**2.1 Ethical aspects**

130 Otolaryngology

chronic edema and in those with hyperfunctional dysphonia (Pinho & Pontes, 2008). Some authors also recommend tongue trills in cases of hypofunctional dysphonia (Behlau e Pontes, 1995). Manieka-Aleksandrovix (2006) collected data regarding 500 patients with aphonia due to psychogenic dysphonia and found that gargling is one of the exercises used

Casper *et al.* (1992) employed tongue trills as a therapeutic resource in individuals with vocal fold paralysis and in those having undergone laryngeal surgery (Woo *et al.*, 1994). The tongue trill is contraindicated for individuals with recent-onset acute inflammation, because the exercise can aggravate the inflammatory phase. In the immediate postoperative period, tongue trills can delay healing (Pinho & Pontes, 2008) and, in cases of papillomatosis,

When performing trill exercises, individuals should keep the tongue (or lips) and the mandible relaxed, coordinating the airflow so that vibration can occur (Scneider & Sataloff, 2007). The tongue trill is maintained by the interaction among the firmness of the body of tongue, control of the tip of the tongue, glottic closure, and control of the exhaled air. The exercise should be performed with the sides of the body of tongue firmly pressed against the dental alveoli and the tip of the tongue positioned in the region of the incisive papilla, free to vibrate (McGowan, 1992). As a result, the entire vocal tract vibrates (Scwarz & Cielo, 2009). For lip vibration (lip trill) to occur, the lips should be held together tightly enough to promote airway occlusion and relaxed enough for air pressure to overcome the resistance (Gaskill e Erickson, 2008). During lip trills, as during tongue trills, there is interaction among the vocal tract, glottal vibration, and the exhaled air during lip trills (Titze, 2006). During all trill exercises, the vibrating organ acts as a valve and creates oscillatory differences in external pressure and in the pressure in the cavity behind the constriction. This produces differences in the pressure, speed, and volume of air in the oral cavity, causing changes in the pharyngeal wall. Therefore, for vocal fold vibration to occur concomitantly with the point of oscillation of the oral cavity, subglottic air pressure must be greater than is that

Tongue and lip vibration follow the same principle as does the vocal fold mass effect: the anterior part of the vocal tract is occluded by the tip of the tongue or the lips. Intraoral pressure becomes greater than the atmospheric pressure and therefore greater than the force that maintains the anterior part of the vocal tract closed. Therefore, the anterior part of the vocal tract opens and is subsequently "sucked out" by the speed of the airflow (McGowan,

According to Gaskill and Erickison (2008), the difference between lip trills and other exercises that focus on the anterior part of the vocal tract is that the lip trill is the only exercise that promotes lip occlusion and non-occlusion (without loss of muscle tone). This causes the lips to vibrate, although at a frequency lower than that of vocal fold vibration. Therefore, airflow and subglottic pressure must adjust in order to allow the lips and the vocal folds to vibrate, overloading the vocal folds. The variations that occur in the pharynx during lip trills can increase the force of mucosal vibration during the wavelike motion of

Because of high vocal demand, professional voice users should maintain the fitness of all of the structures involved in phonation. For professional voice users, voice training should

on the first day of therapy for voice rehabilitation.

during normal phonation (McGowan, 1992).

the vocal folds (McGowan, 1992).

1992).

stimulate the dissemination of the disease (Pinho & Tsuji, 2006).

The present study was approved by the Research Ethics Committee of the *Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo* (HCFMUSP, University of São Paulo School of Medicine *Hospital das Clínicas*; Protocol no. 907/06, February 14, 2007; Appendix 1), located in the city of São Paulo, Brazil. All participating individuals gave written informed consent.

#### **2.2 Study sample**

In the present study, we evaluated 14 individuals (7 males and 7 females). We applied the following criteria:

Comparison Among Phonation of the Sustained Vowel /ε/, Lip Trills,

exercises.

dB for the highest intensity

were performed again before each test.

