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Enzymatic Hydrolysis of a Collagen Hydrolysate Enhances Postprandial Absorption Rate—A Randomized Controlled Trial

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Collagen is characterized by its high content of glycine, proline and hydroxyproline, and is found to exert beneficial effects on joint pain related to activity and osteoarthritis. However, to exert any beneficial effects it is essential that collagen is optimally absorbed. This study aimed to investigate the postprandial absorption of collagen and elucidate the impact of an exogenous enzymatic hydrolysis on absorption rate and bioavailability. A randomized, blinded, cross-over study was conducted where ten healthy male subjects received either 35 g enzymatically hydrolyzed collagen protein (EHC), 35 g non-enzymatically hydrolyzed collagen protein (NC) or placebo (250 mL water) on three nonconsecutive days. Blood samples were drawn before, and up to 240 min following, ingestion and the blood metabolome was characterized by nuclear magnetic resonance (NMR)-based metabolomics. A significant increase in the plasma concentration of nearly all amino acids (AAs) was observed over a 240 min period for both EHC and NC. In addition, the absorption rate and bioavailability of glycine, proline and hydroxyproline were significantly higher for EHC (p < 0.05). In conclusion, ingestion of collagen hydrolysates increases postprandial plasma concentrations of AAs over a period of 240 min, and an enzymatic hydrolysis increases the absorption rate and bioavailability of the collagen-rich AAs glycine, proline and hydroxyproline.
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nutrients
Article
Enzymatic Hydrolysis of a Collagen Hydrolysate
Enhances Postprandial Absorption
Rate—A Randomized Controlled Trial
Kathrine Skov 1, Mikkel Oxfeldt 2, Rebekka Thøgersen 1, Mette Hansen 2and
Hanne Christine Bertram 1, *
1Department of Food Science, Aarhus University, Kirstinebjergvej 10, DK-5792 Aarslev, Denmark;
20095682@post.au.dk (K.S.); rebekka.thoegersen@food.au.dk (R.T.)
2Section for Sport Science, Department of Public Health, Aarhus University, Dalgas Avenue 4,
8000 Aarhus, Denmark; Mikkeloxfeldt@hotmail.com (M.O.); mhan@ph.au.dk (M.H.)
*Correspondence: hannec.bertram@food.au.dk; Tel.: +45-8715-8353
Received: 26 April 2019; Accepted: 10 May 2019; Published: 13 May 2019


Abstract:
Collagen is characterized by its high content of glycine, proline and hydroxyproline,
and is found to exert beneficial eects on joint pain related to activity and osteoarthritis. However,
to exert any beneficial eects it is essential that collagen is optimally absorbed. This study aimed
to investigate the postprandial absorption of collagen and elucidate the impact of an exogenous
enzymatic hydrolysis on absorption rate and bioavailability. A randomized, blinded, cross-over
study was conducted where ten healthy male subjects received either 35 g enzymatically hydrolyzed
collagen protein (EHC), 35 g non-enzymatically hydrolyzed collagen protein (NC) or placebo (250 mL
water) on three nonconsecutive days. Blood samples were drawn before, and up to 240 min following,
ingestion and the blood metabolome was characterized by nuclear magnetic resonance (NMR)-based
metabolomics. A significant increase in the plasma concentration of nearly all amino acids (AAs)
was observed over a 240 min period for both EHC and NC. In addition, the absorption rate and
bioavailability of glycine, proline and hydroxyproline were significantly higher for EHC
(p<0.05)
.
In conclusion, ingestion of collagen hydrolysates increases postprandial plasma concentrations of AAs
over a period of 240 min, and an enzymatic hydrolysis increases the absorption rate and bioavailability
of the collagen-rich AAs glycine, proline and hydroxyproline.
Keywords:
collagen absorption; glycine; proline; hydroxyproline; nutrimetabolomics; collagen
nutriceuticals; sports nutrition; amino acid absorption; dietary protein; collagen uptake
1. Introduction
Chronic tendinopathy, ligament ruptures, and bone fractures are common injuries among athletes.
The consequences can be severe for elite athletes, who may be forced to end their career, since treatment
in a majority of patients is incomplete and unsuccessful from the perspective of a return to play
at the elite level [
1
,
2
]. Furthermore, osteoarthritis (OA) is a chronic condition aecting the joints,
and the progressive degeneration of articular cartilage causes pain and functional disability. It is
estimated that more than 250 million people are aected by OA worldwide and to this day no curative
treatments are available [
3
]. Therefore, there is an obvious need to search for alternative treatment
strategies to optimize rehabilitation. Bones, tendons, ligaments, and cartilage are constituted by a
complex matrix but are characterized by a high content of collagen, and this structural protein is
vital for the structure and biomechanical properties of these musculoskeletal tissues [
4
]. Collagen
is an abundant structural protein present in connective tissue. Besides its nutritional value as a
protein source, dietary supplementation with collagen-derived peptide sources has been suggested to
Nutrients 2019,11, 1064; doi:10.3390/nu11051064 www.mdpi.com/journal/nutrients
Nutrients 2019,11, 1064 2 of 13
provide beneficial eects in patients with tendinopathy [
5
7
], chronic joint instability [
8
], osteoarthritis
(OA) [
3
,
9
12
], and activity-related joint pain [
13
,
14
]. Thus, nutritional interventions focusing on
increasing the amino acid (AA) components of collagen have been suggested to improve collagen
synthesis of collagen-rich tissues such as ligaments and bones [
5
] and potentially slow the degenerative
process in OA aected joints [
11
]. However, in order to exert any potential beneficial eects, optimal
digestion and absorption of AA components of collagen is pivotal. Protein digestibility generally
varies depending on dietary source and processing methods [
15
]. Enzymatic hydrolysis of proteins is
expected to influence protein digestion and the postprandial plasma AA profile, and previous studies
on milk proteins have shown an enhanced postprandial AA absorption and bioavailability when
the protein source was subjected to an enzymatic hydrolysis [
16
]. Congruently, it is expected that
an enhanced absorption rate will be observable when the collagen protein structure is manipulated
through an enzymatic hydrolysis. In nutrition science, metabolomics provides a characterization of the
wide-ranging scale of metabolites (the metabolome) and enables a comprehensive comparison of the
metabolic responses after dierent dietary interventions [
17
].
1
H NMR spectroscopy is able to detect
any mobile proton-containing metabolites with a low-molecular weight [
17
], and
1
H NMR-based
metabolomics spectroscopy has recently been shown to be a valuable tool for the characterization of
postprandial blood plasma AA profiles [
18
,
19
]. Thus,
1
H NMR-based metabolomics applied to blood
plasma samples provides a framework for elucidating the eect of an enzymatic hydrolysis on the
bioavailability and absorption of collagen. To the best of our knowledge, investigations on the eects
of enzymatic hydrolysis of collagen protein on the postprandial absorption rate have not been reported
yet. Consequently, the aim of the present study was to examine the postprandial absorption of collagen
hydrolysates and to elucidate the impact of an exogenous enzymatic hydrolysis on the absorption rate
and bioavailability. For this purpose, an acute human intervention study with a cross-over design
was conducted with two collagen-containing products; an enzymatically hydrolyzed collagen protein
(EHC), and a non-enzymatically hydrolyzed collagen protein (NC), and the blood plasma metabolome
was characterized by
1
H NMR-based metabolomics. We hypothesized that the increase in circulating
amino acids would be faster and more pronounced after ingestion of EHC compared to NC.
