Content uploaded by Amanda J Salacinski
Author content
All content in this area was uploaded by Amanda J Salacinski on Jun 06, 2014
Content may be subject to copyright.
Available via license: CC BY 2.0
Content may be subject to copyright.
R E V I E W Open Access
Protein timing and its effects on muscular
hypertrophy and strength in individuals engaged
in weight-training
Matthew Stark
1
, Judith Lukaszuk
1*
, Aimee Prawitz
1
and Amanda Salacinski
2
Abstract
The purpose of this review was to determine whether past research provides conclusive evidence about the effects
of type and timing of ingestion of specific sources of protein by those engaged in resistance weight training. Two
essential, nutrition-related, tenets need to be followed by weightlifters to maximize muscle hypertrophy: the
consumption of 1.2-2.0 g protein.kg
-1
of body weight, and ≥44-50 kcal
.
kg
-1
of body weight. Researchers have
tested the effects of timing of protein supplement ingestion on various physical changes in weightlifters. In general,
protein supplementation pre- and post-workout increases physical performance, training session recovery, lean
body mass, muscle hypertrophy, and strength. Specific gains, differ however based on protein type and amounts.
Studies on timing of consumption of milk have indicated that fat-free milk post-workout was effective in promoting
increases in lean body mass, strength, muscle hypertrophy and decreases in body fat. The leucine content of a
protein source has an impact on protein synthesis, and affects muscle hypertrophy. Consumption of 3–4gof
leucine is needed to promote maximum protein synthesis. An ideal supplement following resistance exercise
should contain whey protein that provides at least 3 g of leucine per serving. A combination of a fast-acting
carbohydrate source such as maltodextrin or glucose should be consumed with the protein source, as leucine
cannot modulate protein synthesis as effectively without the presence of insulin. Such a supplement post-workout
would be most effective in increasing muscle protein synthesis, resulting in greater muscle hypertrophy and
strength. In contrast, the consumption of essential amino acids and dextrose appears to be most effective at
evoking protein synthesis prior to rather than following resistance exercise. To further enhance muscle hypertrophy
and strength, a resistance weight- training program of at least 10–12 weeks with compound movements for both
upper and lower body exercises should be followed.
Keywords: Protein timing, Muscular hypertrophy, Muscular strength, Body composition, Whey protein, Milk protein,
Protein synthesis
Review
Purpose
Individuals who engage in resistance weight training,
whether as competitive weightlifters or to promote opti-
mal physical outcomes, would benefit by knowing the
ideal nutritional intake protocol needed to maximize
muscle hypertrophy and strength. The type, timing (pre/
post workout) or amount of protein intake required to
meet strength-training goals may not be clear to
weightlifters or their trainers. The purpose of this review
was to determine whether past research provides conclu-
sive evidence about the effects of type and timing of
ingesting specific protein sources by those engaged in
resistance weight training. The review targets the effects
of intake and timing of the following protein sources on
physical outcomes: whey, casein, milk, soy and essential
amino acids.
Protein and calorie intake
For maximal muscle hypertrophy to occur, weightlifters
need to consume 1.2-2.0 grams (g). protein kilogram.
(kg)
-1
and > 44–50 kilocalories (kcal)
.
kg
-1
body weight
* Correspondence: jmlukaszuk@niu.edu
1
School of Family, Consumer, and Nutrition Sciences. Northern Illinois
University, DeKalb, IL, USA
Full list of author information is available at the end of the article
© 2012 Stark et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative
Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and
reproduction in any medium, provided the original work is properly cited.
Stark et al. Journal of the International Society of Sports Nutrition 2012, 9:54
http://www.jissn.com/content/9/1/54
daily [1-9]. This is considerably higher than the recom-
mended dietary allowance (RDA) for protein (currently
0.8 g
.
kg
-1
) which meets the needs of 97.5% of all healthy
adult Americans not engaged in weightlifting with the
intent of gaining muscle mass [8]. Table 1 summarizes
ranges for protein intake for weightlifters based on pre-
vious literature reviews.
Leucine and muscle protein synthesis
The leucine content of a protein source has an impact
on protein synthesis, and affects muscle hypertrophy
[10-15]. This section details the role of leucine in protein
synthesis to illustrate its importance in the process.
Protein synthesis occurs when methionyl-transfer ribo-
nucleic acid (methionyl-tRNA) binds to a eukaryotic
small ribosomal subunit (40S ribosomal unit) resulting
in the formation of a pre-initiation complex (43S pre-
initiation complex) [16]. This initial step is mediated by
eukaryotic initiation factor 2 (eIF2) [16]. The 43S com-
plex subsequently binds to messenger ribonucleic acid
(mRNA) near the cap structure. After successful engage-
ment of the 43S pre-initiation complex to RNA, the
molecule eukaryotic initiation factor 5 (eIF5) removes
eIF2 while a molecule of guanosine triphospahte (GTP)
is hydrolyzed so that eIF2 is recycled to its active form
of eIF2-GTP [16]. This allows eIF2-GTP to continue
with the initial step of protein synthesis. Once eIF2-GTP
is released, the second step can occur. A ribosomal bind-
ing site/translation start site forms once eukaryotic initi-
ation factor 4F (eIF4F) recognizes the molecule [16].
The eIF4F complex binds the eukaryotic initiation factor
4E (eIF4E) subunit of eIF4F to the m
7
GTP cap structure
present in all eukaryotic mRNAs [16]. Replication of the
mRNA strand occurs, thus indicating protein synthesis.