**2.3.2 Data collection and analysis** 

and Tongue Trills: The Amplitude of Vocal Fold Vibration and the Closed Quotient 133

and /з/, at the highest and lowest possible intensities of which they were capable. As shown in Table 2, we chose the lowest of the highest intensities and the highest of the lowest intensities, in order to standardize the intensities during exercise training and data collection. The voiced fricatives were used because the lowest of the highest intensities is commonly achieved during the phonation of those sounds; therefore, the participants were able to perform the tests comfortably without experiencing vocal fatigue or aperiodicity due to the use of threshold-range phonation (Jiang *et al.*, 2001) during the performance of trill

Intensity Vowel /ε/ Lip trill Tongue trill /v/ /z/ /з/ Highest (dB) 51 52 52 <50 <50 51 Lowest (dB) 85 70 71 68 70 71

phonations. The values chosen for data collection were 52 dB for the lowest intensity and 68

In order to measure vocal range, as well as to determine and maintain the selected note, we used a Casio VL-Tone-VL1 keyboard (Casio Computer Co., Ltd., Tokyo, Japan). Intensity was measured with a sound pressure level meter (model 33-2055, RadioShack Corporation,

After the abovementioned procedure, each singer practiced producing the sounds at the requested note and at the intensity selected (maximum variation, 2 dB), with minimal effort and maintaining the larynx in the same (low) position, for ≥ 10 seconds. The practice session was conducted with the aid of a speech-language pathologist specializing in voice, who instructed the singers during a meeting held before data collection. On the day of data collection, previously agreed upon by the singers and the investigators, the phonation tasks

Participants were informed of the objectives and conditions of the present study before the first evaluation was scheduled. If a participant presented with illness, sleep deprivation, vocal fatigue, or dysphonia on the day of data collection, the evaluation was rescheduled.

The individuals underwent videolaryngostroboscopy by an experienced otolaryngologist of the HCFMUSP Clinical Otolaryngology Division Voice Group Outpatient Clinic and EGG at the *Centro de Especialização em Fonoaudiologia Clínica* (CEFAC, Center for Clinical Audiology and Speech-Language Pathology), both located in the city of São Paulo. The tests were performed on different days so that the singers did not experience vocal fatigue. For the tests, the individuals were asked to produce the sustained vowel /ε/ and perform tongue and lip trills for as long as possible at the same frequency and intensities as those used in the training sessions (5th whole step above the lowest note in their range and the lowest of the highest/highest of the lowest, respectively). Frequency was controlled by the singers. In order to do that, they used the same keyboard that was used in the training sessions. Intensity was monitored by a speech-language pathologist in the examination room, using

Table 2. Highest and lowest intensities achieved by individual 1 during sustained

Fort Worth, TX, USA) placed 30 cm from the corner of the mouth of the singer.


A total of 4 individuals were excluded from the present study. Of those, 2 were male and 2 were female. Of the 2 males, 1 was excluded because he could not tolerate the examination of the larynx and 1 was excluded because he had been a professional singer for less than three years (one year and six months). Of the 2 females, 1 was excluded because she could not tolerate the examination of the larynx and 1 was excluded because she presented with speaking voice complaints.


A total of 10 singers were analyzed. Their characteristics are shown in Table 1.

Table 1. Characteristics of the individuals included in the study sample

#### **2.3 Method**

#### **2.3.1 Preparation for data collection**

Before data collection, we measured the vocal range of each participant. Vocal range is the distance between the lowest and the highest note that an individual can produce, excluding the vocal fry register and including the falsetto.

After having measured the vocal range of the participants, we selected the 5th whole step above the lowest possible note that each individual was able to produce (Cooper, 1979). The participants were then asked to produce, in that note and with their larynx in a low position, the sustained vowel /ε/, lip trills, and tongue trills, as well as the voiced fricatives /v/, /z/,

 inclusion criteria—being a healthy, classically trained, professional singer; having laryngeal control; having mastered the techniques for performing lip and tongue trills;

 exclusion criteria—having been a professional singer for less than three years; presenting with singing or speaking voice complaints; presenting with incomplete

A total of 4 individuals were excluded from the present study. Of those, 2 were male and 2 were female. Of the 2 males, 1 was excluded because he could not tolerate the examination of the larynx and 1 was excluded because he had been a professional singer for less than three years (one year and six months). Of the 2 females, 1 was excluded because she could not tolerate the examination of the larynx and 1 was excluded because she presented with

**professionally Voice type** 

glottal closure; having reported intolerance to examination of the larynx

A total of 10 singers were analyzed. Their characteristics are shown in Table 1.