2. Materials and Methods
2.1. Subjects
Ten healthy young males were recruited from the Section of Sports Science, Department of
Public Health at Aarhus University. The recruitment was initiated on 8 November 2018 and the last
experimental day including subjects was completed on 15 December 2018. Participants were healthy
males in the age range 18–35. Participant characteristics are provided in Table 1. Exclusion criteria
included: the use of medication that could influence postprandial absorption; muscular, tendon or
metabolic illnesses; smoking; hypertension; use of supplements during the trial period; weight loss or
gain of more than 2 kg during the trial period; and working night shifts. The trial complied with the
Declaration of Helsinki and was approved by The Central Denmark Region Committees on Health
Research Ethics (Journal 65141, case number 1-10-72-287-18). Furthermore, the study was registered at
ClinicalTrials gov (NCT03749239) and adhered to the CONSORT guidelines. All participants gave
their informed consent before the intervention was carried out.
Table 1. Subject characteristics.
Subjects (n=10)
Age (y) 26 ±1
Weight (kg) 77 ±6
Height (cm) 180 ±5
Activity level (h)
BMI (kg/m2)
10 ±3
24 ±2
y, years; h, hours of physical activity per week. BMI, body mass index. All values are mean ±SD.
Nutrients 2019,11, 1064 3 of 13
2.2. Design and Experimental Protocol
The study was a randomized, blinded, placebo-controlled, cross-over study consisting of three
nonconsecutive experimental days at the Section for Sports Science, Department of Public Health at
Aarhus University. On the experimental days, participants consumed a collagen supplement of 35 g
crude protein enzymatically hydrolyzed collagen (EHC), or non-enzymatically hydrolyzed collagen
(NC), or placebo (water) in restricted random order based on the recruitment order to ensure that an
equal number of subjects received the three intervention beverages on experimental days one, two and
three, respectively. One of the primary investigators (MH), who did not take an active part in the blood
sampling procedures or the following laboratory analysis, performed the randomization. The subjects
were blind to the content and order of the intervention beverages. Furthermore, the laboratory sta
were not informed about the sample id for the intervention beverages until the statistical analysis was
initiated. Participants were instructed to record their dietary intake and physical activity level on the
day before the first trial. These recordings were used as a standard plan before the following trial
days to ensure standardization of dietary intake and physical activity level. On trial days, participants
arrived in the morning after an overnight fast and consumed a drink containing either EHC, NC,
or placebo dissolved in 250 mL water. The drinks were consumed within 2 min and the time was
designated as time zero. Immediately following consumption of the drink, all participants consumed
100 mL water to attenuate the mouthfeel and taste of the trial product. Additional blood samples were
drawn at fixed time points for four hours following beverage consumption (Figure 1). Furthermore,
participants were instructed to answer three questions on a visual analogue scale (VAS) after every
blood sample collection. All participants remained in a resting position and were not allowed to eat
or drink throughout the trial hours. There were at least two days between the experimental days.
The primary outcome measure was the time to peak amino acid concentration and the accumulated
amino acid concentration of the individual amino acids after ingestion of the intervention beverages.
Nutrients 2019, 11, x FOR PEER REVIEW 3 of 13
2.2. Design and Experimental Protocol
The study was a randomized, blinded, placebo-controlled, cross-over study consisting of three
nonconsecutive experimental days at the Section for Sports Science, Department of Public Health at
Aarhus University. On the experimental days, participants consumed a collagen supplement of 35 g
crude protein enzymatically hydrolyzed collagen (EHC), or non-enzymatically hydrolyzed collagen
(NC), or placebo (water) in restricted random order based on the recruitment order to ensure that an
equal number of subjects received the three intervention beverages on experimental days one, two
and three, respectively. One of the primary investigators (MH), who did not take an active part in the
blood sampling procedures or the following laboratory analysis, performed the randomization. The
subjects were blind to the content and order of the intervention beverages. Furthermore, the
laboratory staff were not informed about the sample id for the intervention beverages until the
statistical analysis was initiated. Participants were instructed to record their dietary intake and
physical activity level on the day before the first trial. These recordings were used as a standard plan
before the following trial days to ensure standardization of dietary intake and physical activity level.
On trial days, participants arrived in the morning after an overnight fast and consumed a drink
containing either EHC, NC, or placebo dissolved in 250 mL water. The drinks were consumed within
2 min and the time was designated as time zero. Immediately following consumption of the drink,
all participants consumed 100 mL water to attenuate the mouthfeel and taste of the trial product.
Additional blood samples were drawn at fixed time points for four hours following beverage
consumption (Figure 1). Furthermore, participants were instructed to answer three questions on a
visual analogue scale (VAS) after every blood sample collection. All participants remained in a resting
position and were not allowed to eat or drink throughout the trial hours. There were at least two days
between the experimental days. The primary outcome measure was the time to peak amino acid
concentration and the accumulated amino acid concentration of the individual amino acids after
ingestion of the intervention beverages.
Figure 1. Experimental protocol and fixed time points for blood sample collection and visual analogue
scale (VAS) questionnaire. Time of drink consumption was designated as time zero (0 min).
2.3. Collagen Products
The collagen supplements were manufactured and provided by Essentia Protein Solutions A/S,
Denmark. Two different types of collagen products were included: an enzymatically hydrolyzed
collagen (EHC) (OmniCol
TM
110, Essentia Protein Solutions, Graasten, Denmark) and a non-
enzymatically hydrolyzed collagen-containing product (NC) (HydroBEEF
TM
, Essentia Protein
Solutions, Graasten, Denmark). Both products were manufactured from beef bone, and the collagen
products did not contain any additives, sweeteners or flavors. OmniCol
TM
100 and HydroBEEF
TM
are
commercial products and thus approved for human consumption according to GRAS. Accordingly,
the products’ content of heavy metals has been analyzed and the values are considered safe. We have
not conducted any further testing of the products in relation to safety, but a recent study with a
similar collagen product revealed that the product did not elicit any toxic effects in an in vitro model
[20].
Figure 1.
Experimental protocol and fixed time points for blood sample collection and visual analogue
scale (VAS) questionnaire. Time of drink consumption was designated as time zero (0 min).
2.3. Collagen Products
The collagen supplements were manufactured and provided by Essentia Protein Solutions
A/S, Denmark. Two dierent types of collagen products were included: an enzymatically
hydrolyzed collagen (EHC) (OmniCol
TM
110, Essentia Protein Solutions, Graasten, Denmark) and a
non-enzymatically hydrolyzed collagen-containing product (NC) (HydroBEEF
TM
, Essentia Protein
Solutions, Graasten, Denmark). Both products were manufactured from beef bone, and the collagen
products did not contain any additives, sweeteners or flavors. OmniCol
TM
100 and HydroBEEF
TM
are
commercial products and thus approved for human consumption according to GRAS. Accordingly,
the products’ content of heavy metals has been analyzed and the values are considered safe. We have
not conducted any further testing of the products in relation to safety, but a recent study with a similar
collagen product revealed that the product did not elicit any toxic eects in an in vitro model [20].
Nutritional contents and amino acid profiles are provided as supplementary material. Collagen
powder corresponding to 35 g protein was dissolved in 200 mL ~50
C hot water. Prior to ingestion,
four ice cubes (~50 mL) were added to allow drinks to cool.
Nutrients 2019,11, 1064 4 of 13
2.4. Blood Samples
In each experimental trial, eight blood samples of ~8 mL were collected into evacuated containers
containing lithium heparin. The blood samples were collected 20 min before ingestion of the intervention
beverage and at 20, 40, 60, 90, 120, 180 and 240 min after ingestion of the intervention beverage (Figure 1).
The blood samples were centrifuged at 1200
×
gfor 10 min at 4
C to separate the plasma and stored at
80 C until further analysis.
2.5. Visual Analogue Scale (VAS)
Sensation of hunger, satiety and fullness were measured using a VAS, consisting of three questions.
The VAS consisted of a 73 mm horizontal unbroken line with a statement anchored at each end
describing the extremes, e.g., “I am not hungry at all”, “I have never been more hungry”, “I am
not full at all”, “I am very full”. Each set of VAS questions was answered at every blood sampling.