The processes of protein synthesis appear to be highly
regulated by the amino acid leucine [10-14].
Leucine plays a role in muscle protein synthesis mostly
through stimulation of the mammalian target of rapa-
maycin (mTOR) signaling pathway [15,17,18]. Leucine
interacts with two mTOR regulatory proteins, mTOR
raptor (or raptor) and rashomolog enriched in the brain
(or Rheb) [19,20]. The importance of the regulation of
mTOR is that when activated, it phosphorylates the
proteins eIF4E binding protein 1 (4E-BP1) and riboso-
mal protein S6 kinase (S6K1) complex [21,22]. When
4E-BP1 is phosphorylated, it becomes inactive, which
allows the continuation of the second step initiation
phase of translation by inhibiting its binding to eIF4F
complex [10]. This allows additional translation to occur.
When S6K1 is phosphorylated, it produces additional
eIFs which increases the translation of mRNAs that en-
code components of the protein synthesis pathway
[10,12].
Leucine has been indicated as the sole stimulator of
protein synthesis [10-15]. For example, Dreyer et al. con-
ducted a study on 16 young, healthy untrained men to
determine the effects of post-workout consumption of
either no beverage or leucine-enhanced EAAs [15].
Those consuming the leucine-enhanced beverage one
hour following a single bout of resistance exercise had
greater rates of protein synthesis than did the control
group. Another study conducted by Koopman et al. [23]
concurs with the findings of Dreyer. Eight untrained
men were randomly assigned to consume one of the
three beverages: carbohydrates, carbohydrate and pro-
tein or carbohydrate, protein and free leucine following
45 minutes of resistance exercise. The results indicated
that whole body net protein balance was significantly
greater in the carbohydrate, protein and leucine group
compared with values observed in the carbohydrate and
protein and carbohydrate only groups, indicating the
ability of leucine to augment protein synthesis [23].
Leucine alone appears to be nearly as effective in
stimulating protein synthesis as when all branched chain
amino acids (BCAAs) are consumed [24-26]. Leucine
also seems to have both insulin-dependent and insulin-
independent mechanisms for promoting protein synthe-
sis [27,28]. Approximately 3 to 4 g of leucine per serving
is needed to promote maximal protein synthesis [29,30].
See Table 2 for the leucine content of protein sources
for all protein ingestion timing studies referenced in this
review.
Types of protein
There are numerous protein sources available to the
consumer. This review article focuses on studies that
Table 1 Summary of protein requirements for weightlifters
Research study Recommendation for protein intake Type of study
Lemon [1] 1.6-1.7 g
.
kg
-1
Review of literature
Lemon et al. [2] 12-15% total energy intake Review of literature
Kreider [3] 1.3-1.8 g
.
kg
-1
Review of literature
Phillips [4] 12-15% total energy intake Review of literature
Lemon [5] 1.6-1.8 g
.
kg
-1
Review of literature
Lemon [6] 1.5-2.0 g
.
kg
-1
Review of literature
Campbell et al. [7] 1.4-2.0 g
.
kg
-1
Review of literature
Stark et al. Journal of the International Society of Sports Nutrition 2012, 9:54 Page 2 of 8
http://www.jissn.com/content/9/1/54
have used a variety of dairy- and soy-based protein
sources. This section describes each of these protein
sources and compares their quality on the two scales
most relevant to this review: biological value and protein
digestibility corrected amino acid score (PDCAAS) [44].
Biological value (BV), determines how efficiently exogen-
ous protein leads to protein synthesis in body tissues
once absorbed, and has a maximum score of 100 [44].
PDCAAS numerically ranks protein sources based on
the completeness of their essential amino acid content,
and has a maximum score of 1.0 [44]. The BV and
PDCAAS are both important in understanding bioavail-
ability and quality of different protein sources.
Three sources of dairy protein typically used in studies
of muscle hypertrophy and strength are bovine milk, ca-
sein and whey. Bovine milk is a highly bioavailable
source of protein, comprising 80% casein and 20% whey
[44]. Overall, bovine milk has a BV of 91 and a PDCAAS
of 1.00 indicating that it is readily absorbed by the body,
promoting protein synthesis and tissue repair, and pro-
vides all essential amino acids (EAAs). Casein, with a BV
of 77 and a PDCAAS of 1.00, is the predominate protein
in bovine milk and gives milk its white color [44]. It
exists in micelle form, and within the stomach will gel
or clot, thus resulting in a sustained release of amino
acids [45]. Compared with milk, it is less bioavailable,
but like milk, it provides all EAAs. Whey the other
protein found in milk, is the liquid part of milk that
remains after the process of cheese manufacturing [44].
With a BV of 104 and a PDCAAS of 1.00, whey is super-
ior to both milk and casein. It contains all EAAs, and its
excellent bioavailability leads to rapid protein synthesis
[44,45].
Soy is a vegetable-based protein source that is useful
for vegetarians and individuals who are lactose- or ca-
sein-intolerant. Soy has a BV of 74 and PDCAAS of
1.00, indicating that it is not as bioavailable as milk
based protein, but does contain all EAAs [44].
Whole-food protein intake studies: post workout only
The timing of protein intake has been an important con-
dition in studies on muscle hypertrophy and strength in
weight-trained individuals. In this section, studies using
whole-food protein sources (i.e. bovine and soy milk)
have been reviewed with respect to their intake following
weight-resistance training.
