructose
MoutanCortex
[143]
25mm
IDAm
perometric
8cm
totallength
100mMsodium
hydroxide
100V
sucrose,lactose,trehalose
glucose,fructose,galactose,
mannose,xylose
honey
[134]
1mmID
onmicrochip
DAD
-Diode
ArrayDetector.
198 M. POKRZYWNICKA AND R. KONCKI
-
(effectively in the presence of phosphate ions[166]).
Sometimesadditional enzymes are used in the developed bioanalytical
sys-tems for eliminations of interferences. For example
speciallydesigned bioreactors with immobilized glucose oxidase
(GOx,E.C. 1.1.3.4) and catalase (Cat, E.C. 1.11.1.6) have been
appliedfor elimination influences from glucose in the course of
sucrosedetermination.[26,165]
There are also some enzymatic paths that allow
selectivedetermination only selected disaccharide without
theirhydrolysis to respective
monosaccharides.[5,27,46,54,58,151,156]
Sucrose phosphorylase (SP E.C. 2.4.1.1) decomposes sucroseto
glucose 1-phosphate, a substance which further enzy-matic
conversion (Fig. 3) is not interfered by glucose pres-ent in the
sample.[5,156] Similar biosensing scheme (Fig. 4)has been developed
for maltose using maltose phosphory-lase (MP E.C. 2.4.1.8) in the
first biorecognition step.[58]
Lactose can be oxidized by cellobiose dehydrogenase (CDHE.C.
1.1.99.18) and concentration of acceptor are mea-sured.[46,54]
However such approach is useful only in case ofsamples that do not
contain cellobiose and maltose.[170] Sin-gle enzymatic reaction can
be applied also for sucrose deter-mination after hydrolysis with
invertase, but in these casesuncommon detection method have to be
applied: thermom-etry for measuring of heat produced in the course
of bioca-talyzed[27] reaction or subtle changes of Infra-Red
spectrabetween substrate and product.[151]
Commercially available photometric assay kits for disaccha-ride
determination are based on two steps cascade
enzymaticreactions.[107,171–174] These kits contain soluble
enzymes, how-ever as can be seen from table 6 a large number of
bioanalyticalsystems dedicated for disaccharide determination is
based onimmobilized enzymes. They are immobilized in the form
of
Figure 3. Analytically useful enzymatic pathways for detection
of sucrose.
Figure 4. Analytically useful enzymatic pathways for detection
of lactose.
CRITICAL REVIEWS IN ANALYTICAL CHEMISTRY 199
-
mono- or polyenzymatic bioreactors or as biosensing
layersintegrated with respective detectors (biosensors). These
biosen-sors[27,46,53,54,57,70,146,148,150,156,157,167,175] and
bioreac-tors[3,5,26,64,70,71,155,160,161,165] are often applied in
systemsdesigned for flow analysis. Also measurements with
freeenzyme are performed under flow analysis
conditions,[17,153]
although it is connected with high enzyme consumption.
Suchapproach (flow analysis) allows mechanization of
multistepanalytical procedure and offers highly reproducible
conditionsof biochemical processes (precise control of reaction
time, tem-perature, reagent mixing etc.) and detection, what is
especiallyimportant in case of reported here biocatalytic
analytical meth-ods due to their kinetic, non- stationary
character.
Not only enzymes could be applied as biocatalytic mate-rials for
selective determination disaccharides. Severalmicroorganisms,
natural sources of commercially availableenzyme preparation, were
used for development of biosens-ing devices. Yeast cell were
successfully used instead ofinvertase[166] and coupled with GOx on
surface of biosen-sor. Saccharomyces crevisiae caused fermentation
of glucoseobtained from catalysed by lactase lactose hydrolysis
andallow determination with carbon dioxide electrode.[176]
Con-sumption of oxygen by specially grown mutants of Escheri-chia
coli K12 enable to monitoring of sucrose, maltose andlactose.[175]
A maltase-displayed bacteria and glucose dehy-drogenase-displayed
bacteria were co-immobilized on multi-
Figure 5. Analytically useful enzymatic pathways for detection
of maltose.
Table 5. Possible detection methods defined by the last enzyme
in biocatalytic path, Table 5 Detection type according to last
enzyme type in cascade enzymatic path.
