1. Manchester Metropolitan University
Department of Exercise and Sport Science
Research Project
Does timing of omega-3 supplementation impact its
effectiveness on reducing DOMS?
11021865
Keywords: exercise-induced muscle damage, DOMS, omega-
3, resistance exercise, recovery.
2. Abstract
Exercise-induced muscle damage (EIMD) and symptoms of delayed-onset muscle soreness
(DOMS) affect muscle functionality and athletic performance. Omega-3 fatty acids (omega-3)
exhibit anti-inflammatory properties (Magee et al., 2008). However, it is still unclear whether
timing of omega-3 supplementation would impact its effectiveness in minimising DOMS.
Twelve healthy males (age 20.7±2.4 years, height 181.7±7.2 cm and body mass 80.2±10.3 Kg)
were randomly assigned to either a pre- (n=6; PRE-EX) or post- (n=6; POST-EX) heavy
resistance exercise, omega-3 supplementation group (1528g EPA and 472g DHA). Torque,
soreness, swelling (limb circumference and muscle thickness) and knee range of motion were
assessed prior to exercise (baseline), and 0, 24, 48, 72 and 96hrs following exercise.
Loss of torque in PRE-EX was significantly greater than that in POST-EX at 0hrs (P = 0.02).
Soreness increased in response to exercise in both groups. Differences in reduction in pain
following exercise was observed between groups, with PRE-EX showing lower ratings of
soreness at 0, 24 and 48-hour time points (P > 0.05). Magnitude of muscle thickness increment
relative to baseline suggested a trend for the greatest effect in PRE-EX, particularly in the
rectus femoris at 48hrs (+8.5%, P > 0.05) and 72hrs (+8.5%, P > 0.05).
These findings suggest that PRE-EX supplementation may provide greater benefits in
minimising post-exercise soreness, whereas POST-EX supplementation appear to be more
effective in reducing swelling and loss of torque. Findings from the study would support
recommendations of supplementing an adequate dose of omega-3 prior to heavy resistance
exercise and during the recovery period to effectively minimise DOMS and improve recovery.
3. Acknowledgements
Completion of this research project would not have been possible without a number of people
who have had a significant influence on me in a variety of different ways. I would like to thank
Dr. Gladys Onambele-Pearson, whose encouragement and support have helped me greatly over
the course of this project. You have been an inspiration to me!
My brother Akim, and all of my friends who have had to hear my endless ramblings, but who
have always done so patiently, kindly and encouragingly. My thanks must also go to all the
individuals who agreed to take part during the data collection process. Without your effort and
commitment, this would not have been possible!
My biggest thanks must go to my parents, Mohamed and Kadie. Your love and support has
been limitless, and a huge inspiration to me when things were not going exactly to plan! My
final thanks must go to my late Grandad (RIP), who taught me that with hard work,
commitment, dedication and perseverance, anything is possible. This is for you!
4. Contents
Introduction..............................................................................................................................1
2. Methods .............................................................................................................................5
2.1 Participants.................................................................................................................5
2.2 Study Design................................................................................................................5
2.3 Omega-3 Supplementation..........................................................................................6
2.4 Exercise Protocol........................................................................................................7
2.5 Muscle Soreness ..........................................................................................................8
2.6 Strength Assessment....................................................................................................9
2.7 Muscle Thickness And Limb Circumference .............................................................10
2.8 Range Of Motion .......................................................................................................12
2.9 Statistical Analysis ....................................................................................................12
3. Results..............................................................................................................................14
3.1 Post-exercise soreness...............................................................................................14
3.2 Strength Loss And Recovery......................................................................................15
3.3 Post-Exercise Swelling..............................................................................................16
3.4 Range Of Motion .......................................................................................................20
3.5 Bivariate associations ...............................................................................................20
4. Discussion........................................................................................................................22
5. Conclusion.......................................................................................................................30
References...............................................................................................................................31
5. 1
1. Introduction
Eccentric exercise (EE) plays a major role in exercise training, with evidence demonstrating
significant strength benefits following a strength training programme that include this
contraction modality (Colliander and Tesch, 1990; Hortobagyi et al., 1996; Enoka, 1996;
Reeves et al., 2009; LaStoya et al., 2014). EE is common in sports such as basketball and tennis
where jumping; landing and abrupt changes of direction play important roles in performance
(Hedayatpour and Fallah, 2012).
Although beneficial in promoting strength adaptations, the occurrence of exercise-induced
muscle damage (EIMD) and the subjective experience of delayed-onset muscle soreness
(DOMS) are consequences of EE (Armstrong, 1984). DOMS is the sensation of pain or
discomfort experienced following unaccustomed muscle-lengthening exercises, which affects
individuals by reducing joint mobility and flexibility (Hough, 1902; Nosaka et al., 2006;
Goodhall and Howatson, 2008; Twist and Eston, 2005; Twist and Eston, 2009)
There is a lack of coherent research on the definitive cause of DOMS. During EE, muscle fibres
are lengthened to the point where actin and myosin protein cross-bridges become mechanically
disrupted (Flitney and Hirst, 1978). Consequently, ultra-structural changes such as
sarcolemma, plasma membrane and extracellular matrix damage occur as a result of this
mechanical disruption (Friden et al., 1983; Armstrong, 1990; Friden and Lieber, 1992;
Clarkson and Sayers, 1999; Lieber and Friden, 2002).
Loss of force production following EE has been linked with a reduction in the number of cross-
bridges available for force generation within over-stretched sarcomeres (Enoka, 1996; Morgan
and Allen, 1999; Byrne et al., 2001) and a shift in the length-tension relationship (Lieber and
Friden 1993; Wood et al., 1993; Jones et al., 1997; Child et al., 1998; Talbot and Morgan,
1998).
6. 2
Contrary to the length-tension relationship theory, strength loss immediately following EE has
been associated with excitation-contraction (E-C) coupling impairments occurring at the
interface of the t-tubules and the intracellular sarcoplasmic reticulum (Ballnave et al., 1995;
Ingalls et al., 1998; Warren et al., 1993; Warren et al., 2001; Yeung et al 2002; Balog, 2010).