The intensity of the feeling (distance of the vertical mark from the origin on the left) was measured,
yielding a score in the range 0–73 mm.
2.6. H NMR Spectroscopy
Thawed plasma samples were filtered using 10K Amicon Ultra centrifugal filter units (Merck
Milipore Ltd., Cork, Ireland). Prior to use, the filters were washed four times with 500
µ
L MilliQ water
to remove traces of glycerol. After washing, 500
µ
L plasma was added to the filters and centrifuged at
4
C, 14,000
×
gfor 2 h. Subsequently, 400
µ
L plasma filtrate was transferred to a 5 mm NMR tube
containing 100
µ
L sodium-hydrogen phosphate buer (50 mM Na
2
HPO
4
, pH 7.4), 25
µ
L deuterium
oxide (D
2
O) with 0.05% TSP and 75
µ
L D
2
O.
1
H NMR spectroscopy was performed on a Bruker
Avance III 600 MHz NMR spectrometer operating at a
1
H frequency of 600.13 MHz with a 5 mm
1
H TXI probe (Bruker Biospin, Reinstetten, Germany). Spectra were acquired at 298 K using the
one-dimensional (1D) nuclear Overhauser enhancement spectroscopy (NOESY)-pulse sequence presat
for water suppression (NOESYPR1D). The acquisition parameters used were: 64 scans (NS), spectral
width (SW) =7288 Hz (12.1450 ppm), data points (TD) =32,768, relaxation delay (D1) =5.0 s and
acquisition time (AQ) =2.25 s. The free induction decays (FIDs) were multiplied by a line-broadening
function of 0.3 Hz prior to Fourier transformation. A total of 263 samples were analyzed, and the
spectra obtained were baseline and phase corrected using TopSpin 3.0 (Bruker BioSpin, Rheinstetten,
Germany). Metabolite assignment and quantification were conducted using Chenomx NMR Suite 8.13
(Chenomx Inc., Edmonton, AB, Canada).
2.7. Statistical Analyses
Statistical analysis and graph design were performed using GraphPad Prism (GraphPad Software
Inc., San Diego, CA, USA). Blood sample concentrations and area under the curve (AUC) data were
analyzed using a two-factor RM ANOVA with statistical significance defined as p<0.05. All data sets
were tested for normal distribution and sphericity prior to the ANOVA analysis. Normal distribution
was tested using the Shapiro-Wilk test. Data were transformed by the natural logarithm if deviations
from the normal distribution were observed. Sphericity was tested using Mauchly’s Test of Sphericity
and a Greenhouse–Geisser correction was used if Mauchly’s Test of Sphericity showed a p-value of less
than 0.05.
Previous trials of 35 g hydrolyzed vs. non-hydrolyzed milk protein tests have shown a dierence
in amino acid uptake of 1.4 mmol * 6 h/L (this dierence was only evident over the first 4 h) [
16
].
Assuming that the dierence in amino acid concentration accumulated over four hours after ingestion
of EHC and NC is of a similar magnitude, a test population of 8 would enable detection of a significant
dierence (significance level 0.05, 80% power, SD 1.0 mmol * 4 h/L). Nevertheless, we included 10
subjects to account for possible dropout or missing data.
Nutrients 2019,11, 1064 5 of 13
3. Results
3.1. H NMR Spectroscopic Analysis
1
H NMR spectroscopy was performed on all blood plasma samples and a total of 19 plasma
metabolites were identified and quantified. These included 18 AAs and glucose (Table 2). The four
AAs—aspartic acid, cysteine, tryptophan and hydroxylysine—were not quantified, as their concentration
was below the detection limit of the applied 1H NMR spectroscopic analysis.
Table 2. List of identified metabolites and their chemical shift (δ).
Metabolite δ1H (multiplicity)
Alanine 1.46 (d), 3.76 (q)
Arginine 1.68 (m), 1.90 (m), 3.23 (t), 3.76 (t)
Asparagine 4.00 (dd), 2.94 (m), 2.84 (m)
Glucose
3.233 (dd), 3.398 (m), 3.458 (m), 3.524 (dd), 3.726 (m), 3.824 (m), 3.889 (dd), 4.634 (d), 5.223 (d)
Glycine 3.54 (s)
Histidine 3.16 (dd), 3.23 (dd), 3.98 (dd), 7.09 (d), 7.90 (d)
Isoleucine 0.926 (t), 0.997 (d), 1.246 (m), 1.475 (m), 1.968 (m), 3.661 (d)
Leucine 0.948 (t), 1.700 (m), 3.722 (m)
Lysine 1.46 (m), 1.71 (m), 1.89 (m), 3.02 (t), 3.74 (t)
Methionine 2.157 (m), 2.631 (t), 3.851 (dd)
Phenylalanine 3.19 (m), 3.98 (dd), 7.32 (d), 7.36 (m), 7.42 (m)
Proline 1.99 (m), 2.06 (m), 2.34 (m), 3.33 (dt), 3.41 (dt), 4.12 (dd)
Serine 3.832 (dd), 3.958 (m)
Threonine 1.316 (d), 3.575 (d), 4.244 (m)
Tyrosine 3.024 (dd), 3.170 (dd), 3.921 (dd), 6.877 (m), 7.170 (m)
Valine 0.976 (d), 1.029 (d), 2.261 (m), 3.691 (d)
Hydroxyproline 2.14 (ddd), 2.42 (M), 3.36 (ddd), 3.46 (dd), 4.33 (d), 4.35 (d)
s=singlet, d =doublet, dd =doublet of doublets, ddd =doublet of doublet of doublets, dt =doublet of triplets,
m=multiplet, t =triplet, q =quartet.
3.2. General AAs Absorption
Figure 2shows the total amount of AAs during the postprandial period. A significant increase in
postprandial plasma concentration of total AAs was observed after ingestion of both collagen products
compared to placebo. In addition, after EHC and NC intake, a significantly higher postprandial
plasma concentration was observed compared with placebo (p<0.05) for all AAs except histidine
and methionine (Table 3). Histidine and methionine also increased after intake for both EHC and
NC, however, a significant dierence was only observed between NC and placebo (Table 3). For all
AAs except glycine, proline, hydroxyproline and methionine, no significant dierence in AUC was
observed between EHC and NC. All AA concentrations peaked earlier after intake for EHC compared
with NC, and for many AAs, a significant dierence in AA concentration between EHC and NC was
observed at 20 min after intake. Postprandial concentrations of individual AAs over time can be found
in the supplementary material.
Nutrients 2019,11, 1064 6 of 13
Nutrients 2019, 11, x FOR PEER REVIEW 6 of 13
Figure 2. Plasma concentration of total amino acids (AAs). A significant difference was observed
between enzymatically hydrolyzed collagen (EHC) and placebo as well as between non-
enzymatically hydrolyzed collagen (NC) and placebo, but not between the two collagen supplements.
# p < 0.05, ## p < 0.002 ### p < 0.0002, #### p < 0.0001. All data are mean ± SEM.
Table 3. Pairwise comparisons of dietary treatments for all amino acids (AAs).