1 Female 24 4 Soprano 2 Female 45 20 Soprano

5 Female 48 15 Contralto 6 Male 29 5 Tenor 7 Male 27 6 Tenor 8 Male 33 12 Baritone 9 Male 34 15 Baritone

10 Male 38 18 Bass

Before data collection, we measured the vocal range of each participant. Vocal range is the distance between the lowest and the highest note that an individual can produce, excluding

After having measured the vocal range of the participants, we selected the 5th whole step above the lowest possible note that each individual was able to produce (Cooper, 1979). The participants were then asked to produce, in that note and with their larynx in a low position, the sustained vowel /ε/, lip trills, and tongue trills, as well as the voiced fricatives /v/, /z/,

Table 1. Characteristics of the individuals included in the study sample

3 Female 30 3 Mezzo-soprano 4 Female 30 4 Mezzo-soprano

**Individual Gender Age Years singing** 

and presenting with no vocal fold lesions

speaking voice complaints.

**2.3 Method** 

**2.3.1 Preparation for data collection** 

the vocal fry register and including the falsetto.

and /з/, at the highest and lowest possible intensities of which they were capable. As shown in Table 2, we chose the lowest of the highest intensities and the highest of the lowest intensities, in order to standardize the intensities during exercise training and data collection. The voiced fricatives were used because the lowest of the highest intensities is commonly achieved during the phonation of those sounds; therefore, the participants were able to perform the tests comfortably without experiencing vocal fatigue or aperiodicity due to the use of threshold-range phonation (Jiang *et al.*, 2001) during the performance of trill exercises.


Table 2. Highest and lowest intensities achieved by individual 1 during sustained phonations. The values chosen for data collection were 52 dB for the lowest intensity and 68 dB for the highest intensity

In order to measure vocal range, as well as to determine and maintain the selected note, we used a Casio VL-Tone-VL1 keyboard (Casio Computer Co., Ltd., Tokyo, Japan). Intensity was measured with a sound pressure level meter (model 33-2055, RadioShack Corporation, Fort Worth, TX, USA) placed 30 cm from the corner of the mouth of the singer.

After the abovementioned procedure, each singer practiced producing the sounds at the requested note and at the intensity selected (maximum variation, 2 dB), with minimal effort and maintaining the larynx in the same (low) position, for ≥ 10 seconds. The practice session was conducted with the aid of a speech-language pathologist specializing in voice, who instructed the singers during a meeting held before data collection. On the day of data collection, previously agreed upon by the singers and the investigators, the phonation tasks were performed again before each test.

Participants were informed of the objectives and conditions of the present study before the first evaluation was scheduled. If a participant presented with illness, sleep deprivation, vocal fatigue, or dysphonia on the day of data collection, the evaluation was rescheduled.

## **2.3.2 Data collection and analysis**

The individuals underwent videolaryngostroboscopy by an experienced otolaryngologist of the HCFMUSP Clinical Otolaryngology Division Voice Group Outpatient Clinic and EGG at the *Centro de Especialização em Fonoaudiologia Clínica* (CEFAC, Center for Clinical Audiology and Speech-Language Pathology), both located in the city of São Paulo. The tests were performed on different days so that the singers did not experience vocal fatigue. For the tests, the individuals were asked to produce the sustained vowel /ε/ and perform tongue and lip trills for as long as possible at the same frequency and intensities as those used in the training sessions (5th whole step above the lowest note in their range and the lowest of the highest/highest of the lowest, respectively). Frequency was controlled by the singers. In order to do that, they used the same keyboard that was used in the training sessions. Intensity was monitored by a speech-language pathologist in the examination room, using

Comparison Among Phonation of the Sustained Vowel /ε/, Lip Trills,

tongue trills, 3 photographs

tongue trills, 3 photographs

unedited image; and in b, edited image.

region corresponding to the cuneiform cartilage (Figure 3).

from a total of 220 photographs.