Furthermore, muscle damage and strength loss observed was also attributed to reduced calcium
(Ca2+) activation of myofibrils (Balnave and Allen, 1995) and increased intracellular Ca2+
concentration due to loss of plasma membrane integrity (Armstrong, 1990; Mikkelsen et al.,
2004). Lieber et al. (1996) postulated that increased strain on muscle fibres during lengthening
exercises result in the activation of Ca2+ channels within the membrane, which initiates the
influx of Ca2+ into the plasma, subsequently increasing susceptibility to damage.
Structural damage to skeletal muscle fibres result in swelling lasting up to 10 days following
muscle-damaging exercise as fluid accumulates within the extracellular matrix and the
perimysium due to an inflammatory response (Clarkson et al., 1992; Nosaka and Clarkson,
1996). This response initiates the release of intramuscular proteins into the plasma and
production of cytokines such as tumour necrosis factor alpha (TNF-ɑ) and interleukin-6 (IL-6)
(Armstrong, 1990; Clarkson and Sayers, 1999). Asmussen (1956) hypothesised that these
cytokines may affect nerve endings and activate nociceptors creating the sensation of muscle
soreness. Furthermore, Evans and Cannon (1991) also suggested that cytokines may worsen
damage by facilitating the production of free radicals that cause loss of membrane integrity
(Clarkson and Sayers, 1999). However, contradictory research has suggested that these
cytokines promote the infiltration of lymphocytes, monocytes and neutrophils that bolster the
healing phase of muscle regeneration (Smith, 1991; Tidball, 1995; MacIntrye et al., 1996).
Increased IL-6 has been linked with acute phase inflammation and elevated production of C-
reactive protein (CRP) following skeletal injury (Northoff and Berg, 1991; Pedersen and
Hoffman-Goetz, 2000; Febbraio et al., 2002; Calder, 2006) and there is sufficient evidence
7. 3
associating DOMS with IL-6 and CRP production (Northoff and Berg, 1991; Nosaka and
Clarkson, 1996; Smith et al., 2000). Therefore, it has been suggested that minimising IL-6 and
CRP production may minimise post-exercise inflammation and pain; and promote a quicker
recovery following intense EE (Jouris et al., 2011; Houghton and Onambele, 2012).
Sport medicine and clinical professionals continue to seek and implement methods (massage,
cryotherapy, pharmacological anti-inflammatory drugs, dietary supplements, etc.) to reduce or
prevent DOMS and other symptoms of EIMD, improve recovery and facilitate performance
(see Toress et al. 2012, for review). However, some of these methods are based upon very little
evidence-based scientific research.
Inclusion of omega-3 fatty acids in a balanced diet is essential as humans are unable to naturally
produce these fats from other substances. Omega-3 fatty acids facilitate production of
hormone-like substances called prostaglandins. These are involved in the processes that lead
to symptoms of inflammation; including redness, swelling and pain (Funk, 2001). However,
Calder (2006) discussed that increased levels of prostaglandins as a result of omega-3
supplementation reduces inflammation and improves blood flow to damaged area.
Furthermore, prostaglandins produced from dietary omega-3 fatty acids were also associated
with reduced swelling, reduced sensitivity to pain, and inhibited recruitment of inflammatory
white blood cells (Maroon and Bost, 2006a).
The eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) present in omega-3 fish
oils may be effective in ameliorating DOMS and other markers of EIMD (Tartibian et al., 2009;
Tartibian et al., 2011; Jouris et al., 2011; Lemkbe et al., 2014) as it reduces inflammation; and
modulates serum CRP (Poudel-Tandukar et al., 2009) and TNF-α production (Magee et al.,
2008; Wang et al., 2008; Bloomer et al., 2009; Calder, 2013). Although there are potential
benefits associated with omega-3 fatty acids, there is a limited understanding of the required
8. 4
duration of therapy, timing relative to muscle damage incidence, and dosage of
supplementation suitable to sufficiently minimise inflammation, DOMS and its effects on
performance. Most of the underlying research on omega-3 supplementation has focused on 7
to 30 days supplementation prior to exercise.
Lenn et al. (2002) reported that supplementing omega-3 (1800mg/day) was ineffective in
ameliorating an inflammatory response following eccentric elbow flexion exercises. More
recently, Houghton and Onambele (2012) reported similar results as Lenn et al. (2002),
indicating that supplementing a dose of EPA (360mg/day) is ineffective in reducing DOMS
and other symptoms of inflammation. Findings from these two studies suggest that a much
larger dosage is necessary to reduce the effects of pro-inflammatory cytokines. Some studies
have reported decreased production of TNF-α and IL-6 in healthy humans following
supplementation of 2000mg/day or greater EPA/DHA in the form of fish oil (Endres et al.,
1989; Meydani et al., 1991; Cayghey et al., 1996; Treble et al., 2003). Furthermore, a
2700mg/day dosage of omega-3 (Lembke et al., 2014) and a 3000mg EPA/DHA (2:1) dose
(Jouris et al., 2011) were effective in reducing DOMS, CRP production and other signs of
inflammation.
It is still unclear what effects, if any, an acute 5-7day supplementation of omega-3 immediately
following exercise may have on DOMS and other indirect markers (range of motion, swelling,
isometric strength) of EIMD. The aims of the present study were therefore to investigate the
effects of different timings (prior to exercise and post-exercise) of a dose of omega-3
supplementation on DOMS and other indirect markers of EIMD and inflammation. It was
hypothesised that the pre-exercise supplementation group would report less pain, less swelling
and quicker recovery in torque and range of motion (ROM) because of the immediate
availability of a hospitable environment much more suitable for muscle recovery.
