Effects of molecular characteristics of on konjac glucomannan glass transitions of potato amylose, amylopection and their mixtures

75

8

Research ArticleReceived: 18 May 2010 Revised: 4 October 2010 Accepted: 1 November 2010 Published online in Wiley Online Library: 13 December 2010

(wileyonlinelibrary.com) DOI 10.1002/jsfa.4247

Effects of molecular characteristics of on konjacglucomannan glass transitions of potatoamylose, amylopection and their mixturesLi Guo,a,b Qin Lianga and Xianfeng Dua∗

Abstract

BACKGROUND: The purpose of this study was to explore further the functions of konjac glucomannan (KGM) in starch-basedfoods. Experiments were carried out using the mixed amylose/amylopectin/KGM system as a model. High-speed differentialscanning calorimetry (hyper-DSC) with the support of high-performance size exclusion chromatography (HPSEC) equipped withmulti-angle laser light scattering (MALLS) and differential refractive index (RI), X-ray diffractometry (XRD) and viscosimetry wasused to investigate the effects of KGM on glass transition temperatures (Tgs) of mixtures with different amylose/amylopectinratios.

RESULTS: Hyper-DSC results showed that the Tgs of amylose, amylopection and their mixtures decreased with increasingconcentration of KGM. Based on the molecular characteristics of KGM, HPSEC-MALLS-RI, viscosimetry and XRD results showedthat the molar masses of KGM ranged from 1.023×106 to 1.329×106 g mol−1; the root mean square (RMS) radii were distributedfrom 110.5 to 129.6 nm, and Mw/Mn was 1.017. KGM was a linear molecule with random-coil conformation in solution and thecrystallinity was 0.00%.

CONCLUSION: It is suggested that the addition of KGM has plasticizing effects on the structures of amylose and amylopectin,which can increase free volume and molecular movement of amylose and amylopectin chains, resulting in a decrease in theirTgs.c© 2010 Society of Chemical Industry

Keywords: konjac glucomannan (KGM); amylose; amylopectin; glass transition; molecular characteristics

INTRODUCTIONKonjac glucomannan (KGM) is a high-molecular-weight, water-soluble and non-ionic polysaccharide found in roots and tubersof the Amorphophallus konjac plant. KGM has β-(1 → 4)-linked D-glucose and D-mannose residues as the main chain with branchesjoined through the C-3 carbon of D-glucosyl and D-mannosylresidues.1 Katsuraya et al.2 reported that KGM is a β-(1 → 4)-linked polysaccharide composed of a D-glucosyl and D-mannosylbackbone lightly branched, possibly through β-(1 → 6) glucosylunits. It consists of (1 → 4)-linked β-D-mannose and D-glucoseunits in the molar ratio 1.5–1.6 : 1,3 with about 1 in every 17–19units being acetylated at the C-6 position.4 KGM has been reportedto have a broad range of therapeutic and nutritional values,such as lowering blood sugar and blood lipids, preventing andcuring constipation, promoting intestinal health, lowering weight,retarding caducity and enhancing human immunity.5 Additionally,KGM can be extruded into films6 or form blend membranes7 – 9

for coating and packaging applications in the food industry.Therefore KGM has development potential for a large number ofcommercial uses in the medicine, hygiene and food fields. Themolecular mass and molecular mass distribution of KGM are theessential structural characteristics having a great influence onits functionality in food processing, and its conformation is veryimportant for its rheological and morphological properties.10 Morespecifically, the functionality of KGM is dependent on molecular

weight. Li et al.11 showed that KGM manifests its functionality onlyif its molecular dimension was limited within a specified range. Themechanical properties of KGM films, gels and fiber are associatedwith its molecular weight.12 The molecular weight is also knownto be an important quality control parameter13 in monitoring KGMproduction on a commercial scale. Some studies8,14 have alsoindicated that the functionality of KGM might be affected by itschain geometry and parameters, especially by root mean squareradius and polydispersity.

The glass transition temperature (Tg) is an important parameterin determining the mechanical properties of amorphous polymersand controlling the kinetics of crystallization of amorphousmaterials. The Tg of starch is a critical thermal property instarch-based products. Recently, high-speed differential scanningcalorimetry (hyper-DSC) has attracted much attention due to the

∗ Correspondence to: Xianfeng Du, Anhui Agricultural University, 130 WesternChangjiang Road, Hefei City 230036, Anhui Province, China.E-mail: [email protected]

a Key Laboratory of Tea Biochemistry and Biotechnology, Anhui AgriculturalUniversity, Hefei 230036, China

b Key Laboratory of Cigarette Smoke, Technology Center of Shanghai Tobacco(Group) Corp., Shanghai 200082, China

J Sci Food Agric 2011; 91: 758–766 www.soci.org c© 2010 Society of Chemical Industry

75

9

Effects of konjac glucomannan on glass transitions of potato starch www.soci.org

fact that it can detect glass transitions using a sufficiently highheating rate, up to 500 ◦C min−1. The high heating rate allowsthe weak Tg of starch to be visible since the high rate increasesthe sensitivity of thermal events.15 – 17 KGM is reported to interactsynergistically with individual carrageenan, xanthan, gellan, andcorn starch.9,18 However, publications on synergistic interactions ofKGM with potato starch are scarce. Therefore, to further understandthe functions of KGM on potato starch, some experiments wereconducted using the mixed potato amylose/amylopectin/KGMsystem as a model. In performing these experiments, the structureproperties of KGM were studied using high-performance sizeexclusion chromatography equipped with multi-angle laser lightscattering and differential refractive index detectors (MALLS-HPSEC-RI) and viscosimetry. The glass transition temperatures(Tgs) of the mixed systems were determined using hyper-DSC witha heating rate up to 400 ◦C min−1.

