Electromyography analysis of natural mastication behavior using varying mouthful quantities of two types of gels

Physiology & Behavior 161 (2016) 174–182

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Electromyography analysis of natural mastication behavior using varying mouthful quantities of two types of gels Kaoru Kohyama a,⁎, Zhihong Gao a, Sayaka Ishihara b, Takahiro Funami b, Katsuyoshi Nishinari c a b c

Food Research Institute, NARO, 2-1-12 Kannondai, Tsukuba, Ibaraki 305-8642, Japan San-Ei Gen F. F. I., Inc., 1-1-11 Sanwa-cho, Toyonaka, Osaka, 561-8588, Japan School of Food and Pharmaceutical Engineering, Hubei University of Technology, Wuchang, Wuhan 430068, China

H I G H L I G H T S

G R A P H I C A L

A B S T R A C T

• Natural mastication behavior of gel foods was measured by electromyography. • Two-type gels with similar fracture loads and masses showed similar chewing times. • Smaller mass and more-elastic type of gels were chewed more with a single side. • Contribution of both sides is more effective in consuming a portion of gels. • Smaller mouthfuls induced slower eating with more chews per portion of both types.

a r t i c l e

i n f o

Article history: Received 21 January 2016 Received in revised form 28 March 2016 Accepted 13 April 2016 Available online 19 April 2016 Keywords: Mouthful mass of gel Gel texture Natural mastication Electromyography Masseter muscles Dominant side of chewing

⁎ Corresponding author. E-mail address: [email protected] (K. Kohyama).

http://dx.doi.org/10.1016/j.physbeh.2016.04.030 0031-9384/© 2016 Elsevier Inc. All rights reserved.

a b s t r a c t The objectives of this study were to examine the effects of mouthful quantities and mechanical properties of gels on natural mastication behaviors using electromyography (EMG). Two types of hydrocolloid gels (A and K) with similar fracture loads but different moduli and fracture strains were served to eleven normal women in 3-, 6-, 12-, and 24-g masses in a randomized order. EMG activities from both masseter muscles were recorded during natural mastication. Because of the similar fracture loads, the numbers of chews, total muscle activities, and entire oral processing times were similar for similar masses of both gel types. Prior to the first swallow, the more elastic K gel with a higher fracture strain required higher muscle activities than the brittle A gel, which had higher modulus. Majority of subjects had preferred sides of chewing, but all subjects with or without preferred sides used both masseters during the consumption of gels. Similar effects of masses and types of gels were observed in EMG activities of both sides of masseters. Contributions of the dominant side of chewing were diminished with increasing masses of gels, and the mass dependency on ratio of the dominant side was more pronounced with K gel. More repetitions of smaller masses required greater muscle activities and longer periods for the consumption of 24-g gel portions. Reduction in the masses with an increased number of repetitions necessitated slower eating and more mastication to consume the gel portions. These observations suggest that chewing using both sides is more effective and unconsciously reduces mastication times during the consumption of gels. © 2016 Elsevier Inc. All rights reserved.

K. Kohyama et al. / Physiology & Behavior 161 (2016) 174–182

1. Introduction Food texture plays an important role in food palatability and may thus modify natural eating behaviors [1,2]. Texture is defined as the mechanical, geometrical, and surface properties that are perceptible using human receptors [1], and food textures perceived during oral processing are greatly influenced by the mechanical properties and sizes of the food items [1–3]. Electromyography (EMG) of masticatory muscles has been widely used to quantify mastication behaviors [2,4–7]. Harder foods generally require greater EMG activities of the jaw-closing muscles, and longer mastication times with a greater number of chews have been reported in association with these [4–9]. Moreover, previous EMG studies examining the quantity of each mouthful showed that larger masses or volumes of food required longer oral processing times [4, 10–18]. Forde et al. [19] observed free-eating behaviors and reported that softer foods were generally consumed with larger bites (mouthful masses), leading to faster eating rates. Moreover, smaller bites of semi-solid foods increased oral processing times and decreased food intake [20], whereas harder foods resulted in reduced energy intake that could be sustained till the next meal [21]. Other studies that examined young subjects who ate quickly showed a significantly lower number of chews, shorter durations of chewing, and greater bite size were seen to be associated with a smaller number of bites in subjects who ate rice balls [22]. Finally, a meta-analysis showed that slow eating rates were associated with reduced energy intake [23]. In preceding studies [18,24], EMGs were recorded during natural chewing of model food gels. These experiments showed that mastication times, number of chews, and number of swallows required for gels with various textures were approximately proportional to 0.7 times to the power of gel masses ranging from 3 to 24 g, and this relationship could be used to make predictions. Studies using fixed masses of foods with different shapes reported that the geometrical characteristics of gels did not influence eating behaviors [14]. Specifically, blocks, small pieces, and thin slices of apples with a fixed mass did not showed significant differences in EMG variables during natural mastication [25]. Forced mastication with a fixed number of chews has also been tested by multiple researchers. In general, more chews per mouthful of food were seen to be associated with reduced eating rates [26–29]. Recently, Smit et al. [27] confirmed “Fletcherism,” an idea initially propagated by Fletcher which suggests that chewing one mouthful of food many times reduces the food intake and the eating rate despite faster chewing cycles. Increased number of chews also reduced the energy intake of one meal and modulated the plasma gut hormones, insulin and glucose [26,28]. Slow eating with a greater number of chews reportedly increased diet-induced thermogenesis, postprandial blood flow [29], and postprandial glucose responses in glycemic indices measurements [30,31]. However, if fixed modes of chewing were imposed, subjects could control chewing forces and rates of eating that were unnatural. Eating small masses of gel-type foods appeared to be effective in preventing overeating, which often leads to obesity and other health problems [7]. Moreover, to avoid risk of suffocation in children, preparation of gels in cup volumes greater than the size of their mouth [32] and serving in small spoonful masses were recommended. The objectives of this study were to quantify the effects of gel masses on natural mastication behaviors using portions of gel foods. We hypothesized that smaller mouthful masses and greater number of the mouthfuls increase mastication efforts and reduce rates of eating fixed amounts of food portions. In addition, the chewing side may change more frequently with food items that are more difficult to masticate. The subjects performed natural mastication without being instructed to control chewing forces so as to allow investigation of habitual eating behaviors instead of under controlled conditions such as fixed number of chews and side of chewing.