9. 5
2. Methods
2.1 Participants
Twelve convenience-sampled, 18 to 26 year old male students from Manchester Metropolitan
University Crewe campus were invited to take part in the study. All participants gave written
informed consent to take part in the study, which had ethics approval from the Manchester
Metropolitan University ethics committee. Before taking part in the study, participants
completed a questionnaire detailing health and habitual physical exercise levels. Excluded from
the study were individuals with recent physical injury to the lower extremity and individuals
clinically diagnosed with conditions requiring the use of medications likely to affect muscle
function or musculoskeletal health (Woolf and Pfleger, 2003; Lembke et al., 2014). Further
exclusion criteria included use of anti-inflammatory and/or steroid medication (four weeks
prior to baseline) (Houghton and Onambele, 2012). Participants were randomly assigned to
one of two groups: pre-exercise supplementation (PRE-EX: n = 6) and post-exercise
supplementation (POST-EX: n = 6). Participants were also advised to maintain their habitual
activity levels during the study.
2.2 Study Design
This was a controlled independent measures study lasting between 5-13 days using omega-3
supplementation. Familiarisation with gymnasium and laboratory procedures was provided for
participants prior to baseline measurements. Following the familiarisation process, participants
attended the laboratory either between 9h00 AM and 12h00 PM or 18h00 and 20h00 PM on 5
occasions. Assessments of inflammation included measures of swelling and soreness, as these
are hallmark indications of localized damage and inflammation (Friden and Lieber, 1992).
10. 6
Participants underwent baseline assessments for muscle and adipose tissue thickness, knee
ROM, muscle soreness, limb circumference (LC) and isometric strength prior to the exercise
protocol. Upon completion of baseline testing, participants performed knee flexions, knee
extensions, squats and walking lunges to induce damage and inflammation. They then returned
to the laboratory for follow-up measurements at 0-, 24-, 48-, 72- and 96hrs post exercise.
PRE-EX POST-EX
Age (y)
Height (cm)
Body mass (kg)
BMI (kg∙m-²)
21.5 (2.9)
183 (7.8)
84.8 (11.6)
25.31 (.0)
19.8 (1.60)
180.5 (7.07)
75.7 (7.07)
23.10 (.0)
BMI: body mass index
2.3 Omega-3 Supplementation
Omega-3 supplementation was administered in the form of two Minami MorEPA Platinum
1780mg softgel caps, containing in total for the 2 gels, 1528mg EPA and 472mg DHA (Minami
Nutrition, Greenford, UK). This supplementation was similar to that administered in previous
studies including Jouris et al. (2011) and Lembke et al. (2014). Participants were given clear
instructions regarding the omega-3 dosage. The PRE-EX supplementation group were asked
to take the capsules with a meal for 7 days prior to the exercise protocol whereas the POST-
EX group were asked to consume the capsules immediately following the exercise protocol
and throughout the four days of recovery. Pill counts were performed to assess compliance
Table 1. Individual characteristics.There were no significant
differences observed between the two groups (P > 0.05).Values
represent means (± SD)
11. 7
with the supplementation regimen. Participants were instructed to maintain their normal diet
and physical activity levels, while refraining from unaccustomed strenuous activity.
2.4 Exercise Protocol
The exercise protocol included leg extensions (using a leg-extension machine; Pulse 562E class
‘s’ 8/88. Pulse-fitness, Congleton, England), leg flexions (using a leg flexion machine; Pulse
562E class ‘s’ 8/88. Pulse-fitness, Congleton, England), walking lunges (using free weights)
and squats (with free weights) (Figure 1). Participants’ 1RM (for each of the four exercises)
was determined at the beginning of the exercise session by applying progressively heavier loads
until the participant was unable to fully complete a repetition (Beck et al., 2006). Additional
trials were performed with lighter loads until the 1RM was determined and a two-minute rest
period was provided between trials (Baechle, 2000). Participants were then instructed to carry
out three sets of ten repetitions at 80% of their pre-determined 1RM following a 5-minute break
to maximise the initiation of DOMS. Overall, each exercise session lasted ~60 minutes
including 1RM assessments and 3 sets of 10 repetitions of each of the four exercises. This
protocol has been previously shown to cause significant elevations in indices of muscle damage
(Houghton and Onambele, 2012).
12. 8
2.5 Muscle Soreness
Several studies have assessed the validity and reliability of the use of a visual analogue scale
as a method of measuring perceived muscle soreness (SOR) following exercise (Gallagher et
al., 2002; Bijur et al., 2001). Bijur et al (2001) reported adequate reliability with intraclass
correlation coefficient of 0.97 with 95% confidence intervals. In the present study, participants
were asked to indicate soreness level on a 10cm line following a single squat using a range of
scores representing “no pain” (1cm) and “very painful” (10cm) (Figure 2).
A B
C D
Figure 1. Resistance exercise,A - squat, B - walking lunges, C – leg
extension, D – leg flexion (Authorised use of photos from a study
participant, personal communication, March 25 2015).
13. 9
2.6 Strength Assessment
Maximum voluntary isometric contractions of the quadriceps were performed while
participants were seated on a calibrated (prior to testing) KinCom isokinetic dynamometer
(Chattanooga Group Inc., TN). Participants were positioned in 78° of hip flexion and 60° of
knee flexion, with the epicondylus laterallis of the right knee positioned so that it was aligned
to the centre of rotation of the motor arm. Knee ROM was assessed, and the appropriate
mechanical stops positioned accordingly. Straps were then positioned across the
shoulder/chest, and over the right thigh to prevent any extraneous movement. The appropriate
lever attachment was set at a relative 80% of the lower limb length distally from the lateral
condyle of the tibia. Force application was carried out against the lever arm of the dynamometer
(Drouin et al., 2004). Full knee extension was set at 0°.
Following a warm-up including five sub-maximal repetitions of knee flexions and extension
of the right lower limb, participants performed three isometric trials with 2 minutes between
efforts, with peak knee extension torque used as the participant’s strength score. Both visual
(on the computer screen of the dynamometer) and auditory feedback were provided to
Figure 2. VAS indicating soreness ratings. 0cm indicates ‘no pain’
and 10cm indicates ‘worst possible pain’.