MATERIALS AND METHODSMaterialsAmylose (A0512) and amylopectin (A8515) from potato starchwere purchased from Sigma (St Louis, MO, USA). The purifiedkonjac glucomannan (KJ-30, ≥95 g kg−1, dry basis) was purchasedfrom Wuhan Qiangsen Konjac Production Co., Ltd (Wuhan, China).Sodium chloride, toluene, mannase (10 000 U) and bovine serumalbumin globular protein (BSA, Mw = 66.7 kDa, Rg = 2.9 nm) werepurchased from Sigma; Micro PES (polyethersulfone) membranewas obtained from Membrana Co. (Wuppertal, Germany). All otherchemicals were of the highest purity commercially available.

Molecular characteristics of KGMChromatographic fractionationIn this experiment, one guard column, SB-G (6 × 50 mm, 10 µm)and two analytical size-exclusion columns, SB-805HQ (8×300 mm,13 µm) and SB-804HQ (8 × 300 mm, 10 µm) (Showa Denko KK,Tokyo, Japan) were connected in tandem. The mobile phase usedfor HPSEC was 0.2 mol L−1 NaCl, filtered through a 0.45 µm MicroPES membrane filter, at a flow rate of 0.5 mL min−1 and a refractiveindex of 1.333 at 658.0 nm at 30 ◦C. The injector and columns weremaintained at 30 ◦C with a CH-460 column heater and a TC-50 controller (Eppendorf, Madison, WI, USA). Before injectingit into the HPSEC system, the KGM solution was accuratelyfiltered through 0.8 µm filters and injected (0.3 mL) onto thehigh-performance size-exclusion columns.

HPSEC-MALLS-RI systemThe HPSEC-MALLS-RI system consisted of a pump (515 HPLC,Waters, Milford, MA, USA), a Rheodyne 7725i injector (Waters)with a 200 µL injection loop (Rheodyne , IDEX Corporation, LakeForest, IL, USA), two HPSEC columns, a MALLS detector (DAWNHELEOS, Wyatt Technology, Santa Barbara, CA, USA), and an RIdetector (Optilab Rex, Wyatt Technology). The molar mass andRMS radius of KGM in 0.2 mol L−1 aqueous NaCl were measuredwith a multi-angle laser photometer, which was equipped with a 30mW solid-state laser (λ = 658.0 nm) and a K-5 flow cell at angles of34.9◦, 42.9◦, 51.6◦, 60.0◦, 69.4◦, 79.7◦, 90.0◦, 100.3◦, 110.6◦, 121.2◦

and 132.1◦ at 30 ◦C. The RI calibration constant was measuredwith a series of NaCl standards. The 90◦ photodiode detector ofMALLS was calibrated (calibration constant: 1.4462 × 10−4) usingtoluene (HPLC grade). The rest of the 17 photodiode detectorsat all scattering angles were normalized in accordance with the

90◦ detector using a bovine serum albumin globular protein(Mw = 66.7 kDa, Rg = 2.9 nm).

A monodisperse sample with a polydispersity of<1.05 is neededto determine the volume delay between the DAWN HELEOS andRI instruments.23 In order to obtain accurate results, the collectioninterval was set 0.500 s in this experiment. Normalization wasrequired to correct slight differences in light beam collimation,photodiode sensitivity, and refractive index effects among theMALLS detectors. Use of a monodisperse BSA monomer (BSAStandard 50 000; Rg = 2.9 nm, Sigma) enabled the volume delay(0.145 mL) between MALLS and RI to be determined, permittingcorrect alignment of the MALLS and RI signals as required forcalculation of the molecular weight for each data slice.

The differential refractive index increment (dn/dc) defines howa solution’s refractive index (n) changes with solute concentration(c). This parameter was required for converting RI voltages tosolute concentrations at each data slice across a chromatographicpeak and was determined individually for each product. Thedn/dc value of the KGM sample in 0.2 mol L−1 aqueous NaCl wasdetermined using a calibrated Optilab refractometer (Dawn-DSP,Wyatt Technology) at 658 nm and 30 ◦C to be 0.140 mL g−1.

Experimental data obtained from MALLS and RI detectors wereanalyzed using Astra V software (Version 5.1, M1000 Rev. C, WyattTechnology). The Astra software was utilized to analyze the light-scattering data using a Berry plot and Mw, Mn, Mz, Rn, Rw and Rz aswell as Mw/Mn were obtained. Variability in the data collection wasalso determined using Astra V 5.3.2.7 software (Wyatt Technology)with the standard deviation and coefficient of variation of replicateanalytical runs.

Preparation of KGM with different molecular weightsKGM was dissolved completely in deionized water, and thenhydrolyzed with 200 U g−1 mannase at 30 ◦C for 0 min, 4 min,8 min, 12 min and 16 min, respectively. The hydrolyzed KGM wastreated promptly at 100 ◦C for 10 min to inactivate the mannase.The degraded product solutions were centrifuged, and then thekonjac mannans were precipitated with aqueous methanol anddried under vacuum freeze-dry. Finally, a series of powdered KGMswith different molecular weights were obtained. They were codedas KGM-1, KGM-2, KGM-3, KGM-4 and KGM-5, respectively. Themolecular weight of each fraction was determined by using theHPSEC-MALLS-RI system.

Intrinsic viscosity of KGMThe intrinsic viscosity of KGM in 0.2 mol L−1 NaCl aqueous solutionwas measured using an Ubbelohde viscometer (0.57 mm) at25.0 ± 0.1 ◦C. Every value was measured at least five times andthen averaged.