(Figures 2a and 2b).

and Tongue Trills: The Amplitude of Vocal Fold Vibration and the Closed Quotient 135

USA). The digitized images were analyzed frame by frame, and, for each individual, we

low intensity—sustained vowel /ε/, 5 photographs; lip trills, 3 photographs; and

high intensity—sustained vowel /ε/, 5 photographs; lip trills, 3 photographs; and

The photos were edited with Adobe Photoshop CS2 software, version 9.0 (Adobe Systems Incorporated, San Jose, CA, USA), which allowed better visualization of the amplitude of vocal fold vibration, as well as of the limits of the region of the right arytenoid cartilage

a b Fig. 2. Photograph of the maximum amplitude of vocal fold vibration during lip trills. In a,

In order to measure the maximum amplitude of vibration, we imported the edited images into the program X-Cade, version 2.0, specifically designed for the present study by Dr. Arlindo Neto Montagnoli, an engineer at the Federal University of São Carlos School of Electrical Engineering, located in the city of São Carlos, Brazil. Measurements were taken

In order to reduce the number of measurement errors caused by variations in the position of the fiberoptic laryngoscope, we also measured the length of the anatomical structure near the arytenoid region. The structure is a cuneiform cartilage covered by the arytenoid mucosa and was used as a reference for comparison. That structure was chosen because its configuration is not changed by the phonation task being performed. We first measured the largest diameter of the abovementioned structure, which the software automatically converted into a comparative reference that was used in order to measure the amplitude of vocal fold vibration. Therefore, when we refer to the amplitude of vocal fold vibration as being 0.5, this means that the amplitude value corresponds to half the measurement of the

All of the measurements were taken in a blinded fashion by the same rater, who, for each individual evaluated, adopted the following self-calibration method: before considering a measurement to be valid, the rater measured the images obtained from a given individual

obtained photographs of the following phonations at their peak amplitudes:

the same sound pressure level meter that was used in the training sessions, which was again positioned at 30 cm from the corner of the mouth of the singer. We made intraindividual comparisons. Low-intensity phonations were compared only with one another, as were high-intensity phonations.

#### **2.3.2.1 Videolaryngostroboscopy**

We used videolaryngostroboscopy in order to measure the amplitude of vocal fold vibration during the exercises. The singers were instructed to sit in a chair, with both hips well supported, head held straight, and the chin at a 90° angle to the neck. In order to maintain the head of each participant fixed in position, the chair was equipped with a headrest that was adjusted to the height of the individual and a piece of foam that was covered with comfortable fabric, measuring 14 cm in width, 10 cm in height, and 7 cm in depth. In addition, their heads were secured to the headrest with a headband (Figure 1).

Fig. 1. Photograph of a singer undergoing videolaryngostroboscopy.

Images of vocal fold vibration were obtained with a stroboscopic light source (4914; Brüel & Kjær Sound & Vibration Measurement A/S, Nærum, Denmark) and captured with a 3.2 mm fiberoptic laryngoscope (ENT-30PIII; Machida Endoscope Co., Ltd., Tokyo, Japan), positioned so that the region of the arytenoid cartilages and the vocal folds were fully visible. In addition, we attempted to keep variations in the angle and distance of recording to a minimum. To that end, the otolaryngologist held the fiberoptic laryngoscope between the thumb and middle finger, with the index finger resting on the tip of the nose of the individual being examined (Figure 1).

The fiberoptic laryngoscope was connected to a charge-coupled device camera (IK-M41A; Toshiba, Tokyo, Japan), and the images were recorded on videotape with an NTSC videocassette recorder (NV-FS90; Panasonic Corporation, Osaka, Japan). The images were digitized on a computer (Inspiron 1525; Dell, Inc., Round Rock, TX, USA) equipped with a 1.73-GHz Pentium Duo CPU T2370 processor (Intel Corporation, Santa Clara, CA, USA), 2 GB of RAM, and a 32-bit operating system. In order to digitize the images, we used a video capture card (PCTV Pro USB; Avid Technology, Inc., Burlington, MA, USA) and the program Studio QuickStart, version 10.8 (Pinnacle/Avid Technology, Inc., Burlington, MA,

the same sound pressure level meter that was used in the training sessions, which was again positioned at 30 cm from the corner of the mouth of the singer. We made intraindividual comparisons. Low-intensity phonations were compared only with one another, as were

We used videolaryngostroboscopy in order to measure the amplitude of vocal fold vibration during the exercises. The singers were instructed to sit in a chair, with both hips well supported, head held straight, and the chin at a 90° angle to the neck. In order to maintain the head of each participant fixed in position, the chair was equipped with a headrest that was adjusted to the height of the individual and a piece of foam that was covered with comfortable fabric, measuring 14 cm in width, 10 cm in height, and 7 cm in depth. In

addition, their heads were secured to the headrest with a headband (Figure 1).