Final enzyme type
Oxidases: GOx,GaOx, PyOx
Dehydrogenases: GDh, GaDh,FDh, G6PDh, CDh Peroxidase: HrP
Detection technique Amperometry oxygen
consumption,[17,48,57,153,154]H2O2 detection[3,26,155]
reduction of acceptor[46,54,71,156,157]
oxidation of mediator [47,53]
Conductometry increased in conductivity afterlactone
dissociation [158,159]
— —
Spectrophotometry H2O2 chromogenic reactions[160] with G6PDH
absorbance of
NADPH (340 nm) [29,58,64,161];with FDH reduction of MTTto MTT
formazan (570 nm)[41,162]
product of reaction of 4aminoantrypine and Phenol4 sulphonic
acid salt(500 nm) [163]
Fluorimetry — Fluorescence of NADPH(ex- 340 nm; em 460 nm)
[5]
reduction of Amplex Red tohigh fluorescent resorufin(ex- 550 nm;
em-585 nm)[28]
Chemiluminometry oxidation of luminol by H2O2[164,165]
— —
Abbreviations: CDh- cellobiose dehydrogenase; FDh- fructose
dehydrogenase; G6PDh- glucose-6 phosphate dehydrogenase; GDh-
glucose dehydrogenase; GaDh- galac-tose dehydrogenase; GOx- glucose
oxidase; GaOx- galactose oxdase; HrP- horseradish peroxidase; MTT-
3-(4, 5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide;NADPH-
Nicotinamide adenine dinucleotide phosphate; PyOx- pyranose
oxidase.
200 M. POKRZYWNICKA AND R. KONCKI
-
Table6.
Enzymaticdeterm
inationofdisaccharid
es.
Cascadeenzymaticreactio
nsequ
ence
Target
Analyte
1st
enzyme
2nd
enzyme
3rd
enzyme
4th
enzyme
detection
method
immobilizatio
nmethod
Other
determ
ined
sugars
Sample
Reference
immob
ilizedenzyme(biosensor)
sucrose
invertase
mutarotase
glucoseoxidase
horseradish
peroxidase
EIS
cross-linking
with
GA
glucose
—[145]
sucrose
invertase
mutarotase
glucoseoxidase
—Conductometry
cross-linking
with
BSA
andGA
glucose
orange
andapplejuices
[158]
sucrose
invertase
mutarotase
glucoseoxidase
—ISFET
photo-crosslinkable
polymer(PVA
SbQ)
glucose
—[146]
sucrose
invertase
glucoseoxidase
catalase
—Calorim
etry
covalent
attachmentto
activated
with
GA
glucose
—[150]
sucrose
sucroseph
osph
orylase
phosph
ogluco-m
utase
glucose-6ph
osph
ate
dehydrogenase
—Am
perometry
entrappedincarbon
pasteelectrodematrix
—pineapple,peachapple
juice
[156]
sucrose
invertase
fructose
dehydrogenase
——
Amperometry
entrapmentw
itha
dialysismem
brane
fructose,glucose
cond
ensedmilk,
referencematerial
[157]
sucrose
invertase
glucosedehydrogenase
——
ISFET
cross-linking
with
BSA
andGA
glucose
—[147]
sucrose
invertase
glucokinase
——
Ligh
tadd
ressable
potentiometry
cross-linking
with
BSA
andGA
glucose
—[144]
sucrose
invertase
glucosedehydrogenase
——
ISFET
cross-linking
with
GA
glucose
—[148]
sucrose
invertase
——
—Thermom
etry
cross-linking
with
BSA
andGA
—sugarcanejuice
[27]
lactose
b-galactosidase
glucoseoxidase
horseradishperoxidase
—Am
perometry
entrapmentw
ithin
dialysismem
brane
glucose
chocolate,dairy
samples
[53]
lactose
b-galactosidase
glucoseoxidase
horseradishperoxidase
—Am
perometry
cross-linking
with
GA
glucose
milk
[47]
lactose
b-galactosidase
glucoseoxidase
horseradishperoxidase
—Am
perometry
—glucose
milk,cheese,yogh