X-ray diffractometry (XRD)Diffractograms were obtained at 20 ◦C with an XD-3 X-raygenerator (Purkinje General Instrument Co. Ltd, Beijing, China)operating at 40 kV and 30 mA. The scanning region of thediffraction angle (2θ ) was from 5◦ to 50◦ at a step size of 0.02◦

and scanning speed of 4◦ min−1. Cu Ka radiation (λ = 1.5406 nm)was selected using a graphite monochromator. A divergence slitof 1◦ and a receiving slit of 0.3◦ were chosen. The overall degree ofcrystallinity was calculated as the ratio of the area of the crystallinereflections to the overall area, using the method of Hermans andWeidinger.19 The total area and crystalline area were measuredusing Jade software.

J Sci Food Agric 2011; 91: 758–766 c© 2010 Society of Chemical Industry wileyonlinelibrary.com/jsfa

76

0

www.soci.org L Guo, Q Liang, X Du

Table 1. Sample code, constituents with ±0.0001 g precisionand water contents (mean ± standard deviation, n = 3) ofamylose/amylopectin/KGM

Raw material (g)

Sample code Amylose Amylopectin KGM Water contents (%)

1 – – 0.5000 10.03 ± 0.1

2 0.5000 – – 10.81 ± 0.3

3 0.2501 – 0.2500 10.96 ± 0.1

4 0.1671 – 0.3329 10.51 ± 0.4

5 0.1250 – 0.3750 10.00 ± 0.1

6 0.1013 – 0.4001 11.29 ± 0.1

7 – 0.5000 – 10.25 ± 0.4

8 – 0.2501 0.2500 10.11 ± 0.1

9 – 0.1669 0.3331 10.07 ± 0.2

10 – 0.1250 0.3750 10.81 ± 0.1

11 – 0.1001 0.4000 11.12 ± 0.2

12 0.2500 0.2501 – 10.44 ± 0.2

13 0.1701 0.1700 0.1700 10.23 ± 0.6

14 0.1250 0.1251 0.2500 10.92 ± 0.2

15 0.1001 0.1000 0.3001 10.17 ± 1.9

16 0.0830 0.0834 0.3335 10.81 ± 0.3

17 0.0714 0.0713 0.3572 10.94 ± 0.8

18 0.1667 0.3334 – 10.16 ± 0.1

19 0.1250 0.2500 0.1250 10.61 ± 0.2

20 0.1003 0.2001 0.2001 10.75 ± 1.2

21 0.0833 0.1666 0.2500 10.21 ± 0.7

22 0.0723 0.1421 0.2856 10.75 ± 0.2

23 0.0625 0.1250 0.3124 10.79 ± 0.9

24 0.1250 0.3750 – 10.44 ± 1.1

25 0.1002 0.3000 0.1001 10.14 ± 0.8

26 0.0833 0.2503 0.1664 10.78 ± 0.3

27 0.0713 0.2141 0.2144 11.07 ± 0.1

28 0.0625 0.1875 0.2501 10.69 ± 0.1

29 0.0554 0.1668 0.2778 11.08 ± 0.8

30 0.1000 0.4001 – 10.96 ± 0.2

31 0.0834 0.3332 0.0835 10.26 ± 0.2

32 0.0714 0.2856 0.1431 10.00 ± 0.2

33 0.0623 0.2502 0.1875 10.22 ± 0.1

34 0.0556 0.2223 0.2221 10.99 ± 0.4

35 0.0501 0.2001 0.2500 10.88 ± 0.7

Thermal analysisFilm preparationFilms were prepared by casting the mixture paste on glass sheets.Potato amylose, amylopectin and KGM were mixed in a dry format the appropriate weight ratio (w/w) (Table 1). The 5 g kg−1

suspensions were heated in sealed conical flasks in a water bath,initially at 70 ◦C in order to maintain the same temperaturegradient during gelatinization. Subsequently, the temperaturewas increased to 120 ◦C under 0.1 MPa and maintained for 4 hin an autoclave. Low shear rate (16 × g) was used during theheating process. The pastes were poured onto glass sheets anddried at 60 ◦C in a controlled-environment chamber (25 ◦C and50% relative humidity) for 24 h. The films obtained were removedfrom the glass sheets. The thickness of the films was measuredusing a digital indicator (Mitutoyo IDC-112CB, Mitutoyo Corp,Kawasaki, Kanagawa, Japan) with an accuracy of ±3 µm. The filmswere 180–200 µm thick. The moisture content in each sample was

estimated from weight loss of sample after drying at 130 ◦C until aconstant weight was reached. Triplicate replicate measurementswere performed for each analysis. The moisture content of all filmsranged from 10.00% to 11.29%. The detailed moisture contents ofthe films are listed in Table 1.

Hyper-DSCThe Tgs of the films were determined using hyper-DSC. Calorimetricmeasurements were performed on a DSC Diamond-1 (PerkinElmer,Waltham, MA, USA) with an internal coolant (Intercooler 1P).Measurements were carried out under nitrogen purge gas ata flow rate of 50 mL min−1. The instrument was calibrated fortemperature and heat flow using indium and zinc as standardsat a heating rate of 10 ◦C min−1. A baseline for an empty panwas established for each corresponding heating rate. An emptyaluminium pan was used as a reference. Triplicate samples(3.0 ± 0.1 mg, dry basis) were sealed hermetically in aluminiumpans. Heating rates of 50, 100, 250, 400 and 500 ◦C min−1 wereemployed. The inflection point of the step in the heat flow curve inthe heating scan was taken as the Tg, which corresponded to thetemperature at which one-half of the change in the heat capacityoccurred.14 The samples were scanned between 0 and 150 ◦C.