Comparison EHC vs. NC EHC vs. placebo NC vs. placebo
AA EHC NC p EHC Pl p NC Pl p
Alanine 367.4 351.3 0.69 367.4 266.2 0.0002 351.3 266.2 0.001
Arginine 86.54 82.54 0.67 86.54 50.28 <0.0001 82.54 50.28 <0.0001
Asparagine 40.60 40.43 0.99 40.60 34.58 0.019 40.43 34.58 0.023
Glutamate 39.17 34.34 0.25 39.17 25.68 0.0006 34.34 25.68 0.021
Glutamine 513.5 521.9 0.87 513.5 469.2 0.043 521.9 469.2 0.015
Glycine 448.7 380.7 0.0059 448.7 204.1 <0.0001 380.7 204.1 <0.0001
Histidine 72.85 77.37 0.12 72.85 67.38 0.053 77.37 67.38 0.0006
Hydroxyproline 64.92 48.88 0.0087 64.92 1.484 <0.0001 48.88 1.484 <0.0001
Isoleucine 73.62 68.00 0.21 73.62 58.24 0.0003 68.00 58.24 0.017
Leucine 127.4 120.8 0.42 127.4 99.09 <0.0001 120.8 99.09 0.0014
Lysine 122.5 119.1 0.79 122.5 99.37 0.0008 119.1 99.37 0.0034
Methionine 18.78 22.51 0.034 18.78 17.42 0.59 22.51 17.42 0.0043
Proline 335.4 285.8 0.0074 335.4 186.3 <0.0001 285.8 186.3 <0.0001
Phenylalanine 47.65 48.21 0.93 47.65 39.80 0.0003 48.21 39.80 0.0001
Serine 147.7 145.0 0.89 147.7 108.5 <.0001 145.0 108.5 <0.0001
Threonine 152.2 148.2 0.83 152.2 120.6 0.0006 148.2 120.6 0.0023
Tyrosine 56.47 55.91 0.96 56.47 48.72 0.011 55.91 48.72 0.018
Valine 278.4 254.9 0.09 278.4 213.2 <0.0001 254.9 213.2 0.0027
BCAA 479.5 443.7 0.15 479.5 370.6 <0.0001 443.7 370.6 0.002
Total AA 2994 2806 0.14 2994 2110 <0.0001 2806 2110 <0.0001
EAA 893.4 859.1 0.48 893.4 715.2 <0.0001 859.1 715.2 0.0003
Gly-Pro-Hp 849.1 715.4 0.0003 849.1 391.8 <0.0001 715.4 391.8 <0.0001
EHC, enzymatically hydrolyzed collagen; NC, non-enzymatically hydrolyzed collagen; Pl, placebo;
BCAA, Branched chain amino acids; AA, amino acids; EAA, essential AA; Gly-Pro-Hp, sum of
glycine, proline and hydroxyproline. Data are mean concentrations (μM). p = statistical p-value.
3.3. Specific Collagen AAs
Overall statistical analysis showed a significant difference between EHC and NC for plasma
concentrations of glycine (p = 0.0059), hydroxyproline (p = 0.0087), proline (p = 0.0074) and the sum of
glycine, proline and hydroxyproline (Gly-Pro-Hyp) (p = 0.0003). In addition, plasma concentrations
of these AAs were significantly different for both collagen products compared to placebo (Table 3).
For glycine, a significant difference was observed between EHC and NC from 20–90 min (Figure
3A). A separate analysis of the AUC showed a significant difference between EHC and NC (Figure
Figure 2.
Plasma concentration of total amino acids (AAs). A significant dierence was observed
between enzymatically hydrolyzed collagen (EHC) and placebo as well as between non-enzymatically
hydrolyzed collagen (NC) and placebo, but not between the two collagen supplements. # p<0.05,
## p<0.002, ### p<0.0002, #### p<0.0001. All data are mean ±SEM.
Table 3. Pairwise comparisons of dietary treatments for all amino acids (AAs).
Comparison EHC vs. NC EHC vs. Placebo NC vs. Placebo
AA EHC NC pEHC Pl pNC Pl p
Alanine 367.4 351.3 0.69 367.4 266.2 0.0002 351.3 266.2 0.001
Arginine 86.54 82.54 0.67 86.54 50.28 <0.0001 82.54 50.28 <0.0001
Asparagine 40.60 40.43 0.99 40.60 34.58 0.019 40.43 34.58 0.023
Glutamate 39.17 34.34 0.25 39.17 25.68 0.0006 34.34 25.68 0.021
Glutamine 513.5 521.9 0.87 513.5 469.2 0.043 521.9 469.2 0.015
Glycine 448.7 380.7 0.0059 448.7 204.1 <0.0001 380.7 204.1 <0.0001
Histidine 72.85 77.37 0.12 72.85 67.38 0.053 77.37 67.38 0.0006
Hydroxyproline 64.92 48.88 0.0087 64.92 1.484 <0.0001 48.88 1.484 <0.0001
Isoleucine 73.62 68.00 0.21 73.62 58.24 0.0003 68.00 58.24 0.017
Leucine 127.4 120.8 0.42 127.4 99.09 <0.0001 120.8 99.09 0.0014
Lysine 122.5 119.1 0.79 122.5 99.37 0.0008 119.1 99.37 0.0034
Methionine 18.78 22.51 0.034 18.78 17.42 0.59 22.51 17.42 0.0043
Proline 335.4 285.8 0.0074 335.4 186.3 <0.0001 285.8 186.3 <0.0001
Phenylalanine 47.65 48.21 0.93 47.65 39.80 0.0003 48.21 39.80 0.0001
Serine 147.7 145.0 0.89 147.7 108.5 <.0001 145.0 108.5 <0.0001
Threonine 152.2 148.2 0.83 152.2 120.6 0.0006 148.2 120.6 0.0023
Tyrosine 56.47 55.91 0.96 56.47 48.72 0.011 55.91 48.72 0.018
Valine 278.4 254.9 0.09 278.4 213.2 <0.0001 254.9 213.2 0.0027
BCAA 479.5 443.7 0.15 479.5 370.6 <0.0001 443.7 370.6 0.002
Total AA 2994 2806 0.14 2994 2110 <0.0001 2806 2110 <0.0001
EAA 893.4 859.1 0.48 893.4 715.2 <0.0001 859.1 715.2 0.0003
Gly-Pro-Hp 849.1 715.4 0.0003 849.1 391.8 <0.0001 715.4 391.8 <0.0001
EHC, enzymatically hydrolyzed collagen; NC, non-enzymatically hydrolyzed collagen; Pl, placebo; BCAA, Branched
chain amino acids; AA, amino acids; EAA, essential AA; Gly-Pro-Hp, sum of glycine, proline and hydroxyproline.
Data are mean concentrations (µM). p=statistical p-value.
3.3. Specific Collagen AAs
Overall statistical analysis showed a significant dierence between EHC and NC for plasma
concentrations of glycine (p=0.0059), hydroxyproline (p=0.0087), proline (p=0.0074) and the sum of
glycine, proline and hydroxyproline (Gly-Pro-Hyp) (p=0.0003). In addition, plasma concentrations of
these AAs were significantly dierent for both collagen products compared to placebo (Table 3).
For glycine, a significant dierence was observed between EHC and NC from 20–90 min (Figure 3A).
A separate analysis of the AUC showed a significant dierence between EHC and NC (Figure 3B).
These findings show that overall the glycine concentration was higher for EHC compared to both
NC (p=0.0029) and placebo (p<0.0001). Plasma concentration of proline showed a significantly
Nutrients 2019,11, 1064 7 of 13
higher proline concentration for EHC compared to NC at 20 and 60 min (Figure 3C). A separate
AUC analysis showed that overall the concentration of proline was significantly higher following
consumption of EHC compared to NC (p=0.0028) and placebo (p<0.0001) (Figure 3D). Analysis of
hydroxyproline concentrations showed a significant dierence between EHC and NC collagen at all
time points between 40 and 120 min (Figure 3E). Both EHC and NC showed a significantly higher
concentration of hydroxyproline compared to placebo at all time points between 20 and 240 min.
Furthermore, a separate AUC analysis showed an overall higher concentration of hydroxyproline
following consumption of EHC compared to NC (p=0.0082) and placebo (p<0.0001) (Figure 3F).