Fig. 1. Photograph of a singer undergoing videolaryngostroboscopy.

individual being examined (Figure 1).

Images of vocal fold vibration were obtained with a stroboscopic light source (4914; Brüel & Kjær Sound & Vibration Measurement A/S, Nærum, Denmark) and captured with a 3.2 mm fiberoptic laryngoscope (ENT-30PIII; Machida Endoscope Co., Ltd., Tokyo, Japan), positioned so that the region of the arytenoid cartilages and the vocal folds were fully visible. In addition, we attempted to keep variations in the angle and distance of recording to a minimum. To that end, the otolaryngologist held the fiberoptic laryngoscope between the thumb and middle finger, with the index finger resting on the tip of the nose of the

The fiberoptic laryngoscope was connected to a charge-coupled device camera (IK-M41A; Toshiba, Tokyo, Japan), and the images were recorded on videotape with an NTSC videocassette recorder (NV-FS90; Panasonic Corporation, Osaka, Japan). The images were digitized on a computer (Inspiron 1525; Dell, Inc., Round Rock, TX, USA) equipped with a 1.73-GHz Pentium Duo CPU T2370 processor (Intel Corporation, Santa Clara, CA, USA), 2 GB of RAM, and a 32-bit operating system. In order to digitize the images, we used a video capture card (PCTV Pro USB; Avid Technology, Inc., Burlington, MA, USA) and the program Studio QuickStart, version 10.8 (Pinnacle/Avid Technology, Inc., Burlington, MA,

high-intensity phonations.

**2.3.2.1 Videolaryngostroboscopy** 

USA). The digitized images were analyzed frame by frame, and, for each individual, we obtained photographs of the following phonations at their peak amplitudes:


The photos were edited with Adobe Photoshop CS2 software, version 9.0 (Adobe Systems Incorporated, San Jose, CA, USA), which allowed better visualization of the amplitude of vocal fold vibration, as well as of the limits of the region of the right arytenoid cartilage (Figures 2a and 2b).

Fig. 2. Photograph of the maximum amplitude of vocal fold vibration during lip trills. In a, unedited image; and in b, edited image.

a b

In order to measure the maximum amplitude of vibration, we imported the edited images into the program X-Cade, version 2.0, specifically designed for the present study by Dr. Arlindo Neto Montagnoli, an engineer at the Federal University of São Carlos School of Electrical Engineering, located in the city of São Carlos, Brazil. Measurements were taken from a total of 220 photographs.

In order to reduce the number of measurement errors caused by variations in the position of the fiberoptic laryngoscope, we also measured the length of the anatomical structure near the arytenoid region. The structure is a cuneiform cartilage covered by the arytenoid mucosa and was used as a reference for comparison. That structure was chosen because its configuration is not changed by the phonation task being performed. We first measured the largest diameter of the abovementioned structure, which the software automatically converted into a comparative reference that was used in order to measure the amplitude of vocal fold vibration. Therefore, when we refer to the amplitude of vocal fold vibration as being 0.5, this means that the amplitude value corresponds to half the measurement of the region corresponding to the cuneiform cartilage (Figure 3).

All of the measurements were taken in a blinded fashion by the same rater, who, for each individual evaluated, adopted the following self-calibration method: before considering a measurement to be valid, the rater measured the images obtained from a given individual

Comparison Among Phonation of the Sustained Vowel /ε/, Lip Trills,

accordance with the grading system proposed by Vieira (1997):

 grade 3, able to trace signals with irregular excitation grade 4, able to locate every individual glottal stop

and standard deviation of the closed quotient for each task.

Fig. 4. Phases of the EGG waveform and the closed quotient (CQ).