The temperature profile for each sample involved heating from0 to 150 ◦C, followed by cooling as rapidly as possible in thecalorimeter to 20 ◦C at 50 ◦C min−1 and maintaining at 4 ◦C for 5,10, 15, 20, 25 and 30 days. The stored samples were rescannedfrom 0 to 150 ◦C at a heating rate of 400 ◦C min−1 to study theeffects of KGM on the physical aging of amylose and amylopectin.

Statistical analysisResults were expressed as mean ± standard deviation oftriplicate analyses for each sample, unless otherwise stated.Statistical significance was assessed with one-way analysis ofvariance (ANOVA) using ORIGIN 7.5 for Windows (OriginLabInc., Northampton, MA, USA). Treatment means were consideredsignificantly different at P < 0.05.

RESULTS AND DISCUSSIONMolecular characteristics of KGMThe HPSEC-MALLS-RI system was used to determine the molecularweight of KGM (Fig. 1). It shows the peaks given by the 90◦

light scattering and RI detectors are of similar size and shape,and almost completely overlaid, which suggests the interdetectordelay procedure has been correctly determined when the systemis properly aligned. Figure 1 shows the Berry plots for KGM in0.2 mol L−1 NaCl aqueous solution at 30 ◦C. It is often sufficientto choose a fit degree of 2, which gives the smallest error. In thisanalysis one data slice was collected every second, in which thelight-scattering signal extrapolated to zero scattering angle wasdivided by the signal from the differential refractometer to obtainthe molar mass and RMS radius of KGM, as shown by a typicalBerry plot in Fig. 1. For KGM at elution volume (7.252 mL) for peakslice 11 772 on the Berry plot, the molar mass and RMS radius of1.013 × 106 g mol−1 and 111.4 nm were evaluated, respectively.

SEC chromatograms and the molar mass and RMS radius data asa function of elution volume of KGM in 0.2 mol L−1 aqueous NaClare shown in Fig. 2. Figure 2(a) shows a gradual decrease in molarmass with increasing elution volume. Figure 2(b) indicates that theaggregates and individual molecules exist as a relatively compactand spherical conformation in aqueous solution, thereby resulting

wileyonlinelibrary.com/jsfa c© 2010 Society of Chemical Industry J Sci Food Agric 2011; 91: 758–766

76

1


Figure 1. SEC and LLS chromatograms, and typical Berry plots,√

kc/Rθ versus sin2(θ/2), of KGM in 0.2 mol L−1 NaCl at 30 ◦C detected by 90◦ LS ( )and DRI (- - - - ).

Figure 2. (a) Molar mass versus elution volume and (b) RMS radius versuselution volume for KGM determined by HPSEC–MALLS-RI.

in a very small change in the RMS radius.20 Mw (weight-averagemolecular weight), Mn (number-average molecular weight) andMz (z-average molar mass) are 1.040 × 106, 1.023 × 106 and1.329 × 106 g mol−1, respectively. Weight-average mean squareradius (Rw), number-average mean square radius (Rn) and z-average mean square radius (Rz) are 115.9, 110.5 and 129.6 nm,respectively. The polydispersity index Mw/Mn is 1.017. Molecularcharacteristics of KGM obtained in this experiment are in goodagreement with what were reported in some other studies.21,22

Figure 3 shows the plot of RMS radius versus molar mass for KGM.The slope of such a plot indicates the conformation of KGM insolution. Wyatt23 stated that a slope of less than or equal to 0.33indicated a most compact spherical molecule, whereas for linearmolecules with random coil conformation the slope normallyranges between 0.5 and 0.6. From Fig. 3 we can see that the slopeof the plot for KGM is 0.52 ± 0.00, indicating that KGM is a linearmolecule with a random coil conformation in solution, which is inagreement with the findings of Li et al.11 on the molecular chainmorphology of KGM.

Intrinsic viscosity and molecular morphology of KGMFigure 4 shows Huggins and Kraemer plots (ηsp/C and ln ηr/Cversus C) for KGM in 0.2 mol L−1 NaCl aqueous solution. Intrinsicviscosity was obtained from the Huggins and Kraemer fittings.Huggins and Kraemer constants are related to the slope of thecorresponding plots and the intrinsic viscosity.24 From Fig. 4 it canbe seen that [η] is equal to 19.739 dL g−1, and that kH and kK areequal to 0.448 and 0.077, respectively.

Based on the respective Mw and [η] values of KGM-1, KGM-2,KGM-3, KGM-4 and KGM-5, a plot of log(η) against log Mw yielded


76

2


Figure 3. RMS radius versus molar mass for KGM determined by HPSEC–MALLS-RI; the slope of the plot is 0.52 ± 0.00.

Figure 4. Huggins and Kraemer plots for KGM in 0.2 mol L−1 NaCl aqueoussolution at 25 ◦C.

Figure 5. Relationship between molecular weight and intrinsic viscosityfor the Mark–Houwink equation.

a straight line (Fig. 5). According to the slope and intercept ofthe line, we can obtain K = 4.5394 × 10−2 and α = 0.7704.Consequently, the Mark–Houwink equation could be establishedas

[η] = 4.5394 × 10−2M0.7704w

The character constant α has an intimate relationship withthe rigidity degree and solvent ability of the molecular chain;meanwhile its value depends on the polymer solution systemproperties and polymer chain structure. The α value of KGMis 0.7704, which indicates that the KGM molecule is a kind ofsemi-flexible chain in aqueous solution.25

Glass transitions of the mixed amylose/amylopectin/KGMsystemsTgs of individual biopolymers

The Tg responses for amylose, amylopectin and KGM are shown inFig. 6 for different scan rates. It was observed that the detectionsensitivity of the Tg obtained from the peak of the derivative ofthe heat flow curve increased substantially as the heating ratewas increased. A clear Tg peak cannot be seen as the heating rateranged from 50 to 250 ◦C min−1. For a scan rate of 400 ◦C min−1 itwas very easy to see the detail of the Tg. Due to large distortionsin baseline at a heating rate up to 500 ◦C min−1, we performedthe Tg measurement using hyper-DSC with a heating rate of400 ◦C min−1.