Analysis of the total sum of glycine, proline and hydroxyproline (Gly-Pro-Hyp) showed a significant
dierence between EHC and NC at all time points between 40 and 120 min (Figure 3G). In addition,
both EHC and NC showed a significantly higher Gly-Pro-Hyp concentration than placebo at all time
points between 20 and 240 min. A separate AUC analysis showed a significantly higher Gly-Pro-Hyp
concentration following consumption of EHC compared to NC (p=0.0002) and placebo (p<0.001)
(Figure 3H).
Nutrients 2019, 11, x FOR PEER REVIEW 8 of 13
Figure 3. Postprandial plasma concentration of glycine (A), proline (C), hydroxyproline (E) and Gly-
Pro-Hyp (G) after ingestion of enzymatically hydrolyzed collagen (EHC), non-enzymatically
hydrolyzed collagen (NC) or placebo. Area under the curve (AUC) analysis for each concentration of
glycine (B), proline (D), hydroxyproline (F) and Gly-Pro-Hyp (H) is evaluated from time = 20 min. *
Significant difference between EHC and NC. # significant differences between either of the collagen
products and placebo. * p < 0.05, ** p < 0.002 *** p < 0.0002, **** p < 0.0001, # p < 0.05, ## p < 0.002 ### p
< 0.0002, #### p < 0.0001. Values are mean ± SEM.
Figure 3. Cont.
Nutrients 2019,11, 1064 8 of 13
Nutrients 2019, 11, x FOR PEER REVIEW 8 of 13
Figure 3. Postprandial plasma concentration of glycine (A), proline (C), hydroxyproline (E) and Gly-
Pro-Hyp (G) after ingestion of enzymatically hydrolyzed collagen (EHC), non-enzymatically
hydrolyzed collagen (NC) or placebo. Area under the curve (AUC) analysis for each concentration of
glycine (B), proline (D), hydroxyproline (F) and Gly-Pro-Hyp (H) is evaluated from time = 20 min. *
Significant difference between EHC and NC. # significant differences between either of the collagen
products and placebo. * p < 0.05, ** p < 0.002 *** p < 0.0002, **** p < 0.0001, # p < 0.05, ## p < 0.002 ### p
< 0.0002, #### p < 0.0001. Values are mean ± SEM.
Figure 3.
Postprandial plasma concentration of glycine (
A
), proline (
C
), hydroxyproline (
E
) and
Gly-Pro-Hyp (
G
) after ingestion of enzymatically hydrolyzed collagen (EHC), non-enzymatically
hydrolyzed collagen (NC) or placebo. Area under the curve (AUC) analysis for each concentration of
glycine (
B
), proline (
D
), hydroxyproline (
F
) and Gly-Pro-Hyp (
H
) is evaluated from time =
20 min.
* Significant dierence between EHC and NC. # significant dierences between either of the collagen
products and placebo. * p<0.05, ** p<0.002 *** p<0.0002, **** p<0.0001, # p<0.05, ## p<0.002,
### p<0.0002, #### p<0.0001. Values are mean ±SEM.
3.4. Glucose
Overall analysis of plasma glucose showed no eect of time (p=0.57), treatment (p=0.87) or
time*treatment interaction (p=0.4239). Accordingly, the AUC showed no significant eect of treatment
(p=0.99) or time*treatment interaction (p=0.93) on the plasma concentration of glucose. Data are
shown in Figure S5 in the supplementary material.
3.5. VAS
Overall analysis for VAS 1 (how hungry are you) showed a significant eect of time (p<0.0001),
whereas no eect of treatment (p=0.27) or time*treatment interaction (p=0.2005) were observed.
Similarly, overall analysis of VAS 2 data (how full are you) showed a significant eect of time
(p<0.0001),
but no eect of treatment (p=0.22) or time*treatment interaction (p=0.65). Overall analysis of VAS 3
data (how filled up do you feel) showed a significant eect of time (p<0.0001) but not of treatment
(p=0.275) or time*treatment interaction (p=0.2203). VAS data can be found in the supplementary
material (Figure S6).
4. Discussion
Protein hydrolysates have been suggested to be more readily digested and absorbed compared
to intact proteins, thus creating a more rapid increase in plasma concentrations of AAs [
16
]. In the
present study the postprandial responses to intake of enzymatically hydrolyzed collagen (EHC) and
non-enzymatically hydrolyzed collagen (NC) were examined by
1
H NMR-based metabolomics for the
first time. AA quantification from the
1
H NMR-based metabolomics data displayed a significantly
higher plasma AA concentration following ingestion of EHC and NC compared to placebo, revealing
that both products provided absorbable AAs including essential AAs. In addition, ingestion of EHC
led to significantly higher plasma concentrations of glycine, proline and hydroxyproline compared to
non-hydrolyzed collagen. Notably, these AAs have been connected with the potential beneficial eects
of collagen supplementation on tendinopathy and articular joint pain experienced in both OA patients
and athletes [
7
14
,
21
,
22
]. As net collagen synthesis is negative in OA patients, enhancing collagen
synthesis has been proposed to help regeneration of cartilage [
11
,
22
]. Baar et al. [
23
] showed that
AAs present in high amounts in collagen (glycine, proline, lysine, hydroxyproline and hydroxylysine)
combined with vitamin C could improve collagen synthesis in a tissue-engineered ligament model.
Similar findings were reported by de Paz-Lugo et al. [
20
], who found that glycine and proline enhanced
type II collagen synthesis in bovine articular chondrocytes, suggesting that increased consumption of
Nutrients 2019,11, 1064 9 of 13
these AAs could be beneficial for the regeneration of the articular cartilage matrix. Oesser et al. [
24
]
investigated the absorption of gelatin hydrolysate and its subsequent tissue distribution in mice.
They found that gelatin hydrolysate accumulated in cartilage tissue, suggesting it as a potential
mechanism for clinically observed benefits of gelatin consumption [
24
]. Whether these
in vitro
and animal findings can be transferred to human adults has not been established. Nevertheless,
in a
controlled double-blinded trial including 30 patients with mild knee OA, collagen hydrolysate
supplementation seemed to have a positive eect on cartilage compared to placebo when measured
by delayed gadolinium enhanced magnetic resonance imaging [
11
]. Furthermore, intake of 15 g of a
vitamin C-enriched gelatin supplement prior to an exercise bout of 6 min rope-skipping enhanced
plasma concentration of the amino-terminal propeptide of type I collagen, indicating increased collagen
synthesis likely related to increase in bone type I collagen synthesis [5].
In the present study, the plasma concentrations of proline, glycine and hydroxyproline, following
ingestion of EHC were similar to reported plasma concentrations after intake of 15 g vitamin-C-enriched
gelatin supplement in the trial by Shaw et al. [
5
]. This finding could indicate that there is an upper
threshold in regards to experiencing beneficial eect after increasing doses of collagen-derived peptides.
The concentrations of glycine, proline and hydroxyproline increased significantly at 20 min post-
ingestion of EHC compared to baseline. In contrast, NC led to a significant increase in glycine, proline
and hydroxyproline at 40 min post-ingestion compared to baseline. Glycine content of the EHC
product was slightly higher (1.6 g/100 g, Table S1) than for the NC, whereas the content of proline
and hydroxyproline was similar in EHC and NC, respectively. Consequently, the dierence in the
initial postprandial increase in glycine, proline and hydroxyproline concentrations, and the larger
AUC for EHC compared to NC, indicate that an enzymatic hydrolysis enhances both absorption rate
and bioavailability of these collagen AAs. Previous studies have investigated the absorption rate and
plasma concentration of AAs following ingestion of intact proteins and their hydrolysate. However,
these studies have primarily focused on whey and casein proteins and results have been conflicting.