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Two types of gel foods with similar fracture loads (agar gel and konjac jelly) were prepared as the present food samples. Previous studies have reported that the number of chews and mastication times were alike for the gels with similar masses [18,33]. Accordingly, hydrocolloid gels provide models of solid or semi-solid foods consumed by mastication using the back teeth as the mechanical properties and sizes of these gels can be easily controlled [34–37]. Moreover, gel alternatives for food items such as cooked rice are extensively consumed as staple foods, and jellies are often served to dysphagic patients and elderly people with mastication difficulties [7,35,38,39]. The chemical components of these hydrogels do not change during oral processing, as salivary enzymes do not decompose non-starch ingredients. In addition, these hydrogels contain sufficient water to minimize the amount of saliva required for bolus formation and lubrication. Hence, size reductions with respect to structural threshold [40] are the main requirement for oral processing, and reduction of mouthful masses is easy and does not require changes in the method of food preparation. Finally, EMG recordings provide quantitative evidence of the effects of serving sizes, and also allow simultaneous evaluation of how the mechanical properties of foods influence mastication efforts and rates. 2. Materials and methods 2.1. Participants This study design was approved by the National Food Research Institute Ethics Committee. Eleven women volunteers (mean age, 36.5; range, 22–49 years) participated in this study were common in our previous study [18]. Women were selected as there may be gender differences in mastication behavior [18]. Their heights and body mass indices ranged from 150 to 170 cm and 18 to 23 kg/m2, respectively. The subjects were free of any masticatory or swallowing dysfunctions and did not use removable denture prostheses. Moreover, they were all right-handed. Written informed consent was collected before EMG recordings, and the subjects were asked to eat sample gels as they normally would to allow assessment of habitual eating behavior. 2.2. Sample gels Two types of hydrocolloid gels were prepared in cups (diameter 60 mm and height 25 mm) as described in the preceding study [18]. All ingredients were food grade and provided by San-Ei Gen F. F. I., Inc. (Osaka, Japan). Type one (A) gel contained a 1.2% w/w agar (GEL UP® J-3531), while type two (K) gel was a mixture of 0.26% w/w konjac mannan (VIS TOP® D-2134), 0.46% w/w κ-carrageenan (CARRAGEENAN CS606), and 0.46% w/w locust bean gum (VIS TOP® D-2050). Both gels contained 25% w/w sucrose, 0.15% w/w sodium citrate, and 0.22% w/w anhydrous citric acid. The fracture load of both types of gels was similar, the elastic modulus was about 30-fold higher in A gel, fracture strain and work were much higher in K gel (Table 1), as reported previously [18]. The load at 90% strain and work until 90% strain in A gel have not been reported previously and are significantly greater than those in K gel (Table 1). 2.3. EMG measurements The EMG activities were recorded from the left and right masseter muscles (LM and RM, respectively) using bipolar surface electrodes (EL503, Biopac Systems Inc., Goleta, CA, USA) [17,18,24]. Briefly, the skin was wiped with cotton soaked in 70% ethanol, masseter electrodes were carefully placed along the muscle fibers on both sides symmetrically and approximately 2 cm apart, and a ground electrode was placed on the left wrist. Subjects were asked to clench their jaw, tap their teeth several times, and swallow a small amount of water to check conditions prior to testing the gels. The electrodes were replaced if unstable baseline signals and/or significant noises were detected, or if the magnitudes

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Table 1 Mechanical properties of sample gels measured by a penetration test. Type of gel

Fracture load (N)

Apparent elastic modulus (kPa)

Fracture strain (%)

Fracture work (N.mm)

Load at 90% strain (N)

Work until 90% strain (N.mm)

A-gel K-gel

0.52 ± 0.02 0.51 ± 0.03

980 ± 70 35 ± 6

6.5 ± 0.3 51.3 ± 1.5

0.34 ± 0.02 2.27 ± 0.17

0.46 ± 0.03 0.36 ± 0.04

8.66 ± 0.38 5.40 ± 0.32

Means and standard deviations from 6 replicates. A (agar) or K (mixed gels of konjac mannan, κ-carrageenan, and locust bean gum) gels. Both gels contained 25% w/w sucrose, 0.15% w/w sodium citrate, and 0.22% w/w anhydrous citric acid. Gel samples in cups were compressed at 10 mm/s using a cylindrical probe (3-mm diameter) at 20 °C. Fracture points were determined at the first reduction of load. Strain was calculated as the ratio of deformation to the initial gel thickness. Stress was calculated under small deformations as the load divided by the cross sectional area of the probe. Apparent elastic moduli were determined from the initial slope of the stress-strain curve. Work was calculated as the area under the load–distance curve.

of both masseter EMGs were unbalanced during tapping the electrodes were replaced. The peak to baseline EMG ratios were N30 times those before the first swallow, and the differences in mean peak amplitudes between LM and RM were less than four times. A switch (TSD116A, Biopac Systems Inc.) was then handed to the subject and she was asked to eat the sample gels as she would habitually, and the natural chewing behaviors were recorded. Subjects were also instructed to press the button with the right thumb to indicate the start of chewing, and to indicate every swallow. The end of eating was indicated by longer signals. The EMG signals were then filtered (10–500 Hz), the 50 Hz noise was removed, amplified 1000 times, digitized, and saved in a PC using an MP-150 system (Biopac Systems Inc.) with three EMG 100C amplifiers at 1000 Hz. An additional digital signal from the switch was stored using an analogue-digital converter (UIM100C, Biopac Systems Inc.). A typical EMG is shown in Fig. 1. To study the effects of food mass, the two types of gels were randomly served at least twice to subjects as 3-, 6-, 12-, and 24-g specimens using a plastic spoon. The 3-, 6-, and 12-g gels were cut into cubes, while the 24-g specimens were cut into cuboids with a height of 25 mm. They were prepared based on their weight, and their approximate sizes were 15 × 15 × 15 mm, 19 × 19 × 19 mm, 24 × 24 × 24 mm, and 33 × 33 × 25 mm, respectively. The height of the K-gel specimens was shorter than that of A gels with similar mass due to high deformability of the former [33]. The first side of chewing was observed by the experimenter who served the gels. The subjects were allowed to talk, rinse their mouths, and drink water freely between each trial. EMG recordings were taken for 30 min and the sum of chewing periods analyzed was approximately 10 min for each subject. The entire session, including delivery of instructions, setting, and removal of EMG electrodes was performed in b 60 min [18].