urt
[126]
lactose
b-galactosidase
mutarotase
glucoseoxidase
—Am
perometry
cross-linking
with
GAand
b-cyclodextrin
and
coveredby
nafion
glucose
—[49]
lactose
b-galactosidase
glucoseoxidase
——
Conductometry
cross-linking
with
BSA
andGA
glucose
milk
[159]
lactose
b-galactosidase
glucoseoxidase
——
Amperometry
cross-linking
with
gelatin
eandGA
glucose
milk
[48]
lactose
b-galactosidase
glucoseoxidase
——
Voltammetry
cross-linking
with
GA,
covalent
bond
edwith
polyazetidine
glucose
—[167]
lactose
b-galactosidase
glucosedehydrogenase
——
ISFET
cross-linking
with
GA
glucose
—[149]
lactose
b-galactosidase
galactosedehydrogenase
——
ISFET
cross-linking
with
GA
galactose
—[148]
lactose
lactase
galactoseoxidase
——
Amperometry
Lang
muir-Blodgetfi
lmof
poly(3-hexyl
thioph
ene)/stearic
acid
galactose
—[168]
lactose
cellobiosedehydrogenase
——
—Am
perometry
physicaladsorptio
n,entrapmentw
itha
dialysismem
brane
—milk
[46,54]
maltose
amylo-
glucosidase
mutarotase
glucoseoxidase
horseradish
peroxidase
EIS
cross-linking
with
GA
glucose
—[54]
maltose
a–g
lucosidase
glucokinase
—Ligh
tadd
ressable
potentiometry
cross-linking
with
BSA
andGA
glucose
—[145]
maltose
a–g
lucosidase
glucosedehydrogenase
——
ISFET
cross-linking
with
GA
glucose
—[144]
(Continuedon
nextpage)
CRITICAL REVIEWS IN ANALYTICAL CHEMISTRY 201
-
Table6.
(Continued)
Cascadeenzymaticreactio
nsequ
ence
Target
Analyte
1st
enzyme
2nd
enzyme
3rd
enzyme
4th
enzyme
detection
method
immobilizatio
nmethod
Other
determ
ined
sugars
Sample
Reference
maltose
amylo-
glucosidase
glucoseoxidase
——
Cyclicvoltammetry
physicaladsorptio
nglucose
beer
[148]
maltose
amylo-
glucosidase
glucoseoxidase
——
Amperometry
cross-linking
with
BSA
andGA
glucose
starch
hydrolysate
[60]
maltose
a–g
lucosidase
pyranose
oxidase
——
Amperometry
cross-linking
with
chito
san,carbon
nanotube
andGA
glucose,galactose,xylose
beer
[57]
lactulose
b-galactosidase
(inbioreactor)
fructose
dehydrogenase
——
Amperometry
cross-linking
with
BSA
andGA
fructose
milk
[154]
immob
ilizedenzyme(bioreactor)
sucrose
invertase
phosph
oglucose
isom
erase
hexokinase
glucose-6ph
osph
ate
dehydrogenase
Spectrophotometry
covalent
attachmentto
CPGallkylam
inated
with
APTS
and
form
ylated
with
GA
glucose,fructose
synthetic
samples
[161]
sucrose
invertase
mutarotase
glucoseoxidase
catalase
Amperometry
covalent
attachmentto
Amino-Cellulofine
activated
byGA
glucose,fructose
(byFD
H)C
ocacola,kiwi,apple,
banana,m
andarin
[26]
sucrose
invertase
mutarotase
glucoseoxidase(in
solutio
n)catalase
Chem
iluminescence
covalent
attachmentto
CPGallkylam
inated
with
APTS
and
form
ylated
with
GA
glucose
Pepsi,coke,cereal,cake
mix
[165]
sucrose
invertase
mutarotase
glucoseoxidase
—Am
perometry
cross-linking
with
BSA
andGAon
pig’ssm
all
intestine
glucose
fruitjuices
[3]
sucrose
invertase
mutarotase
glucoseoxidase
—Spectrophotometry
covalent
attachmentto
CPGallkylam
inated
with
APTS
and
form
ylated
with
GA
glucose
—[160]
sucrose
invertase
mutarotase
glucoseoxidase
—Am