Many studies on the effects of protein intake timing on
physical changes have used protein supplements [31-36],
but some studies have used milk and other fluid pro-
tein sources. In a study focused on protein intake fol-
lowing a single resistance training session, Elliot et al.
examined milk consumption post-workout in 24 un-
trained men and women [37]. Subjects were randomly
assigned to one of three groups: 237 g of fat-free milk,
Table 2 Leucine content of protein sources for studies that used a protein ingestion timing method
Research
study
Protein used Leucine
content
Reached 3g
Threshold for Leucine
Hoffman
et al. [31]
42 g of a proprietary blend of protein (enzymatically hydrolyzed collagen protein isolate,
whey protein isolate, and casein protein isolate)
3.6 g Yes
Hoffman
et al. [32]
42 g of a proprietary protein blend (enzymatically hydrolyzed collagen protein isolate, whey
protein isolate, casein protein isolate, plus 250 mg of additional branch chain amino acids)
3.6 g Yes
Cribb et al.
[33]
Whey protein, creatine and dextrose mixture based on individuals bodyweight 3.49 g
1
Yes
Verdijk et al.
[34]
20 g of casein split into two 10 g servings pre- and post-workout 1.64 g total in 2
servings
2
No
Hulmi et al.
[35]
30 g whey split into two 15 g servings pre- and post-workout 3.4 g total in 2
servings
No as only 1.7 g were
given at a time
Andersen
et al. [36]
25 g of a protein blend (16.6 g of whey protein; 2.8 g of casein; 2.8 g of egg white protein;
and 2.8 g of l-glutamine)
2.29 g
2,3
No
Elliot et al.
[37]
237 g of whole milk 0.639 g No
Hartman
et al. [38]
500 mL of fat-free milk 1.35 g No
Wilkinson
et al. [39]
500 mL of fat-free milk 1.35 g No
Rankin et al.
[40]
Chocolate milk based on bodyweight Unknown Unknown
Josse et al.
[41]
500 mL of fat-free milk 1.35 g No
1
3.49 g is based on the amount of leucine that the mean weight (80 kg) of the participants in this study.
2
Leucine content of casein received from Tang et al. [42].
3
Leucine content of egg white received from Norton et al. [43].
Stark et al. Journal of the International Society of Sports Nutrition 2012, 9:54 Page 3 of 8
http://www.jissn.com/content/9/1/54
237 g of whole milk, or 393 g of isocaloric fat-free milk.
The findings indicated that in untrained individuals,
threonine uptake was significantly higher for those con-
suming 237 g whole milk versus those consuming 237 g
fatfreemilk.Threonineuptakeisindicativeofnet
muscle protein synthesis. The results of this study
suggest that whole milk increased utilization of avail-
able amino acids for protein synthesis [37]. Tipton et al.
conducted a study on 23 untrained men and women in
which participants ingested 1) 20 g casein, 2) 20 g whey,
or 3) artificially sweetened water one hour following heavy
leg resistance exercise [46] Positive changes in net muscle
protein balance resulted for both protein groups but not
for the control group. This study indicated that milk pro-
teins (both casein and whey) post-workout increased pro-
tein synthesis [46].
Various studies have compared whole-food protein
sources to determine which is most effective in improv-
ing muscle mass and strength gains. Hartman et al. con-
ducted a study comparing the use of milk, soy protein,
or carbohydrate drinks by 56 young untrained males
[38]. Subjects were assigned to one of three groups; each
consumed 500-milliliter (mL) of a) fat-free milk, b) an
isocaloric, isonitrogenous, and macronutrient- matched
soy-protein beverage, or c) an isocaloric carbohydrate
beverage immediately following and again one hour after
resistance exercise. Body composition, muscle hyper-
trophy, and strength measurements were recorded at
baseline and three days following 12 weeks of training 5
d.wk
-1
. The group using milk post-workout had signifi-
cantly increased body weight and decreased body fat ver-
sus the other two groups, indicating an increase in lean
body mass (LBM). Results indicated that consumption of
fat-free milk post-workout was statistically more effect-
ive than soy protein in promoting increases in LBM
(p<0.01), increases in type II muscle fiber area (p<0.05)
and decreases in body fat (p<0.05) [38]. These results
were similar to those found by Wilkinson et al. [39].
Researchers assigned eight weight-trained men to either
500 mL of skim milk or an isonitrogenous, isocaloric,
and macronutrient-matched soy-protein beverage fol-
lowing resistance exercise [39]. A crossover design was
used so that all participants consumed either milk or soy
on their first trials and alternated to the other supple-
ment on the second trials. Trials were separated by one
week. Both protein drinks increased protein synthesis
and promoted increases in muscle mass; however, the
consumption of skim milk had a significantly greater im-
pact on the development of muscle mass than did con-
sumption of the soy protein [39]. Both Hartman et al.
[38] and Wilkinson et al. [39] demonstrated the super-
iority of milk proteins over soy protein in building
muscle mass. This may be due to the fact that soy has a
lower BV than milk (74 versus 91 respectively), resulting
in lower bioavailability, thus providing less protein syn-
thesis in body tissues.