In order to analyze the results, we adopted a level of significance of 5% (p = 0.05) for all statistical tests. The Statistical Package for the Social Sciences, version 13.0 (SPSS Inc., Chicago, IL, USA) was used for the analysis. In order to determine the differences among phonation of the sustained vowel /ε/, lip trills, and tongue trills, we used Friedman's test.

**2.4 Statistical analysis** 

the variation in that measurement over time.

 grade 1, free from gross errors grade 2, as accurate as possible

and Tongue Trills: The Amplitude of Vocal Fold Vibration and the Closed Quotient 137

The EGG waveforms were submitted to high band-pass filtering with the program delay0.bat, designed by Maurílio Nunes Vieira, an engineer at the Federal University of Minas Gerais, located in the city of Belo Horizonte, Brazil. The waveforms were initially graded by two speech-language pathologists with experience in EGG and one engineer, in

Only grade 1 and grade 2 waveforms were included in the study. In order to take automatic measurements, we used the technique developed by Vieira (1997). We assessed the mean

The closed quotient is the ratio between the closed phase and the complete cycle of the EGG waveform (Figure 4). The mean closed quotient is the measurement of each EGG waveform divided by the number of waveforms analyzed by the software. The standard deviation is

several times until the measurements taken from the same image were within two decimal places of each other. The images from each individual were measured without interruption. Otherwise, the calibration process was restarted, and the measurements were taken again. The data were tabulated and entered into a database for subsequent statistical analysis.

Fig. 3. X-Cade software. The amplitude measurement corresponds to 0.4 of the measurement of the arytenoid cartilage.

#### **2.3.2.2 EGG**

In order to measure the EGG signal, we referred the individuals to a sound-treated booth in a quiet room in the CEFAC Voice and Speech Laboratory. We used an EG2-Standard electroglottograph (Glottal Enterprises, Syracuse, NY, USA). The electroglottograph was connected to a computer audio interface (BCA2000; MUSIC Group Services EU GmbH, Willich, Germany), which was in turn connected to a computer equipped with a 1.66-GB Centron processor (Advanced Micro Devices, Inc., Sunnyvale, CA, USA).

The individuals were asked to remove any metal objects that they might be wearing in the head and neck region and to sit upright in a chair equipped with a headrest. To ensure the safety of the individuals, we placed a rubber mat under the chair. The neck region was cleaned with dry paper towel. We applied a thin layer of hypoallergenic electrically conductive gel (SPECTRA 360®; Parker Laboratories, Inc., Fairfield, NJ, USA) to the electrodes, which were placed over each ala of the thyroid cartilage and secured with a Velcro strap around the neck. To ensure that the electrodes were positioned correctly, we asked the participants to produce the sustained vowel and perform tongue trills. We then observed whether the green LEDs of the top LED array (Electrode Placement/Laryngeal Movement) on the front panel of the electroglottograph were on. To confirm that there was signal, we observed whether the green LEDs of the bottom LED array (Signal) were on. To record the signal, we selected the Vocal Fold Contact Area signal option, and high or low gain was determined by monitoring the LED signal indicator. The signal was saved as a .wav file and edited with the audio editing suite Sound Forge, version 7.0, (Sony Creative Software, Inc., Middleton, WI, USA) at a sampling frequency of 22,050 Hz and a resolution of 16 bits.

The EGG waveforms were submitted to high band-pass filtering with the program delay0.bat, designed by Maurílio Nunes Vieira, an engineer at the Federal University of Minas Gerais, located in the city of Belo Horizonte, Brazil. The waveforms were initially graded by two speech-language pathologists with experience in EGG and one engineer, in accordance with the grading system proposed by Vieira (1997):

grade 1, free from gross errors

136 Otolaryngology

several times until the measurements taken from the same image were within two decimal places of each other. The images from each individual were measured without interruption. Otherwise, the calibration process was restarted, and the measurements were taken again. The data were tabulated and entered into a database for subsequent statistical analysis.