2.4. Data analyses The stored data were analyzed using AcqKnowlegde® ver. 4.1.1 software (Biopac Systems Inc.). Switch signals were used as a guide for the period and to count the number of swallows, and precisely-determined periods and time points were based on EMGs. The time for oral processing was defined as the period elapsed between appearance of the first EMG activity close to the start signal and end of EMG activity close to the end signal (top arrows in Fig. 1). The entire time for oral processing was divided by the first swallowing signal into two stages (T1 and T2), as described in previous papers [18,41]. The initial quantities of gels were masticated in the mouth during the first stage (T1), which extended from the start to the first swallow, and an unknown smaller amount was processed thereafter (T2). The number of chews was counted from the EMGs. As is shown in a magnified view of one chew (Fig. 2), peak-to-peak amplitude, burst duration, cycle time, and muscle activity (time-integrated EMGs) were read per chew for LM and RM [7,17,18,24]. The four EMG parameters of each chew were then averaged for T1 and T2. EMG duration and muscle activities were summed up for each period and for the entire time for oral processing. The dominant side of chewing (DS) was defined as the side which had the greater mean peak-topeak amplitude during T1. Statistical analyses were conducted using the SPSS® package (ver. 17.0 J for Windows®) (SPSS Inc., Chicago, IL, USA), and level of significance was set at p b 0.05. EMG variables were averaged over two trials, and comparisons were made using two-way repeated-measures analysis of variance (ANOVA). EMG variables within a subject were compared, and the types (2) and masses (4) of gels were repeated factors. The presence of preferred side of chewing was determined using binominal tests (two-sided).

Table 2 Numbers of sides of chewing in 16 trials and preferred sides of each subject. Subject ID

Fig. 1. Example of electromyograms during free eating. Sample is 3-g K gel. From the top; electromyogram signals of right masseter (RM), left masseter (LM), and output of button switch. The subject (S8 in Table 2) preferred the left side. She pushed a button to indicate the start of the oral processing and every swallow (two swallows in this case). Time for oral processing is shown at the top. Stages T1 (from the start to the first swallow) and T2 (the following period until the end) were separately analyzed.

S1 S2 S3 S4 S5 S6 S7 S8 S9 S10 S11

First chew

Greater mean amplitudes

Greater total muscle activities

Left–Right

Left–Right

Left–Right

0–16 0–16 4–12 8–8 10–6 11–5 12–4 14–2 15–1 16–0 16–0

0–16 0–16 5–11 15–1 9–7 9–7 15–1 15–1 16–0 16–0 16–0

0–16 0–16 6–10 9–7 10–6 9–7 14–2 15–1 16–0 14–2 16–0

Preferred side of chewing

Right Right ns ns ns ns Left Left Left Left Left

ns, not significant, determined by two-sided binomial test. Each subject consumed 16 gels (3, 6, 12, and 24 g of A and K gels, two replicates of each gel type) using habitual modes of mastication. Mean amplitudes and total muscle activities were calculated for the stage T1 before the first swallow.

K. Kohyama et al. / Physiology & Behavior 161 (2016) 174–182

Fig. 2. Example of variables from raw electromyograms per chew.

3. Results 3.1. Dominant and non-dominant sides of masticatory EMGs Both masseter EMGs were read and the variables were calculated separately. The results showed that the side of the first chew appeared to be fixed for many subjects, as shown in Table 2. Subjects S1 and S2 started chewing on the right side, and the mean amplitudes and total muscle activities calculated for stage T1 were greater on the right side for all samples (Fig. 3). Subject S2 preferred this side over the left for the entire time for oral processing. During stage T1, the LM (non-dominant side NDS) to RM (DS) ratios of amplitude and muscle activity were 0.638 and 0.612, respectively. Conversely, subjects S7–S11 (which includes a subject S8 shown in Fig. 1) started chewing on the left side and masticated samples more with this side. Owing to the small gel size (3 g), subject S8 mainly chewed with the left side for all nine cycles in T1, and the RM (NDS) to LM (DS) ratios of amplitude and muscle activity were 0.678 and 0.649, respectively (Fig. 1). Consequently, the mean EMG amplitudes and total muscle activities of these subjects (S1, S2, and S7–S11) were significantly greater in DS, which was defined as the preferred side of chewing (Table 2). Some subjects exhibited similar EMG amplitudes in both masseters and did not have a preferred side for the first chew. Of 11 subjects, four (S3–S6) did not show any significant preferred side (p N 0.05, determined by binominal test, Table 2). This group included cases with similar amplitudes and muscle activities on both sides with every chew, and the right–left ratios for each chew were 0.9–1.1. In contrast, other subjects of this group exhibited significantly greater EMG amplitude on the chewing side than the opposite side, but used both sides

Fig. 3. Example electromyograms from a subject with a preferred side of chewing. Subject 2 preferring the right side (S2 in Table 2) consumed a 24-g K gel. Signals from the three channels, T1 and T2 are the same as that in Fig. 1. Gray arrowheads present swallowing activities excluded from the mastication analysis.