The hyper-DSC curves for the individual biopolymer are shownin Fig. 7. The Tgs of amylose and amylopectin containing about 10%water were detected to be 85.40 ◦C and 84.19 ◦C, respectively. Bizotet al.26 reported that pure amylose exhibited a somewhat higherTg than branched amylopectin, with the explanation that linearchains appear to favor chain–chain interactions and induce partialcrystallinity.27 The α-(1 → 6) linkages theoretically offer threerotational degrees of freedom whereas α-(1–4) linkages offer twodegrees. Thus amylopectin with the higher degree of branchinghas greater chain flexibility than the relatively linear amylose. Onthe other hand, the Tg value for KGM containing about 10% waterwas 61.08 ◦C. It can be seen that the Tg of KGM is obviously lowerthan the Tgs of amylose and amylopectin. Bizot et al.26 illustratedthe influence of molecular weight, degree of branching, degree ofcrystallinity and (1–4) versus (l–6) glycosidic linkage ratio upon thedepression of Tg; that is, the higher the molecular weight and thelonger the chain length, the higher the Tg; the more branched andflexible, the lower the Tg; the more crystalline, the higher the Tg.The results in Figs 2–5 show that the molar masses of KGM rangedfrom 1.040 × 106 to 1.329 × 106 g mol−1 and it was a branchedand flexible macromolecule in solution, which indicates KGM witha higher degree of branching has greater chain flexibility thanthe relatively linear macromolecule. The X-ray diffractograms andcorresponding crystallinities of amylose, amylopectin and KGM aredepicted in Fig. 8 and Table 2, respectively. It is also seen that thecorresponding crystallinities of amylose, amylopectin and KGMwere 24.08%, 12.78% and 0.00%, respectively. Similar results havebeen reported elsewhere.28 Apparently, KGM has smaller degreesof crystallinity, and is more branched and flexible, comparedwith amylose and amylopectin. These physical properties of thespecimens are summarized in Table 2. The above results indicate


76

3


Figure 6. Normalized heat flow as a function of temperature at differentheating rates for (A) amylose, (B) amylopectin, and (C) KGM at about 10%moisture content.

the reason why the Tg of KGM is lower than the Tgs of amyloseand amylopectin, which suggests that KGM may decrease the Tgsof amylose and amylopectin if they are mixed.

Tgs of mixed amylose/amylopectin/KGM systemsHyper-DSC thermograms of mixed amylose/amylopectin/KGMsystems with low water content are shown in Figs 9 and 10.Generally, the Tg of the amorphous material produces a stepwisechange in the heat flow due to the change in heat capacity at thephase transition temperature. It can be seen that there are clearstep changes detected when the heating rate is at 400 ◦C min−1.In Fig. 9 it is observed that the addition of KGM affects the Tgsof potato amylose and amylopectin. As can be inferred fromthe above, the ratios of KGM to potato amylose or amylopectinhave great effects on the Tgs of the mixture system when theyincrease from 0 : 1 to 4 : 1. Figure 9 shows that the Tgs of themixed amylose/KGM and amylopectin/KGM systems are lowerthan that of amylose and amylopectin, respectively. Figure 10shows that the Tgs of the mixed amylose/amylopectin systems arelower than that of amylose, but higher than that of amylopectin.Some researchers have reported that the Tgs of amylose and

Figure 7. DSC thermograms of individual biopolymers containing about10% moisture under a heating rate of 400 ◦C min−1 for (a) amylose,(b) amylopectin, and (c) KGM.

Figure 8. X-ray diffraction diagrams of specimens: (a) KGM; (b) amy-lopectin; (c) amylose.

amylopectin at different ratios are lower than that of amylose, andhigher than that of amylopectin. This finding can be interpreted interms of internal plasticization by α-(l–6) branching.29 Moreover,the increase in KGM content caused a significant decrease in theTgs of amylose/KGM and amylopectin/KGM. The higher the KGMcontent, the lower are the Tgs of the mixtures. This indicates thatthe addition of KGM has plasticizing effects on potato amyloseand amylopectin. Figure 10 represents the effects of KGM on theTgs of mixtures with different amylose/amylopectin ratios (1 : 1,1 : 2, 1 : 3, and 1 : 4) at low moisture. As the KGM content increasesfrom top to bottom, the glass transition regions of the sampleshift to the left, i.e., the lower-temperature area. This indicatesthat the presence of KGM decreases the Tgs of the mixtures.Figure 10(A) shows that the Tgs of mixture systems containingamylose/amylopectin/KGM at various ratios from 1 : 1:0 to1 : 1:5decrease from 84.97 to 76.62 ◦C. Figure 10(B) shows the Tgs ofthe mixture systems containing amylose/amylopectin/KGM atvarious ratios from 1 : 2:0 to1 : 2:5 decrease from 84.42 to 72.50 ◦C.Figure 10(C) shows that the Tgs of mixture systems containingamylose/amylopectin/KGM at various ratios from 1 : 3:0 to1 : 3:5decrease from 84.27 to 71.97 ◦C. Finally, Fig. 10(D) shows that theTgs of mixture systems containing amylose/amylopectin/KGM atvarious ratios from 1 : 4:0 to1 : 4:5 decrease from 84.23 to 68.84 ◦C.From Figs 9 and 10 we can see that the general trend is that