14. 10
encourage maximal efforts. The highest of the three repeated efforts was used as the
participant’s measure of isometric strength. Strength measurements were expressed in units of
torque (Nm), which was calculated by multiplying the force (N) applied against the lever arm
by the lever arm length (m). Torque and angular position (°) on the KinCom were sampled via
an interfaced A/D system. Acquired data was transferred using the Shelton KinCom Data
Transfer Program v1.0.28 (Shelton Technical Ltd.; Milton Keynes, UK) to a Windows XP
computer (Viglen Genie, 3GHz Duo processor, 1GB Ram).
2.7 Muscle Thickness And Limb Circumference
Images of the rectus femoris (RF), vastus intermedius (VI) and sub-cutaneous adipose tissue
(AT) of the upper leg were obtained in the sagittal plane using b-mode ultrasonography (AU5,
Esaote, Genoa, Italy), with a 7.5-MHz linear phased-array probe (image depth: 53.0-93.0 mm)
applied at 50% of the femur length. Great care was taken to apply minimal pressure onto the
tissue area being scanned in order to avoid any image distortion. This method has been used in
previous studies, with great reliability (Bostock et al., 2013; Onambele et al., 2006). The 50%
point between the proximal and distal insertion of the femur were identified and marked on the
skin before images were obtained with the participant in a relaxed standing position.
Muscle thickness was measured as the distance from the top of the superficial muscle
aponeurosis to the bone (for combined RF plus VI thickness) (Figure 3). Sub-cutaneous adipose
thickness was measured as the distance from the bottom of the epidermis to the top of the
superficial muscle aponeurosis above the RF (Figure 3). These distances were measured at
three standardised points on each ultrasound frame to obtain an average muscle and sub-
cutaneous adipose thicknesses.
15. 11
Participants were asked to assume a relaxed standing position with feet shoulder width apart
as limb circumference was measured using an anthropometric measuring tape. Measurements
were taken at 25, 50 and 75% (proximal to the patella) along the distance between the proximal
and distal insertions of the femur to assess where, if any, changes in limb girth occurred. All
measurement sites used for muscle thickness and limb circumference were identified with a
semi-permanent marker by the same investigator to ensure consistent measurements between
days (Goodhall and Howatson, 2008). Previous research had indicated that using a tape
measure for limb circumference was recommended as accuracy was shown to be within 2mm
(Nosaka and Clarkson, 1996). Ultrasound and limb circumference measurements were taken
with the participant in a relaxed standing position with the knee joint consistent throughout.
Figure 3. Ultrasound image during baseline testing. D0-D2 = Rectus
Femoris thickness; D3-D5 = Vastus Intermedius thickness; D6-D8 = Sub-
cutaneous Adipose Tissue thickness.
16. 12
2.8 Range Of Motion
ROM was evaluated by measuring the difference between the flexed (F-ANG) and extended
angle (E-ANG) of the knee joint using the KinCom dynamometer. F-ANG was measured when
the participant tried to fully flex the knee to touch the gluteus muscle whilst E-ANG was
measured when the participant attempted to fully extend the leg (Nosaka and Clarkson, 1996).
2.9 Statistical Analysis
Statistical analysis was performed using IBM SPSS v21 (IBM Inc., Chicago, IL). Change
scores were calculated for soreness, muscle thickness and limb circumference by subtracting
the final values (96 hr post exercise) from the baseline values (as measured prior to EE).
Percentage change relative to baseline scores were calculated for torque and ROM by dividing
the absolute value for the day by the baseline value and multiplying by 100 (i.e. 112/115 x
100). Data normality was confirmed using a Shapiro-Wilks test due to the small sample size (n
< 50).
A mixed design repeated measures two-way analysis of variance (ANOVA) was used to
measure changes in DVs across time and between conditions (2 x 6). The ‘Within’ factor was
protocol phase which had 6 levels (baseline, 0-, 24-, 48-, 72- and 96 hrs) and the ‘between’
factor was the supplementation group with two levels (PRE-EX vs. POST-EX). Significant
interactions were followed up with Tukey post-hoc, pairwise comparisons with appropriate
Bonferonni corrections. The assumption of sphericity was tested by the Mauchly’s test of
sphericity. Where this assumption was violated, corrections were made using the Greenhouse-
Geisser adjustment to raise the critical value of F.
17. 13
In cases where tests revealed non-parametricity, such as muscle soreness data, data was log
transformed. If normality was still not achieved, then the raw data was analysed using the
Friedman’s test, followed by post-hoc Wilcoxon signed-rank test. Difference in change in
muscle soreness between the PRE-EX group and the POST-EX group were assessed using the
Kruskal-Wallis test with post-hoc Mann-Whitney tests. A p-value of ≤ 0.05 was considered
significant. All data are presented as mean ± standard deviation (SD), unless indicated
otherwise.
18. 14
3. Results
All participants completed the study. BMI was 24.24 ± 2.31 kg∙m-², reflecting that most of the
participants were lean to normal weight. Pill counts indicated a 98.5% compliance with the
supplementation regime. Mean coefficient of variance (CV) for repeated measurements in a
single day (intra-day variability) and inter-day is presented below (Table 2).
Inter-day mean CV Intra-day mean CV
Torque 16-22% 4-16%
ROM 4-7% 3-7%
Limb circumference 8-10% 0-3%
RF thickness 12-19% 2-13%
VI thickness 20-24% 5-10%
AT thickness 23-44% 4-24%
3.1 Post-exercise Soreness
Both groups showed increases in soreness between baseline; 0, 24, 48 and 72 hr following EE
(Figure 3), indicating that the exercise protocol induced leg soreness. Muscle soreness peaked
at 24 hrs in the PRE-EX group (2.33 cm) and 0 hr in the POST-EX group (3.33 cm).
Throughout the duration of the experiment, there was an observed non-significant trend for the
POST-EX group to demonstrate slightly larger ratings of muscle soreness than the PRE-EX
group. Although no significant differences were observed for soreness ratings between groups
(P > 0.05), PRE-EX supplementation was more effective in minimising muscle soreness post-
exercise (0 hr = 1.83 ± 2.14 cm, 24 hrs = 2.33 ± 2.16 cm and 48 hrs = 2.33 ± 1.97 cm) than the
POST-EX supplementation (0 h = 3.33 ± 1.51 cm, 24 h = 3.33 ± 1.86 cm and 48 h = 3.33 ±
2.34 cm) (P > 0.05).