perometry
cross-linking
tocellulose
mem
branewith
BSA
andGA
glucose
—[155]
sucrose
sucroseph
osph
orylase
phosph
ogluco-m
utase
glucose-6ph
osph
ate
dehydrogenase
—Fluorometry
covalent
attachmentto
CPGallkylam
inated
with
APTS
and
form
ylated
with
GA
—ionicsoftdrink,cola,
orange
juice
[5]
sucrose
invertase
glucoseoxidase
——
Chem
iluminescence
covalent
attachmentto
CPGallkylam
inated
with
APTS
and
form
ylated
with
GA
glucose
—[164]
sucrose
invertase
glucoseoxidase
(biosensor)
——
Amperometry
Glucose
oxidasesand
wich
mem
braneand
invertasebearingsilk
reactor
glucose
—[169]
sucrose
invertase
——
—FT-NIRspectroscopy
covalent
attachmentto
silicon
chipsilanized
with
ATPS
and
form
ylated
with
GA
—softdrinks
[151]
lactose
lactase
glucoseoxidase
——
Chem
iluminescence
covalent
attachmentto
CPGallkylam
inated
with
APTS
and
form
ylated
with
GA
glucose
—[164]
202 M. POKRZYWNICKA AND R. KONCKI
-
maltose
amylo-
glucosidase
glucoseoxidase
——
Chem
iluminescence
covalent
attachmentto
CPGallkylam
inated
with
APTS
and
form
ylated
with
GA
glucose
—[164]
lactulose
b-galactosidase
(insolutio
n)fructose
dehydrogenase
——
Amperometry
covalent
attachmentto
CPGallkylam
inated
with
APTS
and
form
ylated
with
GA
fructose
milk
[71]
trehalose
trehalase
hexokinase
(insolutio
n)glucose-6ph
osph
ate
dehydrogenase(in
solutio
n)
—Spectrophotometry
covalent
attachmentto
Epoxyresin
glucose
ferm
entatio
nbroth
[64]
non-
immob
ilizedenzyme
sucrose
invertase
hexokinase
phosph
oglucose
isom
erase
glucose-6ph
osph
ate
dehydrogenase
Spectrophotometry
—glucose,fructose
beet
root
[29]
sucrose
invertase
mutarotase
glucoseoxidase
—Am
perometry
—glucose
sugarb
eetm
olase
[153]
sucrose
invertase
glucoseoxidase
horseradishperoxidase
—Fluorim
etry
—glucose
sugarb
eet
[28]
sucrose
invertase
glucoseoxidase
horseradishperoxidase
—Spectrophotometry
—glucose
coffeebeans
[163]
sucrose
invertase
fructose
dehydrogenase
——
Spectrophotometry
—fructose
bloodserum,urin
e[41]
sucrose
invertase
glucoseoxidase
——
Amperometry
—glucose
orange
andapplejuice,
icecream,condensed
milk,jellies,greenpea,
corn,w
heat,peanuts
[17]
lactose
b-galactosidase
glucokinase
hexokinase
—DifferentialpH
techniqu
e—
glucose
milk
[50]
lactose
lactase
glucoseoxidase
——
Amperometry
—infant
form
ula
[17]
maltose
maltose
epimerase
maltose
phosph
orylase
phosph
ogluco-m
utase
glucose-6ph
osph
ate
dehydrogenase
Spectrophotometry
——
potato
solublestarch
[58]
maltose
a–g
lucosidase
glucoseoxidase
——
Amperometry
—corn
andmaltsyrup
[17]
maltose
a–g
lucosidase
glucoseoxidase
——
Coulom
etry
—glucose
—[152]
lactulose
b-galactosidase
hexokinase
——
DifferentialpH
techniqu
e—
glucose,fructose
milk
[50]
lactulose
b-galactosidase
fructose
dehydrogenase
——
Spectrophotometry
—fructose
milk
[162]
Abbreviatio
ns:APTS-3-am
inopropyltriethoxysilane;BSA-b
ovineserumalbu
min;CPG
-controlledporous
glass;EIS-electrolyteisolator
semicondu
ctor;FT-NIR-FourierTransform
NearInfraredSpectroscopy;G
A-glutaraldehyde;ISFET-
Ion-Sensitive
FieldEffectTransistor.