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equally (Fig. 4). Hence, the corresponding right–left ratios of amplitudes and muscle activities for each chew were sometimes b0.2 or were N3, and the mean values for stage T1 were 0.957 and 0.908, respectively. In subject S5, the side of chewing for each cycle was clearly identified by greater amplitudes in the first 9 chews on the left, followed by 14 chews on the right, and then 6 chews on the left before the first swallow (Fig. 4). The chew indicated by the gray double-headed arrow in Fig. 4 exhibits similar RM and LM amplitudes, suggesting that the chewing cycle was performed using both sides simultaneously (bilateral chewing). Some subjects (S2) exhibited great masseter activities during swallowing (Fig. 3), while others (S5 and S8) did not (Figs. 4 and 1). EMGs were used to confirm the pattern in each subject while swallowing water. Fig. 3 shows that EMG activities while swallowing did not differ greatly between the two masseters, and burst durations were longer than rhythmical chewing times. Three swallowing bursts (arrowheads in Fig. 3) were not included in the assessment of number of chews and chewing EMG activities. DS was defined as the side with greater mean amplitude for each EMG while chewing a gel specimen, and it was more apparent during T1 as the EMG activities of masseter muscles decreased during T2. DS was the same as the preferred side of chewing in some subjects (left in Fig. 1 and right in Fig. 3), but varied in others who did not have a preferred side of chewing (Fig. 4), as indicated by small differences in the amplitudes of left and right sides. Accordingly, the left side was identified as dominant in the example shown in Fig. 4, as the mean amplitudes were 1.03 mV for the right and 1.09 mV for the left side. The differences between both mean amplitudes were slight, and the DS of this subject was left, 9 times out of 16 and right, 7 times out of 16. Therefore, it could be concluded that this subject did not have a preferred side of chewing. The average ratio of masseter muscle activity of NDS to DS was calculated for stage T1 in all subjects, regardless of the presence of a preferred side of chewing. Subjects without a preferred side (S5) exhibited a ratio of approximately one, while those with a preferred side showed smaller values. NDS contributed to natural chewing of gels in all cases, and the minimum NDS/DS ratio of amplitudes was 0.259 (6-g A gel by S10, the NDS/DS ratio of muscle activity was 0.529). Moreover, NDS/DS ratios of natural chewing were not considerably small to ignore the NDS and depended on masses and types of gels. Left–right averaged EMGs reported previously [18] were more indicative than single-side EMGs. 3.2. Effects of mass and type of gels on EMG variables Table 3 shows the EMG results for four different masses of the two types of gels, compared using two-way repeated-measure ANOVA.

Fig. 4. Example electromyograms from a subject without a preferred side of chewing. Subject 5 (S5 in Table 2) consumed a 24-g K gel. Signals recorded from three channels, T1 and T2 are the same as that in Fig. 1. Gray single-headed arrows between RM and LM channels indicate side changes of chews from left to right and right to left, respectively. The cycle with the gray double-headed arrow indicates bilateral chewing.

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The EMG variables were separately derived before and after the first swallow and for the entire period of oral processing (time for oral processing). Amplitudes and muscle activities from the DS were greater than those from the NDS for any masses and type at any stage of oral processing. Moreover, values were greater for the K gel before the first swallow (T1), and increased with masses of gels in both DS and NDS (Table 3). The effects of sample masses on amplitude and muscle activity were less significant in DS than in NDS. Gel type had no significant effect (Table 3) after the first swallow up to the end of oral processing (in T2). It is possible that a most of the gels were swallowed the first time and only a small amount of gel remained in the mouth. A significant increase in the amplitudes of DS and NDS, but not in the muscle activities per chew, was observed with an increase in gel mass. Active durations of masseter muscles were similar for both sides (Figs. 1, 3, and 4), but the amplitudes were greater on the side of chewing. Thus, differences in amplitude and muscle activity were observed between the sides. The absolute values of amplitude and muscle activity were greater in the DS, but the effects of the gel types and masses were similar for both sides. Chewing cycles and duration per chew did not significantly differ between the sides. A longer time was also required for larger masses of gels from the first swallow to the last swallow (T2), and the total muscle activities in T2 increased with masses for both types of gels (Table 3). The total muscle activities, times for oral processing, and total number of swallows and chews increased with increasing masses of both types of gels [18]. Moreover, these did not differ between similar masses of gels, except that the number of swallows was significantly higher for A gels [18].

3.3. Effects of a mouthful mass per cup of gel Table 3 shows that many EMG parameters increased with the mass of gel. However, the effects of gel mass on EMG variables were stronger than those of gel type, and portions of gels, (e.g., cup of jelly) were not generally consumed in one mouthful. It was assumed that a portion of minicup jellies (24 g) was eaten in varying masses (3, 6, 12, and 24 g), and the mouthful mass remained fixed when consuming the gel portion. Thus, eight mouthfuls of 3-g, four mouthfuls of 6-g, two mouthfuls of 12-g, and one mouthful of 24-g gels were compared. Table 4 is a comparison of EMG variables per 24-g portions recalculated in this manner. Mass was found to have a significant effect (Table 4). The smaller mass of gel and increased times of the masses resulted in greater EMG variables. Gel type had a significant effect on the sum of muscle activities and number of chews during T1 (A b K gels), and number of swallows (A N K gels), although mastication of both types showed similar number of chews, time for oral processing, and sum of muscle activities for the entire oral processing time. The effects of gel type per 24-g portions were almost the same as those per one mass of gel, as shown in Table 3. 3.4. Comparison ratio of stage T1 to entire oral processing As shown in Figs. 1, 3, and 4, rhythmic masseter activities were observed during T1, and these muscle activities in T1 weakened in T2 and the chewing cycle became irregular in T2. The initial masses remained in the mouth until the first swallow, allowing quantitative analysis of the effects of gel mass in T1 but not in T2. The amount of gel remaining in the oral cavity after the first swallow was not confirmed, but was probably much smaller than the initial mass. Therefore,

Table 3 Masseter EMGs analyzed for the dominant sides (DS) and non-dominant sides (NDS) per mass of gel. Type of gel

A gel

Mass of gel (g)