Table 2. Analysis of Inter-day and intra-day mean coefficient of
variances (%).
19. 15
-2.0
-1.0
0.0
1.0
2.0
3.0
4.0
5.0
6.0
7.0
Baseline 0Hrs 24Hrs 48Hrs 72Hrs 96Hrs
Musclesoreness(cm)
Pre-exercise Post-exercise
3.2 Post-exercise Strength Loss And Recovery
A reduction in strength occurred immediately following EE in both groups. Maximum torque
loss was observed immediately (0 hr) after exercise in the POST-EX group (-8.23%) and 48
hrs later in the PRE-EX group (-9.7%) (P > 0.05). Torque loss at 0 hrs was significantly greater
in the PRE-EX group than in the POST-EX group (-8.23% vs -7.34%, P = 0.023, F (2.467,
24.667) = 1.930). Between groups 2-way ANOVA revealed no significant difference in torque
loss and recovery between the two groups (P > 0.05), suggesting that the two supplementation
protocols did not have a statistical significance as far as how they affected torque loss and
recovery by themselves (P > 0.05). However, there was an apparent tendency for torque to
remain 2.22% lower than baseline values in the PRE-EX group whereas the POST-EX group
were able to fully return to baseline values by the 96 hr time point (P > 0.05).
Figure 4. Comparisons ofmuscle soreness before and up to 96 h
after the damaging bout of exercise between groups. Values are
mean ± SD; N =12 (P ≤ 0.05)
20. 16
75.0
80.0
85.0
90.0
95.0
100.0
105.0
110.0
0Hrs 24Hrs 48Hrs 72Hrs 96Hrs
Torque(%baseline)
Pre-exercise Post-exercise
3.3 Post-exercise Swelling
Both groups showed increases in LC with the greatest increase occurring at the 50% point of
femur length. LC at the 50% point were highest by the 96 hr time point in the PRE-EX group
and immediately (0 hr) following exercise in the POST-EX group. At peak levels mean
difference between groups was 3.63 cm, 3.22 cm and 3.95 cm at the 25%, 50% and 75% points
respectively (P > 0.05) (Table 2). LC at the 75% point was significantly greater in the PRE-
EX group than in the POST-EX group immediately (0 hr) following exercise (P = 0.035; F
(3.170, 31.70) = 2.015). LC at the 50% and 75% point gradually subsided in the POST-EX
group by the 96 hr time point but remained elevated in the PRE-EX group (Table 2).
Furthermore, POST-EX supplementation was more effective than the PRE-EX group in
minimising swelling and facilitating recovery following exercise by the 96hr time point (-0.2
± 0.38 vs 6.29 ± 2.46; P > 0.05).
Figure 5. Comparisons ofpercentage change in torque following
damaging exercise ofbetween group * indicates significant difference
between groups (P = 0.023). Values are mean ± SD; N =12.
*
22. 18
There was a non-significant trend for an increase in muscle thickness following
damaging exercise. No significant differences were found between groups for changes in both
RF and VI (P > 0.05). The greatest change in muscle thickness was observed in the RF muscle
(Table 3). Peak RF thickness was observed at 48 and 72 hrs in the PRE-EX group (29.87 ±
5.16mm) and 96 hrs in the POST-EX group (25.35 ± 2.73 mm). POST-EX supplementation
was more effective in subsidising RF swelling by the 96 hr time point than PRE-EX
supplementation (+1.76 ± 0.72 mm vs +2.09 ± 0.91 mm; P > 0.05). Peak changes from baseline
in VI occurred at 48 hrs in the PRE-EX group (+2.29) and 0 hr following exercise in the POST-
EX group (+2.4). Change in AT thickness relative to baseline was greatest in the POST-EX
group in comparison to the PRE-EX group (+0.86 mm vs +0.54 mm). AT thickness in the PRE-
EX group was significantly greater than in the POST-EX group at 24 hrs (4.35 ± 1.13 mm vs
3.95 ± 1.45 mm; P = 0.028, F (2.832, 28.232) = 0.484). Furthermore the PRE-EX group was
more effective in minimising increases in muscle and adipose thickness from baseline values
by the 96 hr time point than the POST-EX group (P > 0.05) (Table 3).
24. 20
3.4 Post Exercise Range Of Motion
ROM decreased immediately following EE in both groups. However, no significant differences
in ROM were observed between the two groups (P > 0.05). ROM was lowest at 48 hrs in both
groups when the mean percentage decrease was approximately 5.54% in the PRE-EX group
and 6.14% in the POST-EX group (Figure 5). Decrease in ROM was significantly greater in
the PRE-EX group than the POST-EX group at the 72 hr time point (5.49% vs 3.92%; P =
0.010, F (2.519, 21.428) = 4.490). By the 96 hr time point ROM had returned to baseline level
in the POST-EX group (100.44%) and 99.11% relative to baseline in the PRE-EX group (P >
0.05).
3.5 Bivariate Associations
Bivariate correlations revealed that limb circumference was significantly associated with
changes in muscle thickness, especially in the RF throughout the experiment (r = 0.798
(baseline), r = 0.863 (0 hrs), r = 0.844 (24 hrs), r = 0.774 (48hrs); 72hrs, r = 0.808 (72 hrs);
88
90
92
94
96
98
100
102
104
0Hrs 24Hrs 48Hrs 72Hrs 96Hrs
%relativetobaseline
Pre-exercise Post-exercise
Figure 6. Comparisons ofpercentage change in ROM following
damaging exercise ofbetween group * indicates significant difference
between groups (P = 0.010).