CRITICAL REVIEWS IN ANALYTICAL CHEMISTRY 203
-
walled carbon nanotubes modified glassy carbon electrodeto
obtain electrochemical biosensor for maltose and glu-cose.[177] In
the course of another investigations[178] therepressor of
Escherichia coli lac operon has been engineeredas altered effector
for selective recognition of lactulose.
Whereas biorecognition of various sugars using enzymesand
biocatalytic materials is widely reported in the
analyticalliterature, till now the immunoassays for disaccharide
determi-nation are not developed. It is practically impossible to
grownan antibody for sucrose due to lack of its
immunogenicity.Although maltose and lactose antibodies are
available, theiranalytical application is not reported in the
literature.
3.3. Spectroscopic methods
Disaccharides, like all organic compounds, have reach
characteris-tic spectra in Infra-Red (IR) range of wavelength with
specificabsorption bands (table 7). The application of IR
spectroscopy fordisaccharides determination required absorbance
measurements atmany wavelengths simultaneously and then the use of
special mul-tivariate statistical techniques to relate spectral
data with the con-centration of the chosen component. Such
techniques as multiplelinear regression (MLR) or partial least
squares (PLS) regressionequation can predict the concentration of
each constituent fromthe absorbance values at selected wavelengths.
Because of the struc-tural similarities of sugars the spectra can
overlap for each other.The baseline can variate in various
apparatus and because of ambi-ent conditions (especially
temperature). In case of aqueous solutionthere is high background
spectrum of water. Sample surface imper-fections can effect in
nonlinear, inhomogeneous and anisotropiclight scattering. In some
cases high frequency detector noise canoccur. Obviously, many other
compounds present in the samplecan contain similar functional group
and thus can interfere.[179]
For IR data processing and interpretation rather
sophisticatedchemometric methods are required. In experiments from
two[181]
to more than fifty[182] spectra are recorded and averaged. In
manycases also further data handling are processed. For smoothing
andresolving overlapping peaks first and second derivatives
computa-tion[20,182,183] or Savitzky-Golay (SG)
filter[8,19,30,73,180,184–186] couldbe applied. Because spectra
included very extensive data StandardNormal Variates
(SNV),[8,18,30,184,185,187] Principal ComponentAnalysis
(PCA)[18,56] or Genetic Algorithm (GA)[187] could be
applied to reduce the dimensionality of the data in order to
extractmain and remove insignificant information. GA together
withArti-ficial Neural Network (ANN) could also be coupled to
define ana-lyte origin when group of samples is compared.[20] To
obtaincalibration curves the Partial Least Square Regression (PLS)
aremostly applied, however the use of Multiple Linear
Regression(MLR)[187] and Principal Component Regression (PCR)[4] is
alsoreported in the literature. The PLS and PCR use data
reductiontechniques to extract from all extensive data much smaller
amountof new variables representative for most of the variability
in sam-ples. These newly defined variables can be used to create a
calibra-tion curve or develop a regression equation to predict
theconcentration of disaccharide in sample. In those methods, it is
notnecessary to reduce data dimension, as it is in MLR where only
alimited number of wavelengths are used. Examples of applicationof
IR for disaccharide determination using various chemometrictools
for data processing are presented in table 8. As can be seenfrom
this table, chemometrically supported IR spectroscopy is use-ful
for analysis of real samples having quite complexmatrix.
It is worth to notice that not only IR spectroscopy can be
sup-ported by advanced chemometric methods to estimate
disacchar-ides content in sample. There is also example of
measurements invisible range with the use of second derivative
spectra in combina-tion with PLS regression modelling for
determination of sucroseand trehalose in olive leaves.[34] Some
electrochemical techniquesmay require statistical data treatment to
could be applied for selec-tive disaccharide determination. An
Electrochemical ImpedanceSpectroscopy (EIS) was applied for
determination of sucrose, glu-cose, fructose and total sugar
content in pineapple fruit.[21,189] Afterstandard addition,
measurement was performed and ANN techni-ques was applied to
predict specific mathematical models for eachone of determined
sugars. PLS method was also used to model therelationship between
the EISmeasurements and the s