76

4


Table 2. Physical properties of specimens

Biopolymer Mw (×106 g mol−1) Molecular morphology Glycosidic bonds Crystallinity (%)

Amylose (A0512) 0.150a Linear α-(1–4) 24.08 ± 0.02e

Amylopectin (A8515) 6.000a Branched α-(1–4), α-(1–6) 12.78 ± 0.01e

KGM (KJ-30) 1.040b Semi-flexiblec β-(1–4), β-(1–6)d 0.00 ± 0.01e

a Weight-average molecular weight provided by Sigma.b Weight-average molecular weight measured by HPSEC-MALLS-RI system.c Molecular morphology measured by viscosimetry.d From Katsuraya et al.13

e Degree of crystallinity measured by XRD. Data are expressed as mean ± standard deviation (n = 3).

Figure 9. DSC thermograms of the mixed systems containing about 10%moisture under a heating rate of 400 ◦C min−1 for (A) KGM/amylose and(B) KGM/amylopectin ratios of (a) 0 : 1, (b) 1 : 1, (c) 2 : 1, (d) 3 : 1, and (e) 4 : 1.Vertical lines indicate the position of the middle temperature for the glasstransition obtained from the first scan.

the Tgs of the mixed amylose/amylopectin systems decrease withincreasing concentration of KGM.

Tgs of amylose/KGM and amylopectin/KGM after storageTable 3 lists changes in the Tgs of amylose/KGM and amy-lopectin/KGM mixtures during reheating after storage at 4 ◦Cfor 5, 10, 15, 20, 25, and 30 days. The results demonstrate thatTgs of the mixtures decrease with increasing KGM concentrationand increase with increasing storage time. The higher the KGMconcentration, the smaller is the change in Tgs of the mixturesover the storage period. After 30 days of storage, Tgs decreasedby 13.93 ◦C and 11.43 ◦C for amylose/KGM and amylopectin/KGMmixtures with polysaccharide concentrations of up to 4 g kg−1,respectively. These results indicate that KGM can obviously re-duce recrystallization of amylose and amylopectin. Is the effecton amylose and amylopectin aggregation or recrystallization dueto the potential amylose/KGM or amylopectin/KGM complex for-mation, or not? It should be emphasized that the complex might

be formed on a molecular scale when this aggregation involveshundreds or thousands of molecules.30 Therefore amylose/KGMand amylopectin/KGM mixtures were possibly produced and thispotential mixture lowered the rearrangement ability of amyloseand amylopectin, which may cause the decrease in physical agingof amylose and amylopectin.

The reduction in Tg suggests there to be a reduction in densityand an increase in free volume. This is to be expected on thebasis of the free volume theory, which states that the macroscopicthermal and mechanical properties of amorphous polymers are afunction of the unoccupied space, i.e., the free volume availablefor molecular motion.31 The idea that molecular motion in liquidsand solids requires molecule-sized holes was developed by Eyringet al.32 These holes, which collectively make up the free volume,are constantly moving in the liquid state. Doolittle33 related theviscosity of a liquid to its relative free volume. In the context ofpolymers, a hole, or free volume element, may be considered to besimilar in size to a segment of a polymer molecule and more thanone may be required for mobility (i.e., motion in polymers involvescooperative movement of portions of a polymer chain). Fox andFlory34 suggested that the glass transition, Tg, represents an iso-free volume state, occurring at a critical fractional free volumefor large-scale movement of polymer segments. The fractionalfree volume, fV, in the glassy state may be calculated as 0.025(i.e., 2.5% free volume) on the basis of the Williams–Landel–Ferry(WLF) equation,35 which describes the temperature dependenceof viscosity and relaxation times for amorphous polymers, andwhich may be derived on the basis of the Doolittle relationship.A substantially larger fV at Tg may be inferred from Simha andBoyer’s relationship, (αR − αg)Tg = 0.113, where αR and αg are thecoefficients of thermal expansion in the rubbery and glassy states,respectively.36 Other attempts to quantify free volume give furtherdifferent values, depending on the assumptions made about whatconstitutes free volume.

The effect of branched polysaccharide as a Tg depressor is knownand is often described in terms of internal plasticization.37 It wasreported that some polysaccharides (pullulan, chitosan, cellulosederivatives, carrageenans) could decrease Tgs of starches and thehardness of the paste.26,38,39 Bizot et al.26 have found that the Tg

of pullulan is lower than that of starch polymers and have alsointerpreted this phenomenon in terms of internal plasticizationby α-(1–6) branching. BeMiller and Whistler40 indicated that theeffective volume of the polysaccharide in solution is considerablygreater than the volume occupied by the atoms comprising thepolysaccharide chain because it sweeps out a large volume ofsolvent as it undergoes Brownian motion. Huang et al.41 reportedthat bulky polysaccharides contribute to a free volume increasefor starch chains, thus reducing the glass transition temperature.


76

5


Figure 10. DSC thermograms of the mixed systems containing about 10% moisture under a heating rate of 400 ◦C min−1 for amylose/amylopectin/KGMratios of (A) (a) 1 : 1:0, (b) 1 : 1:1, (c) 1 : 1:2, (d) 1 : 1:3, (e) 1 : 1:4, (f) 1 : 1:5; (B) (a) 1 : 2:0, (b) 1 : 2:1, (c) 1 : 2:2, (d) 1 : 2:3, (e) 1 : 2:4, (f) 1 : 2:5; (C) (a) 1 : 3:0, (b) 1 : 3:1,(c) 1 : 3:2, (d) 1 : 3:3, (e) 1 : 3:4, (f) 1 : 3:5; and (D) (a) 1 : 4:0, (b) 1 : 4:1, (c) 1 : 4:2, (d) 1 : 4:3, (e) 1 : 4:4, (f) 1 : 4:5. Vertical lines indicate the position of the middletemperature for the glass transition obtained from the first scan.