*
25. 21
96hrs, r = 0.720 (96 hrs) P < 0.01. A significant association was reported between torque and
muscle soreness at 24 (r = -0.653) and 96 hrs (r = -0.659) P < 0.05. Furthermore, a significant
association was observed between torque and limb circumference at 50% point of femur length
throughout the experiment (r = 0.591 (baseline), r = 0.713 (0 hrs), r = 0.746 (24 hrs), r = 0.596
(48hrs); 72hrs, r = 0.834 (72 hrs); 96hrs, r = 0.592 (96 hrs) P < 0.05. However, no significant
associations were observed between torque and ROM (r < 0.03; P > 0.05); and swelling and
soreness (r < 0.05; P > 0.05) throughout the experiment.
Force ROM RF VI AT Soreness LC25 LC50 LC75
Force – Baseline r = .941**
.527 .563 .762**
-.137 .702*
-.534 .683*
.584*
p = .000 .078 .057 .004 .671 .011 .074 .014 .046
ROM- Baseline r = -.202 .282 -.633*
-.301 -.368 -.517 .453 -.531 -.513
p = .528 .375 .027 .342 .240 .085 .140 .075 .088
RF – Baseline r = .690*
.326 .883**
.664*
.259 .811**
-.431 .895**
.852**
p = .013 .301 .000 .018 .417 .001 .162 .000 .000
VI – Baseline r = .630*
.370 .509 .864**
-.182 .854**
-.636*
.667*
.540
p = .028 .236 .091 .000 .571 .000 .026 .018 .070
AT – Baseline r = -.113 -.363 .359 -.253 .868**
.223 .395 .253 .398
p = .727 .246 .251 .428 .000 .486 .203 .428 .201
Soreness - Baseline r = .077 -.379 .643*
-.141 .825**
.291 .395 .417 .600*
p = .812 .225 .024 .662 .001 .359 .204 .178 .039
LC25 – Baseline r = .700*
.264 .694*
.647*
.281 .965**
-.507 .956**
.900**
p = .011 .407 .012 .023 .376 .000 .093 .000 .000
LC50 – Baseline r = .636*
.139 .718**
.551 .443 .905**
-.280 .959**
.967**
p = .026 .666 .009 .063 .150 .000 .377 .000 .000
LC75 – Baseline r = .623*
.100 .774**
.435 .543 .825**
-.168 .930**
.987**
p = .030 .756 .003 .157 .068 .001 .602 .000 .000
Table 5. Correlations between baseline values and peak changesin indices ofDOM.
*indicates P < 0.05, ** indicates P < 0.01.
26. 22
4. Discussion
A vast amount of scientific research has provided evidence supporting the effectiveness of EPA
and DHA in preventing and treating inflammation by inhibiting production of inflammatory
inducers (Maroon and Bost, 2006, Calder, 2006; Poudyal et al., 2011). This is the first study
aimed at assessing changes in markers of EIMD and DOMS as a response to heavy EE
following two different protocols of omega-3 supplementation. Findings from this study
partially support the hypothesis that omega-3 supplementation prior to EE may have a greater
protective effect on muscle cells during EE than POST-EX supplementation. The PRE-EX
group was able to show lower ratings of soreness at 24, 48 and 72 hr time points (See Figure
3). However, secondary results showed that loss of torque in the POST-EX group was lower
than the PRE-SUP group immediately following exercise (0 hr) (Figure 4), increase in muscle
thickness and limb circumference was also lower in the POST-EX than the PRE-EX group
(Table 3 and Table 4). No difference in change in ROM was observed between the two groups
(Figure 5).
Previous studies have demonstrated that supplementing omega-3 fatty acids 7-30 days prior to
heavy EE is effective in minimising DOMS and change in other markers of EIMD (Jouris et
al., 2011, Tartibian et al., 2009; Lembke et al., 2014). Increased soreness in response to EE in
both groups in the present study is consistent with previous findings which reported significant
increases in perceived muscle soreness; typically peaking 24-48 hrs following EE (Armstrong,
1984; Friden et al., 1986; Clarkson et al., 1992; Howell et al., 1993; Warren et al., 1999;
Goodall and Howatson, 2008; Kanda et al., 2013).
Friden et al. (1986) discussed that a significant increase in intramuscular pressure associated
with increased force against the muscle components due to swelling of muscles and/or
accumulation of inflammatory fluid within the limb compartments is the main cause for
27. 23
soreness experienced. This was also confirmed in the present study as significant associations
were observed between change in soreness and change in muscle thickness, particularly in the
RF muscle (r = 0.643, P = 0.024). However, Smith (1991) and Miller et al. (2004) suggested
that increased perception of pain may be due to a combination of damage to structural proteins
within muscle fibres and an inflammatory response rather than increased intramuscular
pressure. In compliance with Armstrong (1990), Smith (1991) indicated that movement of
inflammatory substances such as cytokines and interleukins to the damaged area initiate
stimulation of sensory receptors terminating in the myofibrils. More, recently Kanda et al.
(2013) supported findings by Armstrong (1990) and Smith (1991) by reporting significant
increases in muscle soreness accompanied by increased levels of circulating neutrophils and
cytokines in healthy males following EE.
Increased omega-3 levels in healthy human tissue subsequently due to omega-3
supplementation (Bloomer et al., 2009; Poudyal et al., 2011), was associated with reduced
inflammation and DOMS (Lembke et al., 2014). Previous studies have reported significant
decreases in muscle soreness after supplementing a dose of omega-3 (324mg/d EPA + 216mg/d
DHA) for 30 days prior to EE (Tartibian et al., 2009; Tartibian et al., 2011). Similar findings
by Jouris et al. (2011) demonstrated that supplementing a daily dosage of fish oil for seven
days (2000mg/d EPA + 1000mg/d DHA) prior to EE is just as effective in minimising increases
in muscle soreness (15%) as a 30-day protocol.
In this study there was an observed non-significant trend for the PRE-EX group to demonstrate
slightly lower ratings of muscle soreness than the POST-EX group. It is possible that
differences in ratings of soreness may be associated with differences in levels of omega-3
concentration already present within the cell membranes prior to EE. Elevated concentrations
of EPA and DHA in the PRE-EX group reduces arachidonic acid content in muscle cell
membranes and inhibits the production of inflammatory inducers during EE (Calder, 2013).