Rankin et al. studied the effects of milk versus carbo-
hydrate consumption post-resistance exercise on body
composition and strength [40]. Nineteen untrained men
were randomly assigned to one of two groups that pro-
vided 5 kcal
.
kg
-1
body weight of either chocolate milk, or
a carbohydrate-electrolyte beverage. Subjects completed
whole body dual-energy X-ray absorptiometry (DXA)
scans and strength assessments prior to and after follow-
ing a 3 d
.
wk
-1
for 10-weeks weightlifting protocol.
Results indicated that both groups had increases in LBM
and strength, but there were no significant between-
group differences [40]. The addition of a control group
to this study would have helped determine whether
increases in strength were due solely to the weightlifting
program or to the combination of exercise and supple-
mentation. The findings suggest that consumption of
chocolate milk post-exercise may be effective in increas-
ing LBM in weightlifters, but more studies using control
groups are needed.
Milk consumption and resistance training also have
been investigated in women. Josse et al. examined the
effects of milk consumption post-workout on strength
and body composition in 20 healthy untrained women
[41]. Subjects were assigned to 500 mL of either fat-free
milk or isocaloric maltodextrin. The women followed a
weight training protocol 5 d
.
wk
-1
for 12-weeks. Each par-
ticipant completed strength assessments, DXA scans,
and blood tests. The group consuming milk had statisti-
cally greater increases in LBM, greater fat mass losses
and greater gains in strength, providing evidence that
fat-free milk consumption post-workout was effective in
promoting increased LBM and strength in women
weightlifters [41]. The results of this study support those
of previous studies completed in men showing that milk
consumption post-workout has a favorable effect on
MPS [37-40].
Protein supplement intake studies: a comparison of
timing protocols
Protein and amino acid supplements have been used
widely in studies showing their effectiveness on protein
synthesis. Hoffman et al. compared protocols providing
protein supplementation and subsequent effects on
muscle strength and body composition in 33 strength-
trained adult men [31]. Two protein-intake timing strat-
egies were implemented over the course of 10-weeks of
resistance weight-training [31]. One group consumed a
protein supplement comprising enzymatically hydro-
lyzed collagen-, whey-, and casein-protein isolates pre/
post-workout. A second group consumed the same sup-
plement in the morning upon awakening and in the
evening. A control group was not given the protein
Stark et al. Journal of the International Society of Sports Nutrition 2012, 9:54 Page 4 of 8
http://www.jissn.com/content/9/1/54
blend. The average caloric intake of the three groups
was 29.1 ± 9.7 kcal
.
kg body mass-
1.
d
-1
.
Muscle strength was assessed through one-repetition
maximum (1RM) on bench and leg press. Body compos-
ition was assessed using DXA [31]. There were no group
differences in body composition based on timing of sup-
plementation [31]. All groups increased the 1RM for
squats, indicating increased muscle strength. Only the
protein supplement groups also showed significant
increases in the 1RM for bench press, indicating
improved strength [31]. These findings indicated that
supplementation was beneficial for increasing muscle
strength in 1 RM bench press but timing of ingestion
was not important. The results on body composition
may have had different effects if participants had con-
sumed adequate kcal
.
kg
-1
, as greater-than-maintenance-
caloric needs are required for muscular hypertrophy to
occur. Strength did increase, providing evidence to both
the effectiveness of protein supplementation on strength
and the effectiveness of the workout regimen used in
this study. Future studies should ensure that participants
are consuming greater than 44–50 kcal
.
kg
-1
to maximize
muscle hypertrophy [9].
Hoffman et al. conducted a double-blind study focus-
ing on the use of protein supplements to hasten recovery
from acute resistance weight training sessions [32]. Fif-
teen strength-trained men were matched for strength
then randomly assigned to receive 42 g of either a) a
proprietary protein blend (enzymatically hydrolyzed col-
lagen-, whey-, or casein-protein isolates, plus 250 mg of
additional BCAA pre and post workout), or b) a placebo
of maltodextrin pre-and post-workout [32]. Participants
initially performed a 1RM for squat, dead lift, and bar-
bell lunge exercises. On the second visit, subjects per-
formed four sets of at least 10 repetitions at 80% of their
1RM for the exercises with 90 seconds between sets. On
visits three (24 hours from visit two) and four (48 hours
from visit two), participants performed four sets of
squats with the previous weight and performed as many
repetitions per set as possible [32]. Hoffman et al. [32]
found that the group receiving the proprietary protein
blend performed significantly more repetitions at visits
three and four than did subjects receiving the placebo.
These findings provide evidence that protein supplemen-
tation pre- and post-workout is useful in maximizing
weight-training performance, as well as in hastening ex-
ercise recovery 24 and 48 hours post-exercise.
Timing of supplementation in relation to the resist-
ance workout also has been studied [33]. Cribb et al.
assigned 23 male bodybuilders to one of two groups:
those who received a supplement a) before and after a
workout, or b) in the morning and evening. The sup-
plement contained 40 g protein (from whey isolate),
43 g carbohydrate (glucose), and seven g creatine
monohydrate per 100 g. Each participant was given
the supplement in quantities of 1.0 g
.
kg
-1
body weight.
All participants followed a preliminary resistance
weight-training program for 8–12 weeks before base-
line measurements were taken. Participants then
started the 10-week resistance weight-training session
which was divided into three distinct stages: prepara-
tory (70–75% 1RM), overload phase 1 (80–85%1RM),
and overload phase 2 (90–95% 1RM) [33].