Koopman et al. [
16
] and Calbet et al. [
25
] reported an increased absorption rate of hydrolyzed casein
compared to its intact protein, while Schmedes et al. (2018) found no significant dierence between
casein and its hydrolysate [
26
]. Hydroxyproline concentration was observed to peak at 120 min and
90 min post-ingestion of EHC and NC, respectively. Though hydroxyproline peaked later following
EHC, it reached a higher plasma concentration. Plasma concentrations of glycine and proline peaked 60
and 90 min after intake of EHC and NC, respectively. Previous studies have reported similar findings
with a peak in hydroxyproline after 120 min and a subsequent decrease to half the maximum after four
hours following consumption of hydrolyzed collagen [20,27].
Plasma concentrations of alanine, asparagine, glutamate, glutamine, histidine, serine, threonine
and valine were significantly higher over the 240 min period after consumption of both EHC and NC
compared to placebo. In addition, plasma concentrations of total AA, essential AAs and branched chain
amino acids (BCAAs) did not dier between EHC and NC, indicating that an enzymatic hydrolysis
did not have a significant eect on bioavailability and absorption rate for these AAs. Similar findings
have previously been observed for hydrolyzed casein and intact casein [
26
]. Plasma concentrations
of arginine, leucine, lysine, tyrosine, isoleucine and phenylalanine increased significantly from 20 to
60 min following ingestion of EHC and NC, with no significant dierences between the two products.
Plasma concentrations of alanine, asparagine, glutamate, glutamine, histidine, serine, threonine and
valine showed an increased absorption from 20 to 120 min, after which levels decreased back to
baseline. These findings are in accordance with previous studies on gelatin [
5
] and studies comparing
hydrolyzed casein to intact casein [
15
], where the concentration of most AAs increased significantly
after approximately 30 min. The results suggest that an enzymatic hydrolysis is not necessary, if the
main goal of consumption is to provide the body with essential AAs (EAAs), BCAAs or any of the
less abundant AAs of collagen. As a result of their low content of BCAA and tryptophan, collagen
sources have not been anticipated to strongly stimulate muscle protein synthesis. However, a study
by Zdzieblik et al. [
26
] found that ingestion of collagen peptides following exercise led to higher
Nutrients 2019,11, 1064 10 of 13
fat free mass (FFM), bone mass (BM), muscle strength and loss of fat mass compared to placebo in
elderly sarcopenic men [
26
]. Taken together with the findings of the present study, it is plausible
that an enzymatic hydrolysis is not vital if using collagen as a protein supplement following exercise
or for combating sarcopenia. Zdzieblik et al. [
26
] speculated that the observed benefits of collagen
ingestion on muscle protein synthesis could be due to the high content of glycine and its role as a
substrate in creatine synthesis [
28
]. Others have proposed that glycine plays a vital role for collagen’s
beneficial eects in relation to OA [
22
], but also as a metabolite that plays a crucial role in epigenetic
regulation and exerts important physiological functions including antioxidative, anti-inflammatory
and immunomodulatory eects [29].
NC displayed a significantly higher postprandial plasma concentration of histidine compared
to placebo, while EHC did not dier significantly from placebo. This finding could be explained by
the fact that the NC product contained a slightly higher amount of histidine (1.1 g/100 g) compared
to the EHC product (0.8 g/100 g). NC also displayed a significantly higher postprandial plasma
concentration of methionine. This can probably be ascribed to oxidation of methionine to methionine
sulfoxide and methionine sulfone taking place during the enzymatic hydrolysis processes, consistent
with observations in other mildly processed foods [30,31].
Glucose quantification from the 1H NMR metabolomics data displayed no significant dierence
between postprandial glucose concentrations in EHC, NC and placebo. This is in accordance with
previous findings reporting that postprandial blood glucose was not significantly aected in healthy
individuals following protein ingestion [
31
33
], and other studies comparing postprandial glucose
levels following ingestion of intact proteins and their hydrolysate have found similar results. Koopman
et al. [
15
] found no dierence in glucose response following consumption of a casein hydrolysate and
its intact protein. Similarly, Claessens et al. [
34
] found no significant dierences when investigating
postprandial glucose levels following the ingestion of intact soy protein, whey protein and their
respective hydrolysates [
34
]. Thus, an insulin-associated response of protein intake on glucose is
probably most strongly associated with dierences in the protein content of BCAAs [35].
Proteins receive considerable attention in relation to satiety and high-protein diets have been
proposed to be ecient in weight-loss strategies [
36
]. In association with this, former studies have
indicated that collagen-rich AAs may exert unique eects on satiety. Thus, Veldhorst et al. (2009)
investigated the eects of dierent proteins on satiety and subsequent energy intake and found that
ad libitum intake was 20% lower after consumption of a breakfast gelatin compared to casein, soy or
whey [
37
]. Similar findings by Hochstenbach-Waelen et al. (2010) reported beneficial short-term eects
of gelatin on hunger suppression but no long-term eects on weight maintenance [
38
]. To the best of
our knowledge, these former studies have never been corroborated by studies investigating the eect
of collagen processing on satiety potential. We therefore decided to investigate if EHC and NC exerted
the same eects on feelings of satiety after intake. Our study revealed no significant dierences in
feelings of satiety after intake of EHC and NC suggesting that it is merely the AA composition rather
than molecular size that determine collagen-derived products’ satiety potential.
5. Conclusions
In conclusion,
1
H NMR-based metabolomics revealed that an enzymatic hydrolysis of collagen
was associated with an enhanced absorption rate of glycine, proline and hydroxyproline 20 min
post-ingestion. In addition, a greater AUC for glycine, proline and hydroxyproline for an enzymatically
hydrolyzed collagen hydrolysate (EHC) suggested a higher bioavailability for EHC compared to a
non-enzymatically hydrolyzed collagen hydrolysate (NC). However, an enzymatic hydrolysis did
not significantly aect the postprandial absorption rate of other AAs, indicating that an enzymatic
hydrolysis only aects absorption and bioavailability of the most abundant AAs of collagen.
Nutrients 2019,11, 1064 11 of 13
Supplementary Materials:
The following are available online at http://www.mdpi.com/2072-6643/11/5/1064/s1,
Table S1. Amino acid composition of the two collagen products included in the study, Figure S1: Plasma
concentration of methionine for EHC, NC and placebo, Figure S2. Plasma concentrations of alanine, asparagine,
glutamate, glutamine, histidine, serine, threonine and valine over time following ingestion of EHC, NC and
placebo, Figure S3. Plasma concentrations of arginine, leucine, isoleucine, lysine, phenylalanine and total AAs
over time following ingestion of EHC, NC and placebo, Figure S4. Plasma concentrations of EAA and BCAA over
time following ingestion of EHC, NC and placebo, Figure S5. (A) Plasma concentrations of glucose over time for
EHC, NC and placebo. (B) AUC is evaluated as incremental AUC from time
20 min (Pre) to 240 min, Figure S6.
Eects of EHC, NC and placebo on feelings of hunger over time.
Author Contributions:
Conceptualization, M.H. and H.C.B.; Methodology, M.H. and H.C.B.; Software, K.S. and
R.T.; Formal analysis, K.S.; Investigation, K.S. and M.O.; Writing—original draft preparation, K.S.; Writing—review
and editing, K.S., R.T., M.O., M.H. and H.C.B.; Supervision, M.H. and H.C.B.; Project administration, H.C.B.;
Funding acquisition, H.C.B.
Funding:
This research received partial financial support from Essentia Protein Solutions A/S. The financial
support consisted of support for running costs in connection with the intervention study and no funds were
received for salary.
Acknowledgments:
The authors thank the subjects for their eorts and compliance during the protocol. In addition,
the authors acknowledge Gitte Kaiser Hartvigsen, Janni Mosgaard Jensen and Nina Eggers for their assistance in
the laboratory work and NMR analyses.
Conflicts of Interest:
The authors declare no conflict of interest. Essentia Protein Solutions A/S had no role in the
design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the
decision to publish the results.
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©
2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/).