3

6

12

24

3

6

12

24

0.93 0.88 0.58 0.43 0.0234 0.0160 0.0151 0.0067 0.244 0.166 0.157 0.070

1.14 1.25 0.77 0.73 0.0256 0.0189 0.0171 0.0095 0.339 0.253 0.232 0.145

1.23 1.08 0.89 0.71 0.0294 0.0188 0.0204 0.0100 0.498 0.356 0.336 0.179

1.39 1.30 0.86 0.44 0.0340 0.0253 0.0226 0.0079 0.679 0.654 0.403 0.248

1.44 0.88 0.92 0.58 0.0296 0.0163 0.0186 0.0085 0.365 0.196 0.226 0.099

1.39 0.81 0.94 0.53 0.0290 0.0137 0.0189 0.0061 0.451 0.246 0.295 0.129

1.53 1.05 1.07 0.64 0.0330 0.0201 0.0222 0.0087 0.594 0.375 0.398 0.181

1.78 1.29 1.32 0.81 0.0383 0.0211 0.0287 0.0125 0.796 0.525 0.556 0.302

0.43 0.34 0.40 0.35 0.0213 0.0139 0.0195 0.0133 0.066 0.069 0.052 0.030

0.58 0.44 0.57 0.59 0.0211 0.0112 0.0177 0.0092 0.119 0.090 0.102 0.091

0.76 0.89 0.51 0.43 0.0243 0.0187 0.0153 0.0059 0.227 0.202 0.141 0.073

1.06 1.33 0.61 0.37 0.0291 0.0256 0.0182 0.0060 0.439 0.285 0.308 0.111

0.65 0.56 0.38 0.25 0.0272 0.0289 0.0272 0.0355 0.066 0.056 0.076 0.152

0.73 0.55 0.60 0.54 0.0228 0.0166 0.0188 0.0068 0.081 0.060 0.066 0.044

1.43 0.97 0.88 0.67 0.0325 0.0185 0.0199 0.0080 0.241 0.109 0.157 0.083

0.314 0.207 0.210 0.078

0.458 0.328 0.334 0.188

0.725 0.537 0.478 0.217

1.118 0.907 0.711 0.282

0.436 0.206 0.302 0.181

0.532 0.255 0.361 0.124

0.835 0.444 0.555 0.197

Before the first swallow (T1) Amplitudes, DS (mV) Amplitudes, NDS (mV) Muscle activities per chew, DS (mV.s) Muscle activities per chew, NDS (mV.s) Total muscle activities, DS (mV.s) Total muscle activities, NDS (mV.s) After the first swallow (T2) Amplitudes, DS (mV) Amplitudes, NDS (mV) Muscle activities per chew, DS (mV.s) Muscle activities per chew, NDS (mV.s) Total muscle activities, DS (mV.s) Total muscle activities, NDS (mV.s)

Entire oral processing Total muscle activities, DS (mV.s) Total muscle activities, NDS (mV.s)

K gel

Mass effect

Gel type effect

Cross effect

*S b L

*A b K

ns

***S b L

*A b K

ns

**S b L

*A b K

ns

***S b L

*A b K

ns

*S b L

**A b K

ns

**S b L

**A b K

*

1.49 0.94 1.00 0.71 0.0321 0.0165 0.0216 0.0117 0.508 0.268 0.386 0.228

*S b L

ns

*

**S b L

ns

ns

ns

ns

ns

ns

ns

ns

***S b L

ns

ns

***S b L

ns

ns

1.304 0.724 0.943 0.458

**S b L

ns

ns

***S b L

ns

ns

Mean (upper row) and standard deviation (lower row) values were calculated from duplicate measurements of 11 subjects; ns, not significant, *, p b 0.05, **, p b 0.01, ***, p b 0.001; analyzed by two-way repeated measure ANOVA. DS, dominant sides of chewing showing greater mean amplitudes during T1 for each chewing sequence: NDS, non-dominant (opposite) side. S (small) to L (large) masses of A (agar) or K (mixed gels of konjac mannan, κ-carrageenan, and locust bean gum) gels.

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Table 4 EMG variables during consumption of 24-g gels. Type of gel

A gel

Mass (g) × multiplication

3×8

Before the first swallow T1 (s) Sum of muscle activities (mV.s) Number of chews

Entire oral processing Time for oral processing (s) Sum of muscle activities (mV.s) Number of chews Number of swallows

K gel

Mass effect

Gel type effect

Cross effect

12.3 4.8 1.37 0.80 20.5 8.2

***S N L

ns

*

***S N L

*A b K

*

***S N L

*A b K

*

24.1 8.8 2.27 1.15 39.0 13.7 3.6 1.2

***S N L

ns

ns

***S N L

ns

ns

***S N L

ns

ns

***S N L

** A N K

*

6×4

12 × 2

24 × 1

3×8

6×4

12 × 2

24 × 1

43.6 13.6 3.21 1.83 86.2 25.6

30.6 12.8 2.40 1.72 56.5 23.4

18.9 7.1 1.67 1.05 34.2 12.7

10.9 4.5 1.11 0.87 19.0 8.3

56.3 20.1 4.73 2.25 105.5 35.3

34.2 12.0 2.99 1.46 64.2 21.1

19.8 6.6 1.98 1.07 36.5 11.6

64.1 17.9 4.19 2.21 110.2 27.6 14.5 3.7

49.4 12.6 3.22 2.11 77.3 23.8 8.9 1.4

33.8 9.5 2.43 1.47 53.9 13.7 6.3 1.9

24.7 7.9 1.84 1.16 37.6 11.0 3.9 1.3

67.6 26.5 5.48 2.35 121.8 38.0 10.9 4.4

44.3 15.5 3.57 1.44 80.4 24.3 7.5 2.5

32.0 10.5 2.78 1.21 54.6 14.6 5.4 2.4

Measured EMG variables for 3-g gels were multiplied by eight to estimate the EMG variables for 24-g portions. Similarly, those for 6-g gels were multiplied by four and those for 12-g gels were doubled. Sum of muscle activities were estimated as the sum of total muscle activities of both masseters. Mean (upper row) and standard deviation (lower row) values were calculated from duplicate measurements of 11 subjects; ns, not significant, *, p b 0.05, **, p b 0.01, ***, p b 0.001; analyzed by two-way repeated measure ANOVA. S (small) to L (large) masses of gels, and A (agar) or K (mixed gels of konjac mannan, κ-carrageenan, and locust bean gum) gels.