Table 3. Glass transition temperature of amylose/KGM and amylopectin/KGM after storage

Tg (◦C)

Sample code 5 days 10 days 15 days 20 days 25 days 30 days

2 (control) 86.93 ± 0.10 88.04 ± 0.02 89.64 ± 0.02 90.27 ± 0.03 93.11 ± 0.01 94.65 ± 0.05

3 84.06 ± 0.03 86.84 ± 0.01 88.12 ± 0.01 89.72 ± 0.02 91.02 ± 0.01 92.73 ± 0.02

4 82.36 ± 0.01 83.21 ± 0.03 84.44 ± 0.01 84.73 ± 0.02 85.39 ± 0.01 85.52 ± 0.03

5 79.35 ± 0.02 80.82 ± 0.01 81.68 ± 0.02 81.87 ± 0.02 82.23 ± 0.15 82.58 ± 0.01

6 77.12 ± 0.02 78.09 ± 0.02 79.16 ± 0.03 79.91 ± 0.01 80.02 ± 0.12 80.72 ± 0.05

7 (control) 85.53 ± 0.14 86.79 ± 0.02 86.84 ± 0.22 87.31 ± 0.04 88.08 ± 0.32 88.69 ± 0.20

8 83.87 ± 0.06 84.28 ± 0.05 85.07 ± 0.10 85.95 ± 0.01 86.75 ± 0.02 87.02 ± 0.01

9 79.93 ± 0.02 80.25 ± 0.01 81.54 ± 0.03 82.14 ± 0.01 82.48 ± 0.02 82.96 ± 0.10

10 77.87 ± 0.01 78.13 ± 0.10 78.76 ± 0.05 79.07 ± 0.01 79.74 ± 0.02 79.91 ± 0.04

11 75.56 ± 0.01 75.88 ± 0.01 76.15 ± 0.01 76.82 ± 0.01 76.97 ± 0.02 77.26 ± 0.01

Data are expressed as mean ± standard deviation (n = 3).

The mixed amylose/amylopectin/KGM system involves not onlyα-(1–6) linkages and α-(1–4) linkages but also β-(1–4) and β-(1–6) linkages. Studies on the molecular morphology of KGMhave shown that it exists as linear molecules with random coilconformation and semi-flexible chain in aqueous solution. Intheory, KGM can offer high rotational degrees of freedom of themixed amylose/amylopectin system, thus decreasing the Tg of themixed amylose/amylopectin system.

CONCLUSIONSKGM, a kind of natural polysaccharide, a has random coilconformation and semi-flexible chain in aqueous solution, and

its crystallinity is 0.00%. Therefore, with regard to the KGMproperties obtained above, these results lead to the conclusionsthat KGM can increase plasticization and molecular movementof the potato amylose and amylopectin chains, and decrease therelation between the macromolecules, thereby decreasing the Tgsand recrystallization of the mixed amylose/amylopectin system.

ACKNOWLEDGEMENTSThis research has been supported by the National Natural ScienceFoundation of China (Grant No. 20776002) and the hi-techresearch and development program of China (863 program)(Grant No. 2006AA10Z340). We thank Key Laboratory of Cigarette


76

6


Smoke, Technology Center of Shanghai Tobacco (Group) Corp. forinstrument support.

REFERENCES1 Smith F and Srivastava HC, Constitution studies on the glucomannan

of konjac flower. J Am Chem Soc 81:1715–1718 (1959).2 Katsuraya K, Okuyama K, Hatanaka K, Oshima R, Sato T and

Matsuzaki K, Constitution of konjac glucomannan: chemical analysisand 13C NMR spectroscopy. Carbohydr Polym 53:183–189 (2003).

3 Kato K and Matsuda K, Studies of the chemical structure of konjacmannan. I. Isolation and characterization of oligosaccharidesfrom the partial hydrolyzate of the mannan. Agric Biol Chem33:1446–1453 (1969).

4 Maekaji K, Relationship between stress relaxation and syneresis ofkonjac mannan gel. Agric Biol Chem 42:177–178 (1978).

5 Levrat VMA, Behr S, Mustad V, Remesy C and Demigne C, Low levelsof viscous hydrocolloids lower plasma cholesterol in rats primarilyby impairing cholesterol absorption. J Nutr 130:243–248 (2000).

6 Dave V, Sheth M, McCarthy SP, Ratto JA and Kaplan DL, Crystalline,rheological and thermal properties of konjac glucomannan. Polymer39:1139–1148 (1998).

7 Xiao C, Gao S, Wang H and Zhang L, Blend films from chitosan andkonjac glucomannan solutions. J Appl Polym Sci 76:509–515 (2000).

8 Cheng LH, Karim AA, Norziah MH and Seow CC, Modification of themicrostructure and physical properties of konjac glucomannan-based films by alkali and sodium carboxymethylcellulose. Food ResInt 358:828–836 (2002).

9 Chen JG, Liu CH, Chen YQ, Chen Y and Chang PR, Structural char-acterization and properties of starch/konjac glucomannan blendfilms. Carbohydr Polym 74:946–952 (2008).

10 Nishinari K, Konjac glucomannan. Dev Food Sci 41:309–330 (2000).11 Li B, Xie BJ and Kennedy JF, Studies on the molecular chain mor-

phology of konjac glucomannan. Carbohydr Polym 64:510–515(2006).