... The treatment options for OA are currently limited; however, several clinical trials have shown that ingestion of collagen hydrolysates (CHs) allows for decreased pain and increased mobility [12][13][14][15][16][17][18]. CH supplements contain a cocktail of peptides and amino acids (AAs); however, it is possible that these peptides are further broken down into Nutrients 2021, 13, 2720 2 of 17 bioactive peptides (BAPs) in the stomach and small intestine (SI) [19][20][21][22][23]. BAPs found in collagen products, such as Pro-Hyp, have been shown to decrease the loss of chondrocytes, prevent cartilage thinning, regulate genes associated with joint integrity, reduce the loss of subchondral bone as well as regulate inflammation by inhibiting cytokines such as tumor necrosis factor-α [24][25][26]. ...
... Studies investigating the bioavailability of CHs are needed, to verify if peptides from CH-GL formed during digestion are absorbed locally at the GI tract and survive after they permeate across the intestinal epithelium to enter the systemic blood circulation. Furthermore, investigations focusing on lower MW CH peptides are needed, as di-and tri-peptides from collagen have known bioactivity, and increased bioavailability compared to greater MW CH peptides [19,22,35]. Analysis identifying lower MW peptides continues to be a limitation of "peptide-centric" proteomic work, seeing as di-and tri-peptides are too small for sequencing. ...
... The treatment dose used in this study was based on the daily dose of the Genacol Original Formula ® that was shown to reduce joint pain in clinical trials [12,13,18]. Other clinical studies, however, have used much greater doses ranging from 5 to 35 g of hydrolyzed collagen products [14,15,17,19,81,82]. It is conceivable that with a higher initial dose of CHs, greater microbial fermentation could have occurred due to more substrate availability for fermentation with subsequent greater increases in SCFAs, BCFAs, colonic gas production and antioxidant capacity. ...
Article
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Osteoarthritis (OA), the most common form of arthritis, is associated with metabolic diseases and gut microbiome dysbiosis. OA patients often take supplements of collagen hydrolysates (CHs) with a high peptide content. Following digestion, some peptides escape absorption to induce prebiotic effects via their colonic fermentation to generate short-chain fatty acids (SCFAs), branched-chain fatty acids (BCFAs) and colonic gases (NH4 and H2S). The capacity of CHs to generate microbial metabolites is unknown. Proteomic analysis of two CHs (CH-GL and CH-OPT) demonstrated different native peptide profiles with increased peptide diversity after in vitro gastric and small intestinal digestion. Subsequent 24 h fermentation of the CH digests in a dynamic gastrointestinal (GI) digestion model containing human fecal matter showed that CH-OPT increased (p < 0.05) H2S, SCFAs (propionic, butyric and valeric acids), BCFAs, and decreased NH4 in the ascending colon reactor with no major changes seen with CH-GL. No major effects were observed in the transverse and descending vessels for either CH. These findings signify that CHs can induce prebiotic effects in the ascending colon that are CH dependent. More studies are needed to determine the physiological significance of CH-derived colonic metabolites, in view of emerging evidence connecting the gut to OA and metabolic diseases.
... Collagen hydrolysates (CHs) have been shown to provide multiple health benefits, which have been primarily attributed to their bioactive peptide (BAP) content [1][2][3]. These BAPs can be found in the hydrolysate products, although an increase in the diversity and content of peptides can result from gastrointestinal (GI) digestion [4,5]. The BAPs released after the digestion of collagen products, such as Pro-Hyp and Gly-Pro-Hyp, can possess multiple health properties, which include antimicrobial and antihypertensive effects, regulating inflammation, reducing pain associated with osteoarthritis, promoting bone synthesis, stimulating wound healing, as well as antioxidant properties and angiotensin-I-converting enzyme inhibitory effects [3,4,6,7]. ...
... These BAPs can be found in the hydrolysate products, although an increase in the diversity and content of peptides can result from gastrointestinal (GI) digestion [4,5]. The BAPs released after the digestion of collagen products, such as Pro-Hyp and Gly-Pro-Hyp, can possess multiple health properties, which include antimicrobial and antihypertensive effects, regulating inflammation, reducing pain associated with osteoarthritis, promoting bone synthesis, stimulating wound healing, as well as antioxidant properties and angiotensin-I-converting enzyme inhibitory effects [3,4,6,7]. ...
... The bioactivity of BAPs depends heavily on their ability to reach the general circulation intact after oral ingestion, otherwise called bioavailability [9]. Clinical studies have consistently shown that peptides generated from orally ingested collagen precursors, such as gelatin, or collagen hydrolysates, can reach the systemic circulation and be excreted in the urine [4,6,[10][11][12]. Importantly, the clinical efficacy of CHs has been demonstrated in multiple trials showing reduction of joint discomfort in athletes with functional knee problems and decreased joint pain in osteoarthritis patients [1,3,13]. ...
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Collagen hydrolysates (CHs) are composed of bioactive peptides (BAPs), which possess health enhancing properties. There is a knowledge gap regarding the bioavailability of these BAPs that involves intestinal transport and hepatic first pass effects. A simulated gastrointestinal model was used to generate digesta from two CHs (CH-GL and CH-OPT), which were applied to a novel transwell co-culture of human intestinal epithelium cell line-6 (HIEC-6) and hepatic (HepG2) cells to simulate in vivo conditions of absorption and first pass metabolism. Peptide transport, hepatic first pass effects, and bioavailability were determined by measuring BAPs (Gly-Pro, Hyp-Gly, Ala-Hyp, Pro-Hyp, Gly-Pro-Hyp) using an innovative capillary electrophoresis method. All peptides were transported across the intestinal cell layer to varying degrees with both CHs; however, Gly-Pro-Hyp was transported only with CH-GL, but not CH-OPT. Notable hepatic production was observed for Ala-Hyp with both CH treatments, and for Pro-Hyp and Gly-Pro with CH-GL only. All peptides were bioavailable (>10%), except for Gly-Pro-Hyp after CH-OPT. Overall, a high degree of transport and hepatic first pass effects on CH-derived BAPs were observed. Further research is needed to explore the hepatic mechanisms related to the production of BAPs and the bifunctional effects of the bioavailable BAPs noted in this study.
... Nutraceutical supplements derived from collagen can be made from beef, pork, or fish bones and skins, which undergo processing to increase the bioavailability of their amino acids and/or peptides; the enzymatic hydrolysis of collagens enhances the postprandial absorption of its processed components [22]. Processed and pre-digested collagen products are called collagen hydrolyzates CHs and are sold in the form of collagen capsules at pharmacies and health food suppliers. ...
... Due to its lower molecular weight, hydrolyzed collagen has been proposed to have higher bioavailability and solubility, and thus better absorption from the small intestine compared to undenatured collagen [22,29]. Absorption of orally administered hydrolyzed collagen has been evaluated by studying vascular-perfused rat intestine in situ. ...
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Purpose of Review Osteoarthritis (OA) is the most common forms of arthritis in the general population, accounting for more pain and functional disability than any other musculoskeletal disease. There are currently no approved disease modifying drugs for OA. In the absence of effective pharmacotherapy, many patients with OA turn to nutritional supplements and nutraceuticals, including collagen derivatives. Collagen hydrolyzates and ultrahydrolyzates are terms used to describe collagens that have been broken down into small peptides and amino acids in the presence of collagenases and high pressure. Recent Findings This article reviews the relevant literature and serves as a White Paper on collagen hydrolyzates and ultrahydrolyzates as emerging supplements often advertised to support joint health in OA. Collagen hydrolyzates have demonstrated some evidence of efficacy in a handful of small scale clinical trials, but their ability to treat and reverse advanced joint disease remains highly speculative, as is the case for other nutritional supplements. Summary The aim of this White Paper is to stimulate research and development of collagen-based supplements for patients with OA and other musculoskeletal diseases at academic and industrial levels. This White Paper does not make any treatment recommendations for OA patients in the clinical context, but simply aims to highlight opportunities for scientific innovation and interdisciplinary collaboration, which are crucial for the development of novel products and nutritional interventions based on the best available and published evidence.