Results indicated significant differences in body com-
position in the group consuming the supplement pre-
and post-workout [33]. This group experienced
increased LBM and decreased body fat. Both groups
demonstrated increases in strength, but the pre- and
post-workout group demonstrated significantly greater
gains [33], indicating that timing of the ingestion of the
protein supplement was crucial. This is contradictory to
the findings of Hoffman et al. [31] with respect to
changes in body composition. This could be because
Cribb et al. [33] used a supplement that was a combin-
ation of protein, carbohydrate and creatine whereas,
Hoffman et al. [31] supplemented with protein only. The
major finding of this study was that after 10 weeks of
training, supplementation pre/post each workout
resulted in greater improvements in 1RM strength and
body composition (increased LBM and decreased body
fat percentage) compared with a matched group who
consumed supplement in the morning and evening, out-
side of the pre- and post-workout time frames.
The majority of studies of protein intake and resist-
ance exercise have been conducted on younger adult
males [31-33]. In contrast, Verdijk et al. investigated
the impact of the protein, casein hydrolysate, on
muscle hypertrophy in healthy untrained elderly men
[34]. Researchers randomly assigned 28 elderly men to
consume either a protein supplement or a placebo
pre- and post-workout. Subjects performed a 12-week
resistance weight-training program requiring weightlift-
ing 3 d
.
wk
-1
. Baseline and ending measurements were
obtained, including strength assessments, CT scans,
DXA scans, blood samples, 24-hour urine samples,
muscle biopsies, and immunohistochemistry tests.
Results indicated no differences in ending measure-
ments between the protein group and placebo group
in muscle hypertrophy, strength, or body composition
[34], suggesting that for elderly men, intake of 20 g ca-
sein hydrolysate before and after resistance training
does not increase muscle hypertrophy or strength. In
this study, however, only 20 g of casein was used, and
it was divided into two servings. This protocol would
not have provided participants with the required 3 g
of leucine needed to maximize protein synthesis. Add-
itionally, since casein is slow digesting [44,45], it may
not have been ideal for use in a study of elderly men.
Stark et al. Journal of the International Society of Sports Nutrition 2012, 9:54 Page 5 of 8
http://www.jissn.com/content/9/1/54
Future studies with this population should incorporate
whey protein, which is highly bioavailable in an
amount that would provide at least 3 g leucine [29,30].
Studies comparing the effects of supplementation with
adequate protein and those with creatine-enhanced
protein pre-and post-workout also should be con-
ducted to determine whether creatine is needed to
produce the desired outcomes, as has been demon-
strated in younger men [33] (See Table 2).
The long-term use of whey protein pre- and post-
resistance exercise was investigated by Hulmi et al.
[35], by assigning participants to one of three
groups:1) 15 g of whey protein before and after resist-
ance exercise, 2) a placebo before and after resistance
exercise, or 3) no supplement no participation in
weightlifting but continued habitual exercise as they
did prior to the study. Participants in the first two
groups completed two resistance exercise sessions per
week for 21 weeks consisting of both upper and lower
body multi-joint lifts. All participants then had biopsies
performed on their vastus lateralis. Results indicated
that the whey protein group had significantly greater
increases than the other groups in vastus lateralis
hypertrophy, and greater overall muscle hypertrophy
[35]. These findings provide evidence that whey pro-
tein supplementation pre- and post-workout is useful
in increasing muscle hypertrophy.
Andersen et al. examined the effects of a mixed
blend of proteins on muscle strength and muscle fiber
size [36]. They studied the ingestion pre- and post-
workout of 25 g of a protein blend (whey, casein, egg-
white proteins, and l-glutamine), compared with a
maltodextrin supplement, over the course of a 3 d
.
wk
-1
14-week resistance-training program. Results of muscle
biopsies from the vastus lateralis indicated that the
protein supplementation group had greater increases
in muscle hypertrophy and in squat jump height [36].
Results of this study provide evidence that supple-
mentation with a blend of whey, casein, egg-white
proteins, and l-glutamine pre- and post-workout helps
promote muscle hypertrophy and improved physical
performance.
Training effects
The effects of training protocols also are very import-
ant on increases in strength and muscle hypertrophy.
All studies used in this review followed a resistance
weight-lifting protocol [31-36,38-41]. It appears from
the studies referenced in this review that a training
protocol tailored for muscle hypertrophy and strength
should be at least 10–12 weeks in length and involve
three to five training sessions weekly, consisting of
compound lifts that include both the upper and lower
body [31,33,35,36,38,40,41].
Conclusions
Researchers have tested the effects of types and timing
of protein supplement ingestion on various physical
changes in weightlifters. In general, protein supplemen-
tation pre- and/or post-workout increases physical per-
formance [31-34,38-41], training session recovery [32],
lean body mass [33,38-41], muscle hypertrophy [35,38-41],
and strength [31,33,38,40,41]. Specific gains, however, differ
based on protein type and amounts [31-36]. For example,
whey protein studies showed increases in strength [31,33],
whereas, supplementation with casein did not promote
increases in strength [34]. Additional research is needed on
the effects of a protein and creatine supplement consumed
together, as one study has shown increases in strength
and LBM [33].
Studies on timing of milk consumption have indicated
that fat-free milk post-workout was effective in promot-
ing increases in lean body mass, strength, muscle hyper-
trophy and decreases in body fat [38-41] Milk proteins
have been shown to be superior to soy proteins in pro-
moting lean body mass [38] and muscle mass develop-
ment [39]. What is interesting about the milk studies
[38-41] is that not one of them provided the 3–4gof
leucine needed to promote maximal MPS (See Table 2),
yet they all showed improvements in LBM and strength.