many studies have measured masseter EMGs with a fixed mass or volume of gel and for T1 only [4,5,14,25,42]. The mass of gel remaining after the first swallow increased with the total mass of gel. To confirm the hypothesis, the ratios of EMG parameters obtained in T1 to those obtained in the entire oral processing time were compared (Table 5). When the value was one, the gels were completely processed before the first swallow. Muscle activities for 3g gels were almost completed before the first swallow, and 3-g K gels were often swallowed whole without the need for T2. In contrast, the 24-g gels were processed to almost 50% before the first swallow (Table 5). Although there were significant cross-effects in oral processing times, the K gel generally exhibited higher ratios than the A gel, corresponding to a higher fracture strain. Masseter activities mainly appeared in T1 and were nearly zero in T2 for small mass of gel. Total muscle activities of the masseter muscle were used as an indicator of mastication effort [13], and the greatest decrease was seen upon increasing the sample mass from 3 g to 24 g. However, the doubling of gel mass did not lead to a two-fold increase in EMG variables. 4. Discussion 4.1. Gel properties and natural mastication behaviors The present study examined two types of gels (A and K) that had similar fracture loads. Both types of gels were expected to easily fracture

during the first chew under a similar force. However, other mechanical properties (Table 1) significantly differed between the two gels (p b 0.001). Specifically, fracture strain and work of elastic K gel were nearly seven times greater than those of brittle A gel, and fracture during the human chew-cycle of K gel would occur later than that of A gel. Apparent elastic modulus indicates the amount of force required to slightly deform the gels, and this was approximately 30 times lower in more deformable K gel compared to brittle A gel. However, as the gels were maximally deformed by chewing between the upper and lower teeth after fracture, the EMG variables were correlated with very large deformations, including compression strains of 80% or higher [9,41]. Accordingly, load at 90% strain and work until 90% strain were significantly greater for A gel compared with K gel, but these differences were relatively small (less than two times). This suggest that they may not significantly affect human mastication kinematics where compression speeds, ratios of deformation, temperatures in the mouth, saliva, tooth compositions, and other factors differ from the conditions of instrumental tests. Based on these properties, A gel was expected to require more muscle effort in the early stages of the first chew than K gel. However, in the following chews, the muscle effort was seen to decrease more rapidly for A gel because of its brittle characteristics, as indicated by lower fracture strain. Moreover, this gel also easily broke into fragments and lost structure faster than K gel during mastication. In contrast, small K gels did not separate into more than one fragment prior to swallowing as observed in other gels with similar textures [43]. These observations may

Table 5 Ratios of EMG variables before the first swallow to those during entire oral processing. Type of gel

A gel

Mass of gel (g)

3

6

12

24

3

6

12

24

Oral processing times, T1/(T1 + T2)

0.708 0.121 0.711 0.108 0.782 0.134 0.791 0.114

0.602 0.124 0.631 0.111 0.718 0.128 0.715 0.095

0.556 0.108 0.565 0.116 0.675 0.128 0.625 0.116

0.442 0.147 0.457 0.157 0.568 0.135 0.503 0.163

0.851 0.142 0.816 0.109 0.869 0.105 0.871 0.113

0.784 0.143 0.757 0.123 0.823 0.118 0.804 0.129

0.626 0.150 0.629 0.139 0.694 0.123 0.669 0.141

0.522 0.187 0.534 0.178 0.593 0.159 0.539 0.177

Total durations Total muscle activities Number of chews

K gel

Mass effect

Gel type effect

Cross effect

***S N L

***A b K

ns

***S N L

***A b K

ns

***S N L

*A b K

ns

***S N L

**A b K

ns

Mean (upper row) and standard deviation (lower row) values were calculated from duplicate measurements of 11 subjects; ns, not significant, *, p b 0.05, **, p b 0.01, ***, p b 0.001; analyzed by two-way repeated measure ANOVA. S (small) to L (large) masses of gels, and A (agar) or K (mixed gels of konjac mannan, κ-carrageenan, and locust bean gum) gels.

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reflect significant differences in the number of swallows required for A and K gels, as fragmented A gels cannot be swallowed in one movement unlike non-fragmented, smaller K gels. The texture of K gels is similar to that of typical mini-cup konjac jellies sold in Japan [18] and, as is indicated by smaller load values at 90% strain and work until 90% strain (Table 1), A gels lose their structure faster than K gels during oral processing. A previous study [18] reported rapid decreases in masseter EMG amplitudes for A gels during the early chewing cycles, reflecting smaller fracture strains and fracture energies. Furthermore, the above gel properties resulted in greater EMG amplitudes, muscle activities per chew, and total muscle activities before the first swallow for K gels compared to A gels (Table 3). In this study, we were unable to prepare cube samples for 24-g gels as the original gel height was only 25 mm. Moreover, K gels with low moduli were easily deformed under gravity, and specimens on spoons were therefore not cubes. Both gel types were transported to the back teeth on one side and chewed. However, samples thicker than 10 mm were difficult to bite with molars because of limited space between the maxillary and mandibular molars [44]. However, differences in the shapes of the gels did not significantly influence mastication variables, as previously discussed [24] and the original shape was completely destroyed during the first chew. Therefore, the study focused on the entire chewing sequence instead of only on the first chew. 4.2. EMGs from left and right masseter muscles Normalization to maximum voluntary contractions or submaximal reference voluntary contractions is recommended when analyzing surface EMGs from different muscles, subjects, and on different days [45, 46]. However, as discussed in our previous paper, maximum voluntary contractions are not a good standard for mastication EMGs of gels, as muscle action required for gel consumption is much less than maximum contractions [18]. Moreover, use of artificial foods that maintain similar physical properties, such as chewing gum, is not suitable for comparison as the textures of the gel samples change rapidly within single chews, and a reduction in size and structural breakdown becomes apparent [40]. EMGs should be tested using a two-factorial design (4 masses × 2 2 types of gel), and this makes it difficult to set a control sample for standardization. In this study, all samples were sequentially tested for differences between masseter EMGs of the two sides without changing recording conditions. Raw EMGs analyzed by repeated-measure ANOVA and standardized EMGs using a control sample provided similar results for the comparison of different foods [47]. Although masseter EMG activities increased with chewing force [4,5, 7,48], the conversion of these chewing forces to EMG voltages differed between LM and RM. Accordingly, symmetrically placed electrodes on both masseters did not exhibit similar voltages and both masseter activities appeared concomitantly during the jaw-closing phase in each chewing cycle in all subjects. Errors between different sides may have been caused by biological factors, such as conductivity of skin, muscle anatomy, and fat thickness, as well as instrumental conditions such as electrodes, codes, and amplifiers [45]. Previous studies have reported that the masseter EMG voltages appeared greater on the side of chewing compared to the opposite side in a majority of subjects, although the muscle activities of the opposite side were not zero [5–8,34,42,49,50]. Some subjects used both sides almost equally, resulting in similar RM and LM amplitudes in a single chew. However, the patterns of EMGs differed between subjects and the amplitudes and muscle activities per chew varied between RM and LM, as shown in Figs. 1, 3, and 4. The mean values of left and right masseter EMGs per chew and the sums of muscle activities were calculated. Table 3 shows that EMG variables from DS and NDS were similar with different samples. Rey et al. [51] analyzed EMGs from DS for side-imposed chewing gum tasks and reported that chewing behaviors of each subject were stable for over 2 months. However, as mentioned above, subjects masticated gel-type foods using both sides, including bilateral chews in natural chewing