28. 24
Lembke et al. (2014) hypothesised that a 30-day dose of omega-3 supplementation
(2700mg/day) prior to exercise correlated with elevated omega-3 concentration in the muscle
cell. It can be hypothesised that the PRE-EX group had an availability of greater concentrations
of EPA and DHA within the muscle cell available for immediate metabolism. This would have
resulted in greater elasticity and flexibility of muscle cell walls and a subsequent reduction in
the degree of muscle damage than the POST-EX group (Poudyal et al. 2011).
It is also possible to associate lower ratings of soreness in the PRE-EX GROUP with a greater
reduction in concentrations of circulating IL-6 and TNF-α than the POST-EX group (Bloomer
et al., 2009; Phillips et al., 2003; Trebble et al., 2003). However, Houghton and Onambele
(2012) reported that a supplementation period longer than the one used in this study (320mg/d
EPA for 3 weeks) is not sufficient to ameliorate DOMS and IL-6 mediated inflammation from
EE. Lenn et al. (2002) also reported no significant differences in participants’ perceived
soreness after supplementing fish oil (1.8g/d) for 30 days prior to exercise protocol and in the
week during the exercise protocol.
Discrepancies in IL-6 production may be due to differences in supplementation implemented.
Houghton and Onambele (2012) used isolated EPA, whereas Bloomer et al. (2009) used a
combination of EPA and DHA in the supplement groups. It should also be noted that Philips
et al. (2003) supplemented a combination of EPA, DHA, tocopherols and flavonoids, all which
play independent roles in regulation of inflammation. Mickleborough (2013) suggested that
differences in the findings between the studies (Lenn et al., 3002; Tartibian et al., 2009) may
also be due to differences in the damage protocol and the muscle groups studied. Furthermore
the difference in the dosage between the studies should be taken into account as significant
reductions in muscle soreness and swelling were reported in studies implementing a higher
dosage of omega-3 supplementation (Jouris et al., 2011; Lemkbe et al., 2014). The current
29. 25
study provides evidence that supplementing a greater than recommended dosage of omega-3
prior to EE is more effective in reducing soreness than POST-EX supplementation.
As an indicator of inflammation, increased limb circumference in both groups in the present
study is consistent with findings from previous studies which reported that swelling is a
consequence of EIMD following EE (Cleak and Eston, 1992; Clarkson et al., 1992; Howell,
1993; Nosaka and Clarkson, 1995; Nosaka and Clarkson, 1996; Chleboun et al., 1998;
Hortobagyi et al., 1998; Warren et al., 1999; Nosaka and Clarkson, 2002; Goodall and
Howatson, 2008). Greatest increase in limb circumference occurred at the 50% point of the
limb, which is in accordance with findings by Cleak and Eston (1992), who reported
significantly higher circumferences at the distal musculotendinous junction and mid-belly
biceps following EE.
In this present study, significant associations were also reported between change in LC and
muscle thickness, especially in the RF (r = 0.895, P < 0.001) which provides support for
findings by Friden et al. (1986). Friden et al. (1986) reported that increased limb size following
EE was due to swelling of muscles. Change in muscle thickness in this present study (Table 4)
is also in accordance with findings reported by Howell et al. (1993). They observed that 65%
of swelling was located within the muscle compartment rather than just fluid accumulation in
the intracellular space of the muscle fibres (Nosaka and Clarkson, 1996).
Omega-3 supplementation has been linked with reduced swelling in healthy adult men
following a 7-day (3,000 mg/d) and a 30-day (1,800mg/d) supplementation protocol prior to
EE (Tartibian et al., 2009; Jouris et al., 2011). According to findings from the present study,
both supplementation protocols are ineffective in preventing an increase in LC following heavy
EE. However, POST-EX supplementation was more effective than the PRE-EX group in
minimising swelling and facilitating recovery following EE by the 96hr time point (-0.2 ± 0.38
30. 26
cm vs 6.29 ± 2.46 cm; P > 0.05) (Table 2). The anti-inflammatory effects of elevated omega-3
content within the cells could explain the greater reduction in thigh circumference in the PRE-
EX group. However, the mechanism for this is not fully understood.
Loss of torque observed in the two groups following resistance EE in the present study is
consistent with findings from previous research (Morgan and Allen, 1999; Byrne et al., 2001;
Warren et al., 2001). Loss of force and impairments in force recovery has been associated with
increases in intracellular Ca2+ concentration due to loss of plasma membrane integrity
(Clarkson et al., 1992; Friden and Lieber, 1992; Stauber, 1989; Morgan et al., 1996; Mikkelsen
et al., 2004). However, Ingalls et al. (1998) and Yeung et al. (2002) concluded that force
decrements (57-75%) were primarily due to E-C coupling impairments in the ruptured
junctions between t-tubules and sarcoplasmic reticulum. This was subsequently confirmed by
Balog (2010), who discussed that fatigue and reduced force following EE was due to E-C
coupling impairments occurring at the interface of the t-tubules and the intracellular
sarcoplasmic reticulum.
Loss of torque was significantly greater in the PRE-SUP group than in the POST-SUP group
at 0 hr (-8.23% vs. -7.34%, P = 0.02, F (2302, 313.30) = 0.488) (Figure 5). Though there was
an apparent tendency for torque to remain 2.22% lower than baseline values in the PRE-EX
group by the 96 hr time point there were no significant differences in overall torque loss and
recovery observed between the two groups (Figure 5). From this, we can suggest that the two
supplementation protocols did not have a statistical significance as far as how they affected
torque loss and recovery by themselves (P > 0.05).
There was a significant association between soreness and torque loss, indicating that
differences in torque loss and recovery between the groups may be attributed to the difference
in soreness experienced (r = 0.702, P = 0.011). However, it should be noted that though the
31. 27
PRE-EX group reported lower ratings of soreness than the POST-EX group, they also reported
greater loss of torque and slower recovery to baseline values. Therefore, it is possible to suggest
that difference in torque loss and recovery between the two groups may be attributed to
differences in muscle activation and force generating capacity of the muscle fibres rather than
the experience of soreness (Stebbings et al., 2014). Although there were no significant
differences between individual characteristics of the two groups it is possible that the POST-
EX group may have had greater levels of activation of muscle fibres thereby resulting in a
subsequent increase in their force generating capacity (Folland and Williams, 2007).