This raises the question of whether other components in
milk could have contributed to the changes observed.
Future researchers should investigate whether other
properties of milk help increase LBM when leucine in-
take is suboptimal to provide maximal MPS. Researchers
should also investigate the effects of protein supple-
ments when participants are consuming adequate kcal
.
kg
-1
and g
.
kg
-1
of protein to maximize muscle
hypertrophy.
The effects of timing of ingestion of EAAs on physical
changes following exercise also have been studied
[47,48]. Tipton et al. [47] found that the ingestion of
EAAs prior to resistance exercise was more beneficial
than post-ingestion in promoting protein synthesis [47],
but these results did not hold true with respect to whey
protein ingestion [48]. Once a protein has been con-
sumed by an individual, anabolism is increased for about
three hours postprandial with a peak at about 45–90
minutes [14]. After about three hours postprandial, MPS
drops back to baseline even though serum amino acid
levels remain elevated [14]. These data show that there
is a limited time window within which to induce protein
synthesis before a refractory period begins. With this in
mind, an ideal protein supplement after resistance exer-
cise should contain whey protein, as this will rapidly di-
gest and initiate MPS, and provide 3–4 g of leucine per
serving, which is instrumental in promoting maximal
MPS [29,30]. A combination of a fast-acting carbohy-
drate source such as maltodextrin or glucose should be
Stark et al. Journal of the International Society of Sports Nutrition 2012, 9:54 Page 6 of 8
http://www.jissn.com/content/9/1/54
consumed with the protein source, as leucine cannot
modulate protein synthesis as effectively without the
presence of insulin [27,28] and studies using protein
sources with a carbohydrate source tended to increase
LBM more than did a protein source alone [33,37-41].
Such a supplement would be ideal for increasing muscle
protein synthesis, resulting in increased muscle hyper-
trophy and strength. In contrast, the consumption of es-
sential amino acids and dextrose appears to be most
effective at evoking protein synthesis prior to rather than
following resistance exercise [47]. To further enhance
muscle hypertrophy and strength, a resistance weight-
training program of at least 10–12 weeks 3–5d
.
wk
-1
with compound movements for both upper and lower
body exercises should be followed [31,33,35,36,38,40,41].
Abbreviations
ml: Milliliter; d: Day; g: Gram; kg: Kilogram; wk: Week; RDA: Recommended
dietary allowance; LBM: Lean body mass; MPS: Maximal protein synthesis;
1RM: One-repetition maximum; DXA: Dual-energy X-ray absorptiometry;
CT: Computed tomography; kcal.kg
-1
: Kilocalories per kilogram; g.kg
-1
: Gram
per kilogram; BCAA: Branched chain amino acids; EAA: Essential amino acids;
Hr: Hour; methionyl-tRNA: Methionyl-transfer ribonucleic acid; 40S: Eukaryotic
small ribosomal subunit; 43S: Pre-initiation complex; eIF2: Eukaryotic initiation
factor 2; mRNA: Messenger ribonucleic acid; GTP: Guanosine triphosphate;
eIF4F: Eukaryotic initiation factor 4F; eIF4E: Eukaryotic initiation factor 4E;
mTOR: Mammalian target of rapamycin; raptor: mTOR raptor;
Rheb: Rashomolog enriched in the brain; 4E-BP1: eIF4E binding protein 1;
S6K1: Ribosomal protein S6 kinase.
Competing interests
The authors declare that they have no competing interests.
Authors' contributions
MS was the primary author of the manuscript. JL, AP and AS played an
important role in manuscript preparation and revisions. All authors have read
and approved the final manuscript.
Author details
1
School of Family, Consumer, and Nutrition Sciences. Northern Illinois
University, DeKalb, IL, USA.
2
The Department of Kinesiology and Physical
Education, Northern Illinois University, DeKalb, IL, USA.
Received: 25 July 2012 Accepted: 10 December 2012
Published: 14 December 2012
References
1. Lemon P: Effects of exercise on dietary protein requirements. Int J Sport
Nutr 1998, 8:426–447.
2. Lemon PW, Proctor DN: Protein intake and athletic performance.
Sports Med 1991, 12:313–325.
3. Kreider R: Effects of protein and amino acid supplementation on athletic
performance. Sportscience 1999, 3(1). sportsci.org/jour/9901/rbk.html.
4. Phillips SM: Protein requirements and supplementation in strength
sports. Nutrition 2004, 20:689–695.
5. Lemon PW: Beyond the zone: protein needs of active individuals. JAm
Coll Nutr 2000, 19(Suppl):513S–521S.
6. Lemon PW: Protein requirements of strength athletes. In Sports
Supplements. Edited by Antonio J, Stout J. Philadelphia, PA: Lippincott,
Williams, & Wilkins Publishing Co; 1996.
7. Campbell B, Kreider R, Ziegenfuss T, Bounty P, Roberts M, Burke D, Landis J,
Lopez H, Antonio J: International society of sports nutrition position
stand: protein and exercise. J Int Soc Sports Nutr 2007, Available at:
http://www.jissn.com/content/4/1/8.
8. Gropper S, Smith J, Groff J: Protein. In Advanced Nutrition and Human
Metabolism. 5th edition. California: Wadsworth Cengage Learning;
2009:179–250.