12 Yang G, Zhang L, Yamane C, Miyamoto I, Inamoto M andOkajima K, Blend membranes from cellulose/konjac glucomannancuprammonium solution. J Memb Sci 139:47–56 (1998).

13 Katsuraya K, Okuyama K, Hatanaka K, Oshima R, Sato T andMatsuzaki K, Constitution of konjac glucomannan: chemical analysisand 13C NMR spectroscopy. Carbohydr Polym 53:183–189 (2003).

14 Liu P, Yu L, Liu HS, Chen L and Li L, Glass transition temperature ofstarch studied by a high-speed DSC. Carbohydr Polym 77:250–253(2009).

15 McGregor C and Bines E, The use of high-speed differential scanningcalorimetry (hyper-DSCTM) in the study of pharmaceuticalpolymorphs. Int J Pharm 350:48–52 (2008).

16 Saunders M, Podluii K, Shergill S, Buckton G and Royall P, The potentialof high speed DSC (hyper-DSC) for the detection and quantificationof small amounts of amorphous content in predominantlycrystalline samples. Int J Pharm 274:35–40 (2004).

17 Yoshimura M, Takaya T and Nishinari K, Rheological studies onmixtures of corn starch and konjac-glucomannan. Carbohydr Polym35:71–79 (1998).

18 ASTRA for Windows User’s Guide for the DAWN EOS, mini DAWN, andWyatt Light Scattering Instruments, Version 5.1. Wyatt Technology,Santa Barbara, CA, pp. 56–78 (2004).

19 Hermans PH and Weidinger A, Quantitative x-ray investigations onthe crystallinity of cellulose fibers, a background analysis. J ApplPhys 19:491–506 (1948).

20 Tao YZ and Zhang LN, Determination of molecular size andshape of hyperbranched polysaccharide in solution. Biopolymers83:414–423 (2006).

21 Kohyama K, Sano Y and Nishinari K, A mixed system composedof different molecular weights konjac glucomannan andκ-carrageenan. II. Molecular weight dependence of viscoelasticityand thermal properties. Food Hydrocoll 10:229–238 (1996).

22 Sugiyama N, Shimahara H, Andoh T, Takemoto M and Kamata T,Molecular weight of konjac mannans of various sources. AgricBiol Chem 36:1381–1387 (1972).

23 Wyatt PJ, Light-scattering and the absolute characterization ofmacromolecules. Anal Chem Acta 272:1–40 (1993).

24 Wanchoo RK and Sharma SK, Viscometric study on the compatibilityof some water-soluble polymer–polymer mixtures. Eur Polym J39:1481–1490 (2003).

25 Zhu SN, Polymer Chain Structure. Science Press, Beijing, pp. 123–132(1996).

26 Bizot H, Bail PL, Leroux B, Davy J, Roger P and Buleon A, Calorimetricevaluation of the glass transition in hydrated, linear and branchedpolyanhydroglucose compounds. Carbohydr Polym 32:33–50(1997).

27 Liu P, Yu L and Wang XY, Glass transition temperature of starcheswith different amylose/amylopectin ratios. J Cereal Sci 51:388–391(2010).

28 Myllarinen P, Buleon A, Lahtinen R and Forssell P, The crystallinity ofamylose and amylopectin films. Carbohydr Polym 48:41–48 (2002).

29 Slade L and Levine H, Beyond water activity: recent advances basedon an alternative approach to the assessment of food quality andsafety. CRC Crit Rev Food Sci Nutr 30:115–360 (1991).

30 Richardson G, Kidman S, Langton M and Hermansson AM, Differencesin amylose aggregation and starch gel formation with emulsifiers.Carbohydr Polym 58:7–13 (2004).

31 Algers J, Suzuki R, Ohdaira T and Maurer FHJ, Characterization of freevolume and density gradients of polystyrene surfaces by low-energypositron lifetime measurements. Polymer 45:4533–4539 (2004).

32 Eyring H, Viscosity, plasticity and diffusion as examples of absolutereaction rates. J Chem Phys 4:283–291 (1936).

33 Doolittle AK, Studies in newtonian flow. II. The dependence of theviscosity of liquids on free-space. J Appl Phys 22:1471–1475 (1951).

34 Fox JTG and Flory PJ, Second-order transition temperatures andrelated properties of polystyrene. I. Influence of molecular weight.J Appl Phys 21:581–591 (1950).

35 Williams ML and Landel RF, The temperature dependence ofrelaxation mechanisms in amorphous polymers and other glass-forming liquids. J Am Chem Soc 77:3701–3707 (1955).

36 Simha R and Boyer RF, On a general relation involving the glasstemperature and the coefficients of expansion of polymers. J ChemPhys 37:1003–1007 (1962).

37 Zimeri JE and Kokini JL, Phase transitions of inulin–waxy maizestarch systems in limited moisture environments. Carbohydr Polym51:183–190 (2003).

38 Athina L and Costas GB, Thermophysical properties of chitosan,chitosan–starch and chitosan–pullulan films near the glasstransition. Carbohydr Polym 48:179–190 (2002).

39 Jetnapa T, Manop S and James NB, Effects of cellulose derivatives andcarrageenans on the pasting, paste, and gel properties of ricestarches. Carbohydr Polym 73:417–426 (2008).

40 BeMiller JN and Whistler RL, Carbohydrates, in Food Chemistry, ed. byFennema OR. Marcel Dekker, New York, pp. 157–159 (1997).

41 Huang RM, Chang WH, Chang YH and Lii CY, Phase transition of ricestarch and flour gels. Cereal Chem 66:411–415 (1994).


Documents

Effects of molecular characteristics of on konjac glucomannan glass transitions of potato amylose, amylopection and their mixtures