behaviors. In particular, the example in Fig. 4 shows that nearly half of the information was lost from EMGs of DS, even though subjects with preferred chewing sides used NDS considerably (Table 3). Moreover, all subjects without a preferred habitual side of chewing and two subjects with preferred chewing sides changed their DS over 16 trials (Table 2). These observations suggest that self-reported preferred sides and DS determined initially may not be entirely accurate. Thus, averaged EMGs were influenced more by the side of chewing in all cases (Figs. 1, 3, and 4), and we suggest that means of masseter EMGs should be considered in analyses of natural chewing behaviors. 4.3. Dominant and non-dominant sides As observed frequently with small gels, the DS possibly plays a dominant role if a mouthful of food can be consumed with fewer chews (Fig. 1). Hence, differences in EMG amplitudes and muscle activities between DS and NDS are probably greater in such cases. Moreover, as the present subjects were allowed to change the side of chewing freely, this probably occurred more frequently for larger masses that required more chewing. NDS/DS ratios of amplitudes and muscle activities were calculated as means of the values from 11 subjects. These values were similar for 3-g gels, namely, 0.62 and 0.65 for A gel, and 0.64 and 0.63 for K gel, respectively (data not shown). NDS/DS ratios of amplitudes and muscle activities per chew for K gels logarithmically increased with the mass of gel (coefficients of determination of the best-fitting curves R2 = 0.995 and 0.902, respectively). They increased by 5% with doubled gel mass and the corresponding values for 24-g K gels were 0.74 and 0.75, respectively. In contrast, A gels did not exhibit this relationship, although ratios of amplitudes and muscle activities tended to increase from 3 g to 12 g of A gels. However, the corresponding values of 24 g of A gel were low at 0.62 and 0.66, respectively. With increasing gel mass, the contributions of NDS became more marked as subjects changed the sides of chewing more frequently as part of natural bilateral chewing. However, the present results for larger A gel was not in accordance with this general tendency, perhaps reflecting early swallow following easy fragmentation in one chew. Sides with greater EMG amplitudes and muscle activities were similar, and the first chew was often performed on this side, as shown in Table 2. Accordingly, subjects with a preferred habitual side started chewing gels with this side more during natural mastication. In addition, their chewing performance was higher on that side compared to the other side. In the present study, 7 of the 11 subjects started their first chew on a fixed side and performed subsequent chews on the same side, and the chewing patterns of individuals were common for all masses and types of gels. In the present study, A gels were easily broken into small pieces during the first chew, and the fragments were probably chewed on both sides during subsequent chewing cycles. At constant masses, these gels were masticated more easily and bilaterally than K gels, potentially leading to observation of greater muscle activities for K gels compared to A gels during T1, as well as and greater values of apparent modulus, load at 90% strain, and work until 90% strain, suggesting a greater chewing effort. In contrast, A gels were more brittle and broke with a fewer number of chews. However, although the average EMG amplitudes and total muscle activities were calculated for T1 and for entire time for oral processing, they did not correspond with rapid decreases during early stages of mastication. Usually, the first and later chews of non-fragmented gels are performed on a single side, while fragmented gels may be chewed using both sides. As the fragmentation of K gels was difficult, small K gels were often chewed using DS until the first swallow (Fig. 1). However, contribution of both sides allowed wider sectional areas between post-canine teeth for compression of the gels. Accordingly, more brittle A gels tended to be broken down with higher probability than K gels, and were processed by both sides, leading to a higher NDS/DS ratio.

K. Kohyama et al. / Physiology & Behavior 161 (2016) 174–182

These observations may explain clear logarithmical dependencies of the NDS/DS ratios on the mass of K gels compared to A gels. We assumed that natural mastication was performed in the most effective way [47]. Subjects were likely to start chewing a mouthful of gel with the DS, and use this side more than NDS. Separation of gel into broken pieces occurred more easily in A gel, and the spread of the bolus to the NDS may contribute to mastication. Moreover, chewing fatigue may warrant changes in the side of chewing, and the effects are likely to be more significant in larger masses of gels. Thus, the strongest EMG voltages are probably detected in the DS at the early stages of mastication as the gel structures are strongest at this point. The maximum voltages of NDS are probably shown during later stages when the required muscle power has decreased. The amplitudes of both masseters do not appear to have similar muscle power. Hence, the maximum amplitudes of the DS probably correspond with greater muscle work required for breakdown of the initial structure than those of the NDS. The absolute values of amplitudes and muscle activities from the NDS were smaller than those from the DS (Table 3). However, the effects of mass on these EMG variables were greater in the NDS than in the DS, as reflected by amplitudes, muscle activities per chew, total muscle activities in stage T1, amplitudes in stage T2, and total muscle activities during the entire oral process (Table 3). These results were confirmed by smaller contributions of the NDS than the DS during mastication of smaller mass of gels, although 3-g gels were sometimes chewed with only the DS until the first swallow, as shown in Fig. 1.