9. American Dietetic Association, Dietitians of Canada, & American College of
Sports Medicine: Position stand: nutrition and athletic performance.
Med Sci Sports Exerc 2000, 32:2130–2145.
10. Kimball SR, Jefferson LS: New functions for amino acids: effects on gene
transcription and translation. Am J Clin Nutr 2006, 83:500S–507S.
11. Anthony JC, Anthony TG, Kimball SR, Vary TC, Jefferson LS: Orally
administered leucine stimulates protein synthesis in skeletal muscle of
postabsorptive rats in association with increased eIF4F formation.
J Nutr 2000, 130:139–145.
12. Anthony JC, Yoshizawa F, Anthony TG, Vary TC, Jefferson LS, Kimball SR:
Leucine stimulates translation initiation in skeletal muscle of
postabsorptive rats via a rapamycin-sensitive pathway. J Nutr 2000,
130:2413–2419.
13. Norton L, Layman D, Garlick P: Isonitrogenous protein sources with
different leucine contents differentially effect translation initiation and
protein synthesis in skeletal muscle. FASEB J 2008, 22:869–875.
14. Norton L, Layman D, Bunpo P, Anthony T, Brana D, Garlick P: The Leucine
content of complete meal directs peak activation but not duration of
skeletal muscle protein synthesis and mammalian target of rapamycin
signaling in rats. J Nutr 2009, 139(6):1103–1109.
15. Dreyer H, Drummond, Pennings B, Fujita S, Glynn E, Chinkes D, Dhanani S,
Volpi E, Rasmussen B: Leucine-enriched essential amino acid and
carbohydrate ingestion following resistance exercise enhances mTOR
signaling and protein synthesis in human muscle. Am J Physiol Endocrinol
Metab 2008, 294:E392–E400.
16. Stipanuk M: Leucine and protein synthesis: mTOR and beyond. Nutr Rev
2007, 65(3):122–129.
17. Norton L, Layman D: Leucine regulates translation initiation of protein
synthesis in skeletal muscle after exercise. J Nutr 2006, 136:533S–537S.
18. Crozier S, Kimball S, Emmert S, Anthony J, Jefferson L: Oral leucine
administration stimulates protein synthesis in rat skeletal muscle.
J Nutr 2005, 135:376–382.
19. Hara K, Maruki Y, Long X, Yoshino K-I, Oshiro N, Hidayat S, Tokunaga C,
Avruch J, Yonezawa K: Raptor, a binding partner of target of rapamycin
(mTOR), mediates TOR action. Cell 2002, 110:177–189.
20. Kim D, Sarbassov D, Ali SM, King J, Latek R, Erdjument-Bromage H,
Tempst P, Sabatini D: mTOR interacts with raptor to form a nutrient-
sensitive complex that signals to the cell growth machinery. Cell 2002,
110:163–175.
21. Atherton PJ, Babraj J, Smith K, Singh J, Rennie MJ, Wackerhage H: Selective
activation of AMPK-PGC-1_ or PKB-TSC2-mTOR signaling can explain
specific adaptive responses to endurance or resistance training like
electrical muscle stimulation. FASEB J 2005, 19:786–788.
22. Baar K, Esser K: Phosphorylation of p70S6k correlates with increased
skeletal muscle mass following resistance exercise. Am J Physiol Cell
Physiol 1999, 276:C120–C127.
23. Koopman R, Wagenmakers A, Manders R, Zorenc A, Senden J, Gorselink M,
Keizer H, Loon C: Combined ingestion of protein and free leucine with
carbohydrate increases postexercise muscle protein synthesis in vivo in
male subjects. Am J Physiol Endocrinol Metab 2005, 288:E645–E653.
24. Fulks RM, Li JB, Goldberg AL: Effects of insulin, glucose, and amino acids
on protein turnover in rat diaphragm. J Biol Chem 1975, 250:290–298.
25. Li JB, Jefferson LS: Influence of amino acid availability on protein
turnover in perfused skeletal muscle. Biochim Biophys Acta 1978,
544:351–359.
26. Buse MG, Reid SS: Leucine, a possible regulator of protein turnover in
muscle. J Clin Invest 1975, 56:1250–1261.
27. Byfield MP, Murray JT, Backer JM: hVps34 is a nutrient-regulated lipid
kinase required for activation of p70 S6 kinase. J Biol Chem 2005,
280:33076–33082.
28. Nobukuni T, Joaquin M, Roccio M, Dann SG, Kim SY, Gulati P, Byfield MP,
Backer JM, Natt F, Bos JL, Zwartkruis FJ, Thomas G: Amino acids mediate
mTOR/raptor signaling through activation of class 3 phosphatidylinositol
3OH-kinase. Proc Natl Acad Sci USA 2005, 102:14238–14243.
29. Paddon-Jones D, Sheffield-Moore M, Zhang X, Volpi E, Wolf S, Aarsland A,
Ferrando A, Wolfe R: Amino acid ingestion improves muscle protein
synthesis in the young and elderly. Am J Physiol Endocrinol Metab 2004,
286:E321–E328.
30. Tipton K, Ferrando A, Phillips S, Doyle D, Wolfe R: Postexercise net protein
synthesis in human muscle from orally administered amino acids.
Am J Physiol 1999, 276:E628–E634.