Smith et al. (2011) proposed that there is evidence of interaction between omega-3 fatty acids
and protein synthesis in human muscle after reporting increased protein synthesis in older
adults following omega-3 supplementation. This is consistent with previous findings by Lo et
al. (1999), who reported that EPA regulates inflammation at the molecular level by decreasing
localisation of the protein complex involved in protein degradation, known as nuclear factor-b
(NF-b). We can hypothesise that there was a greater reduction of NF-b in the POST-EX group,
which provided an anabolic environment more suitable to facilitate protein synthesis required
for repair of muscles following muscle-damaging EE. Additional protein as a result of elevated
protein synthesis may result in improvement in the response to muscle damage and the
modulation of post-exercise protein balance (Koopman, 2004; Levenhagen, 2002).
As an indicator of passive muscle stiffness and soreness (Tokmakidis et al., 2003), ROM
decreased immediately following exercise in both groups. This is consistent with previous
studies that reported shortening of muscle sarcomeres and decreases of 8-25 degrees in resting
angles following EE as a result of EIMD (Clarkson and Dedrick, 1988; Cleak and Eston, 1992;
Nosaka and Clarskon, 1995; Warren et al., 1999; Goodall and Howatson., 2008). However, it
should be noted that analysis of bivariate associations revealed that there were no significant
32. 28
associations between baseline ROM and peak change in muscle soreness in the present study
(r = -.517, P > 0.05).
Chleboun et al (1998) suggested that increased swelling and myofibrils disruption following
EE results in decreased ROM due to passive muscle stiffness. More recently, Nosaka et al.
(2006) and Chen et al. (2009) also reported significant decreases in ROM immediately
following four bouts of 30 maximal isokinetic eccentric contractions of the elbow flexors.
Findings from this current study provides evidence that POST-EX supplementation of omega-
3 is more effective in facilitating recovery and preventing reductions in ROM than PRE-EX
supplementation even though soreness ratings were greater in the POST-EX group. Therefore,
it is possible to suggest that POST-EX supplementation has a greater effect on reducing passive
muscle stiffness than minimising soreness.
As a marker of inflammation, increased LC in both groups following EE in the current study
is consistent with that of previous studies which reported that swelling is a consequence of
EIMD following eccentric exercise (Cleak and Eston, 1992; Clarkson et al., 1992; Howell,
1993; Nosaka and Clarkson, 1995; Nosaka and Clarkson, 1996; Chleboun et al., 1998;
Hortobagyi et al., 1998; Warren et al., 1999; Nosaka and Newton, 2002; Goodall and
Howatson, 2008). Greatest increase in LC occurred at the 50% point of the limb, which is in
accordance with findings by Cleak and Eston (1992), who reported significantly higher
circumferences at the distal musculotendinous junction and mid-belly biceps following
eccentric exercise. However, differences in the muscle groups studied should be taking into
account as the biceps are more susceptible to greater damage than the quadriceps group.
Omega-3 supplementation has been linked with reduced swelling in healthy adult men
following a 7-day (3,000 mg/d) and 30-day (1,800mg/d) omega-3 fatty acids supplementation
prior to exercise (Tartibian et al., 2009; Jouris et al., 2011). According to findings from the
33. 29
current study, both supplementation protocols are ineffective in preventing an increase in LC
following heavy EE. LC at the 50% and 75% point gradually subsided in the POST-EX group
by the 96 hr time point but remained elevated in the PRE-EX group (Table 2). POST-EX
supplementation was more effective than the PRE-EX group in minimising swelling and
facilitating recovery following EE by the 96hr time point (-0.2 ± 0.38 cm vs 6.29 ± 2.46 cm; P
> 0.05). The decrease in LC in the PRE-EX group could be explained by the anti-inflammatory
effects of elevated omega-3 content within the cells. Bivariate correlations revealed that
changes in LC were significantly associated with changes in muscle thickness, especially in
the RF. This is consistent with findings of Howell et al. (1993), who observed that 65% of
swelling was located within the muscle compartment rather than fluid accumulation in the
intracellular space of the muscle fibres (Nosaka and Clarkson, 1996).
It should be noted that the intra-day and inter-day CVs reported in this study were considerably
higher than those in other studies (Table 2). Bostock et al. (2013) reported intra-day variances
between 3.6% and 8.9% and inter-day variances between 4.0% and 7.4% for muscle thickness
in the biceps and triceps. Bemben (2002) also reported CVs for muscles measured using
ultrasonography between 3.5-6.75%. Houghton and Onambele (2012) reported intra-day CV
between 0-2.7% and inter-day CVs between 1.5% and 1.75% for isometric strength
measurements. The difference in the coefficient of variances between this study and those in
other research studies may be associated with the effect of unfamiliarity with the equipment
used in the study. Further training with equipment is recommended to minimise the difference
in CVs.
34. 30
5. Conclusion
These findings suggest that timing of omega-3 supplementation may play a role in response to
EE. PRE-EX supplementation has a greater effect on minimising soreness whereas POST-EX
supplementation proved more effective in minimising swelling, strength loss and decrements
in ROM following EE. These findings would support recommendations to include omega-3
fatty acids as part of a healthy diet both prior to EE and throughout the recovery period
following EE to minimise inflammation and facilitate recovery from heavy EE.
However, findings from this current study is limited due to the small sample size given the time
requirement of the supplementation protocol and possible differences in adaptations to EE
between the two groups. Furthermore, participation in physical activity that may induce further
inflammation during the recovery period could have been followed up. This would minimise
the possibility of attributing difference in response to EE and recovery between the two groups
to their physical activity during the recovery period. Further in-vivo work is required to
investigate the biochemical interaction between omega-3 fatty acids and mediators of
inflammation in both supplementation protocols with aims of providing more conclusive data
to help understand the differences in the inflammatory response to EE.
Word count: 6448
35. 31
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