Fig. 3. X-Cade software. The amplitude measurement corresponds to 0.4 of the measurement

In order to measure the EGG signal, we referred the individuals to a sound-treated booth in a quiet room in the CEFAC Voice and Speech Laboratory. We used an EG2-Standard electroglottograph (Glottal Enterprises, Syracuse, NY, USA). The electroglottograph was connected to a computer audio interface (BCA2000; MUSIC Group Services EU GmbH, Willich, Germany), which was in turn connected to a computer equipped with a 1.66-GB

The individuals were asked to remove any metal objects that they might be wearing in the head and neck region and to sit upright in a chair equipped with a headrest. To ensure the safety of the individuals, we placed a rubber mat under the chair. The neck region was cleaned with dry paper towel. We applied a thin layer of hypoallergenic electrically conductive gel (SPECTRA 360®; Parker Laboratories, Inc., Fairfield, NJ, USA) to the electrodes, which were placed over each ala of the thyroid cartilage and secured with a Velcro strap around the neck. To ensure that the electrodes were positioned correctly, we asked the participants to produce the sustained vowel and perform tongue trills. We then observed whether the green LEDs of the top LED array (Electrode Placement/Laryngeal Movement) on the front panel of the electroglottograph were on. To confirm that there was signal, we observed whether the green LEDs of the bottom LED array (Signal) were on. To record the signal, we selected the Vocal Fold Contact Area signal option, and high or low gain was determined by monitoring the LED signal indicator. The signal was saved as a .wav file and edited with the audio editing suite Sound Forge, version 7.0, (Sony Creative Software, Inc., Middleton, WI, USA) at a sampling frequency of 22,050 Hz and a resolution

Centron processor (Advanced Micro Devices, Inc., Sunnyvale, CA, USA).

of the arytenoid cartilage.

**2.3.2.2 EGG** 

of 16 bits.


Only grade 1 and grade 2 waveforms were included in the study. In order to take automatic measurements, we used the technique developed by Vieira (1997). We assessed the mean and standard deviation of the closed quotient for each task.

The closed quotient is the ratio between the closed phase and the complete cycle of the EGG waveform (Figure 4). The mean closed quotient is the measurement of each EGG waveform divided by the number of waveforms analyzed by the software. The standard deviation is the variation in that measurement over time.

Fig. 4. Phases of the EGG waveform and the closed quotient (CQ).

#### **2.4 Statistical analysis**

In order to analyze the results, we adopted a level of significance of 5% (p = 0.05) for all statistical tests. The Statistical Package for the Social Sciences, version 13.0 (SPSS Inc., Chicago, IL, USA) was used for the analysis. In order to determine the differences among phonation of the sustained vowel /ε/, lip trills, and tongue trills, we used Friedman's test.

Comparison Among Phonation of the Sustained Vowel /ε/, Lip Trills,

\*Wilcoxon signed rank test

trills.

intensity

**3.1.2 High intensity** 

Variable N

\*Friedman's test, among all three variables

and Tongue Trills: The Amplitude of Vocal Fold Vibration and the Closed Quotient 139

Table 4. Pairwise comparisons among phonation of the sustained vowel /ε/, lip trills, and

a 0.08 b 0.11 c 0.12

Fig. 5. Maximum amplitude of vocal fold vibration during phonation tasks at low intensity in relation to the cuneiform cartilage, as assessed by videolaryngostroboscopy. In a, phonation of the sustained vowel /ε/; in b, sustained lip trills; and in c, sustained tongue

/ε/ 10 0.16 0.05 0.11 0.30 0.15

Table 5. Maximum amplitude of vocal fold vibration during phonation of the sustained vowel /ε/ compared with that of vocal fold vibration during tongue and lip trills at high

Tongue trills 10 0.26 0.12 0.15 0.50 0.21

Lip trills 10 0.29 0.08 0.16 0.40 0.30 0.001\*

Vibration amplitude

Mean SD Minimum Maximum Median

P

Pair p\* Lip trills versus sustained vowel /ε/ phonation 0.007 Tongue trills versus sustained vowel /ε/ phonation 0.005 Tongue trills versus lip trills 0.677

tongue trills at low intensity in terms of the amplitude of vocal fold vibration

For the cases in which the difference was statistically significant, we used the Wilcoxon signed rank test in order to identify the types that differed.

For the presentation of the results, the measurements taken automatically by the program were considered to constitute the intraindividual means and standard deviations, whereas the values obtained by statistical analysis of those results were considered to constitute the interindividual means and standard deviations.