Stark et al. Journal of the International Society of Sports Nutrition 2012, 9:54 Page 7 of 8
http://www.jissn.com/content/9/1/54
31. Hoffman J, Ratamess N, Tranchina C, Rashti S, Faigenbaum A: Effect of
protein-supplement timing on strength, power, and body-composition
changes in resistance-trained men. Int J Sport Nutr Exerc Metab 2009,
19(2):172–185.
32. Hoffman J, Ratamess N, Tranchina C, Rashti S, Kang J, Fiagenbaum A: Effects
of a proprietary protein supplement on recovery indices following
resistance exercise in strength/power athletes. Amino Acids 2010,
38:771–778.
33. Cribb P, Hayes A: Effects of supplement timing and resistance exercise on
skeletal muscle hypertrophy. Med Sci Sports Exerc 2006, 38(11):1918–1925.
34. Verdijk L, Jonkers R, Gleeson B: Protein supplementation before and after
exercise does not further augment skeletal muscle hypertrophy after
resistance training in elderly men. Am J Clin Nutr 2009, 89(2):608–616.
35. Hulmi J, Koyanen V, Selanne H, Kraemer W, Hakkinen K, Mero A: Acute and
long-term effects of resistance exercise with or without protein
ingestion on muscle hypertrophy and gene expression. Amino Acids
2009, 37:297–308.
36. Andersen L, Tufekovic G, Zebis M, Crameri R, Verlaan G, Kjaer M, Suetta C,
Magnusson P, Aagaard P: The effect of resistance training combined with
timed ingestion of protein on muscle fiber size and muscle strength.
Metabolism 2005, 54:151–156.
37. Elliot T, Cree M, Sanford A, Wolfe R, Tipton K: Milk ingestion stimulates net
muscle protein synthesis following resistance exercise. Med Sci Sports
Exerc 2006, 38(4):667–674.
38. Hartman J, Tang J, Wilkinson S, Tarnopolsky M, Lawrence R, Fullerton A,
Phillips S: Consumption of fat-free fluid milk after resistance exercise
promotes greater lean mass accretion than does consumption of soy or
carbohydrate in young, novice, male weightlifters. Am J Clin Nutr 2007,
86(2):373–381.
39. Wilkinson S, Tarnopolsky M, MacDonald M, MacDonald J, Armstrong D,
Phillips S: Consumption of fluid skim milk promotes greater muscle
protein accretion after resistance exercise than does consumption of an
isonitrogenous and isoenergetic soy-protein beverage. Am J Clin Nutr
2007, 85(4):1031–1040.
40. Rankin J, Goldman L, Puglisi M, Nickols-Richardson S, Earthman C,
Gwazdauskas F: Effect of post-exercise supplement consumption on
adaptations to resistance training. J Am Coll Nutr 2004, 4(23):322–330.
41. Josse A, Tang J, Tarnopolsky M, Phillips S: Body composition and strength
changes in women with milk and resistance exercise. Med Sci Sports Exerc
2010, 42(6):1122–1130.
42. Tang J, Moore D, Kujbida G, Tarnopolsky M, Phillips S: Ingestion of whey
hydrolysate, casein, or soy protein isolate: effects on mixed muscle
protein synthesis at rest and following resistance exercise in young men.
J Appl Physiol 2009, 107(3):987–992.
43. Norton L, Wilson G, Layman D, Moulton C, Garlick P: Ingestion of whey
hydrolysate, casein, or soy protein isolate: effects on mixed muscle
protein synthesis at rest and following resistance exercise in young men.
Nutr Metab 2012, 9:67.
44. Hoffman J, Falvo M: Protein—which is best? J Sports Sci Med 2004,
3:118–130.
45. Boirie Y, Dangin M, Gachon P, Vasson MP, Maubois JL, Beaufrere B: Slow
and fast dietary proteins differently modulate postprandial accretion.
Proc Natl Acad Sci 1997, 94:14930–14935.
46. Tipton K, Elliot T, Cree M, Wolf S, Sanford A, Wolfe R: Ingestion of casein
and whey proteins result in muscle anabolism after resistance exercise.
Med. Sci. Sports Exerc 2004, 36:2073–2081.
47. Tipton KD, Rasmussen BB, Miller SL, Wolf SE, Owens-Stovall SK, Petrini BE,
Wolfe RR: Timing of amino acid-carbohydrate ingestion alters anabolic
response of muscle to resistance exercise. Am J Physiol Endocrinol Metab
2001, 281:E197–E206.
48. Tipton K, Elliott T, Cree M, Aarsland A, Sanford A, Wolfe R: Stimulation of
net muscle protein synthesis by whey protein ingestion before and after
exercise. Am J Physiol Endocrinol Metab 2007, 292:E71–E76.
doi:10.1186/1550-2783-9-54
Cite this article as: Stark et al.:Protein timing and its effects on
muscular hypertrophy and strength in individuals engaged in weight-
training. Journal of the International Society of Sports Nutrition 2012 9:54.
Submit your next manuscript to BioMed Central
and take full advantage of:
• Convenient online submission
• Thorough peer review
• No space constraints or color figure charges
• Immediate publication on acceptance
• Inclusion in PubMed, CAS, Scopus and Google Scholar
• Research which is freely available for redistribution
Submit your manuscript at
www.biomedcentral.com/submit
Stark et al. Journal of the International Society of Sports Nutrition 2012, 9:54 Page 8 of 8
http://www.jissn.com/content/9/1/54