4.4. Mastication of mouthful masses and cup portions of gels In natural settings, side changes during chewing occur more frequently near the end of the chewing sequence [52], and bilateral chewing may be also involved. Moreover, subjects with preferences usually start chewing on the preferred side, as it is more effective in the early stages of mastication. Fractured gels separated into increasing numbers of fragments with increase in gel mass, and a greater number of fragments were observed with A gels compared to K gels. Unlike initial block gels, fragmented gels can be chewed bilaterally, and fatigue of muscles and oral organs probably motivates changes in the side of chewing, potentially contributing to eating pleasure in terms of time and space. Hence, during the later stages of mastication, contributions of the NDS may increase to avoid fatigue on the preferred side and to enlarge contact areas between the teeth and fragmented food. The largest mass was set at 24 g in this study as it was approximately the largest portion size of commercial mini-cup jellies available on the Japanese market [18]. In addition, the 24-g gels used in this study had a volume of approximately 27 mL. This is slightly smaller than the oral cavity volume, which is reported to be 30.1 ± 0.4 mL [53] and 33 ± 6 mL [54]. We explored the effects of mouthful mass when it was halved three times from 24 g (12, 6, and 3 g) in the previous study [18]. Further halved gel (1.5 g) was too small and not realistic as a mass of gel because it could be swallowed immediately without chewing. When comparing the results given in Tables 3 and 4, smaller masses of gels were seen to lead to lower muscle activities per mass and greater activities per 24 g. Both amplitudes and muscle activities per chew increased with increasing masses for both types of gels (Table 3), suggesting that the chewing force increased with the mass of gels. In contrast, although amplitudes remained constant, muscle activities increased proportionally with numbers of small masses (for example 3 g), and the time for oral processing and number of chews per cup significantly increased with smaller masses. Goto et al. [55] recently tested number of chews for boiled rice, fish sausages and peanuts, and reported that the number of chews increased with increasing masses from 0.5- and 1.5-folds of natural mouthful sizes of each subject. They also reported that the number of chews per gram of food was significantly reduced for the rice and sausages, which are regarded as gel-type foods. Their observations were the same as ours, but EMGs could also provide

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information about chewing force in addition to number of chews and time for oral processing. As recently suggested by Sun et al. [56], eating methods influence eating rate. Hence, eating gels using a small spoon may reduce gel masses and increase number of ingestions, and require more chews and longer times for oral processing without using stronger muscle power. Thus, reductions of gel masses were the easiest way to enhance chewing and slow eating rates as this did not require modifications of the mechanical properties of foods by changing formulas and/or preparation methods. The present study was limited by the inability to quantify measured mastication variables in experiments with eight 3 g masses, four 6 g masses, or two 12 g masses. Moreover, fullness and satiety of subjects probably influenced the results because sequential recordings for four different masses of 24-g portions, two-types of gels, and repetitions were difficult to perform. Accordingly, habitual eating without fixed masses is necessary to confirm the present effects of gel mass of gels on natural eating in future studies. 5. Conclusions In this study, we clarified the effects of mouthful masses of cups of gels (24 g) on natural eating behaviors using EMG. Differences in EMG variables for two types of gels (A and K) with similar fracture loads were used to adjust times for oral processing and were analyzed during the entire oral processing period, and before and after the first swallow. EMG observations suggest that the number of chews, time for oral processing, and total masseter muscle activities are likely to be determined by the fracture load of gel as the fracture load of A and K gels did not differ significantly. Despite similar fracture loads, K gels with greater fracture strain and work required greater mastication efforts before the first swallow, and also required fewer swallows than A gels, which had a higher modulus. Subsequent single-side EMG analysis showed similar effects of masses and types of gels, regardless of the side of chewing (DS or NDS). Therefore, it was reasonable to calculate average LM and RM EMGs as both sides were used when foods were naturally eaten. Thus, regardless of chewing with the DS, NDS or bilateral chewing, average values were more strongly influenced by the DS. Multiple subjects performed the first chew with the DS, as previously reported, and the ratio of muscle activities of NDS to DS increased with masses of K gels. However, the dependency of mouthful masses was less significant for A gels. Finally, with decreasing masses of gels from 24 g to 3 g, the number of chews, time for oral processing, and mastication efforts per 24 g of gel increased, and the ratio of mastication before the first swallow to that during the entire oral process increased. However, reduced masses of gels and an increased number of mouthfuls resulted in greater mastication per gel portion (24 g). Small mouthful masses may be useful to prevent overeating as this slows the eating process. References [1] M.C. Bourne, Body–texture interactions, Food texture and viscosity: concept and measurement, second ed. Academic Press, San-Diego 2002, p. 48. [2] J. Chen, Food oral processing – a review, Food Hydrocoll. 23 (2009) 1–25, http://dx. doi.org/10.1016/j.foodhyd.2007.11.013. [3] K. Kohyama, Oral sensing of food properties, J. Texture Stud. 46 (2015) 138–151, http://dx.doi.org/10.1111/jtxs.12099. [4] A. Woda, W. Foster, A. Mishellany, M.A. Peyron, Adaptation of healthy mastication to factors pertaining to the individual or to the food, Physiol. Behav. 89 (2006) 28–35, http://dx.doi.org/10.1016/j.physbeh.2006.02.013. [5] Y. Espinosa, J. Chen, Applications of electromyography (EMG) technique for eating studies, in: J. Chen, L. Engelen (Eds.), Food oral processing fundamentals of eating and sensory perception, Wiley-Blackwell, Oxford 2012, pp. 289–317, http://dx.doi. org/10.1002/9781444360943.ch13. [6] H. Koç, C.J. Vinyard, G.K. Essick, E.A. Foegeding, Food oral processing: conversion of food structure to textural perception, Ann. Rev. Food Sci. Technol. 4 (2013) 237–266, http://dx.doi.org/10.1146/annurev-food-030212-182637.

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