... The krill protein used in this trial was enzymatically hydrolyzed, and we therefore expected the postprandial absorption of AAs to be faster compared to non-hydrolyzed protein isolates. The effect of hydrolyzing a protein isolate on postprandial AA bioavailability has previously been investigated in both whey, soy and collagen protein sources [27][28][29], but never in relation to krill protein in humans. In addition, to the best of our knowledge, studies investigating the effect of krill protein on appetite have not yet been reported. ...
... Since adequate protein intake is essential to humans [1], identifying new high-quality and sustainable protein sources is needed. Therefore, we investigated the acute postprandial bioavailability of AAs from a new KRILL protein hydrolysate as compared to two commercial protein sources, a SOY and a WHEY protein isolate. 1 H NMR-based metabolomics has previously been shown to be a useful tool to determine postprandial AA concentrations in blood [26,29]. Thus, in the present study, 1 H NMR spectroscopy was used to determine the serum AA concentrations during a 180-min postprandial period. ...
Article
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Background: adequate protein intake is essential to humans and, since the global demand for protein-containing foods is increasing, identifying new high-quality protein sources is needed. In this study, we investigated the acute postprandial bioavailability of amino acids (AAs) from a krill protein hydrolysate compared to a soy and a whey protein isolate. Methods: the study was a randomized, placebo-controlled crossover trial including ten healthy young males. On four non-consecutive days, volunteers consumed water or one of three protein-matched supplements: whey protein isolate, soy protein isolate or krill protein hydrolysate. Blood samples were collected prior to and until 180 min after consumption. Serum postprandial AA concentrations were determined using 1H NMR spectroscopy. Hunger and satiety were assessed using visual analogue scales (VAS). Results: whey and krill resulted in significantly higher AA concentrations compared to soy between 20-60 min and 20-40 min after consumption, respectively. Area under the curve (AUC) analyses revealed that whey resulted in the highest postprandial serum concentrations of essential AAs (EAAs) and branched chain AAs (BCAAs), followed by krill and soy, respectively. Conclusions: krill protein hydrolysate increases postprandial serum EAA and BCAA concentrations in a superior manner to soy protein isolate and thus might represent a promising future protein source in human nutrition.
... For instance, different hydrolyzation protocols are known to produce peptides of larger molecular weights. 95 Skov et al 103 recently demonstrated that ingestion of 35 g of enzymatically hydrolyzed collagen protein results in greater plasma glycine, proline, and hydroxyproline availability (incremental area under the curve) in comparison with the ingestion of 35 g of nonenzymatically hydrolyzed collagen. Work comparing digestibility after different hydrolyzation protocols of the same source of collagenderived protein is required to identify characteristics that maximize the uptake of collagen protein-derived amino acids into the circulation. ...
Article
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Collagen is the central structural component of extracellular connective tissue, which provides elastic qualities to tissues. For skeletal muscle, extracellular connective tissue transmits contractile force to the tendons and bones. Connective tissue proteins are in a constant state of remodeling and have been shown to express a high level of plasticity. Dietary-protein ingestion increases muscle protein synthesis rates. High-quality, rapidly digestible proteins are generally considered the preferred protein source to maximally stimulate myofibrillar (contractile) protein synthesis rates. In contrast, recent evidence demonstrates that protein ingestion does not increase muscle connective tissue protein synthesis. The absence of an increase in muscle connective tissue protein synthesis after protein ingestion may be explained by insufficient provision of glycine and/or proline. Dietary collagen contains large amounts of glycine and proline and, therefore, has been proposed to provide the precursors required to facilitate connective tissue protein synthesis. This literature review provides a comprehensive evaluation of the current knowledge on the proposed benefits of dietary collagen consumption to stimulate connective tissue remodeling to improve health and functional performance.
... (Ahead of Print) approximately 12% and 26% of proline and glycine, respectively (Skov et al., 2019). Recently, Shaw et al. (2017) demonstrated that human serum collected after gelatin ingestion promotes greater collagen synthesis when exposed to engineered tissue constructs in an ex vivo setting. ...
Article
Protein ingestion and exercise stimulate myofibrillar protein synthesis rates. When combined, exercise further increases the postprandial rise in myofibrillar protein synthesis rates. It remains unclear whether protein ingestion with or without exercise also stimulates muscle connective tissue protein synthesis rates. The authors assessed the impact of presleep protein ingestion on overnight muscle connective tissue protein synthesis rates at rest and during recovery from resistance-type exercise in older men. Thirty-six healthy, older men were randomly assigned to ingest 40 g intrinsically L -[1- ¹³ C]-phenylalanine and L -[1- ¹³ C]-leucine-labeled casein protein (PRO, n = 12) or a nonprotein placebo (PLA, n = 12) before going to sleep. A third group performed a single bout of resistance-type exercise in the evening before ingesting 40 g intrinsically-labeled casein protein prior to sleep (EX+PRO, n = 12). Continuous intravenous infusions of L-[ ring- ² H 5 ]-phenylalanine and L -[1- ¹³ C]-leucine were applied with blood and muscle tissue samples collected throughout overnight sleep. Presleep protein ingestion did not increase muscle connective tissue protein synthesis rates (0.049 ± 0.013 vs. 0.060 ± 0.024%/hr in PLA and PRO, respectively; p = .73). Exercise plus protein ingestion resulted in greater overnight muscle connective tissue protein synthesis rates (0.095 ± 0.022%/hr) when compared with PLA and PRO ( p < .01). Exercise increased the incorporation of dietary protein-derived amino acids into muscle connective tissue protein (0.036 ± 0.013 vs. 0.054 ± 0.009 mole percent excess in PRO vs. EX+PRO, respectively; p < .01). In conclusion, resistance-type exercise plus presleep protein ingestion increases overnight muscle connective tissue protein synthesis rates in older men. Exercise enhances the utilization of dietary protein-derived amino acids as precursors for de novo muscle connective tissue protein synthesis during overnight sleep.
Article
Collagen hydrolysates (CHs) are composed of bioactive peptides (BAPs) and amino acids (AAs), which contribute to their health enhancing properties. Post digestion profiling of CHs typically evaluates either BAP or AA content in blood but not within digests. Existing methods for peptide analysis are optimized for blood samples and rely on costly methods that require substantial sample preparation and data interpretation. A capillary electrophoresis (CE) method was developed as a rapid, cost effective, and reliable method for analysis of BAPs (Ala-Hyp, Pro-Hyp, Pro-Hyp-Gly, Gly-Pro-Hyp) within digests. Coupled to LC-MS, a hydrophilic interaction liquid chromatography (HILIC) column was used to quantify 19 AAs in digests, without derivatization. Two bovine CHs (CH-GL and CH-OPT) underwent in vitro digestion and their BAP and AA content was assessed. The Gly-Pro-Hyp and Pro-Hyp-Gly content was greater in CH-GL versus CH-OPT with values of 19.82 ± 4.25 and 8.969 ± 2.742 μg/mL respectively. The two CHs had distinct peptide profiles; 13 unidentified peptide peaks from each CH were not found in the other. No differences in AA content were observed. The present work describes sensitive and rapid methodology for concurrent analysis of BAPs and AAs after digestion of CHs, which can support further understanding of the bioactive components of CHs.
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Background Food proteins are a source of hydrolysates with potentially useful biological attributes. Bioactive peptides from food-derived proteins are released from hydrolysates using exogenous industrial processes or endogenous intestinal enzymes. Current in vitro permeability assays have limitations in predicting the oral bioavailability (BA) of bioactive peptides in humans. There are also difficulties in relating the low blood levels of food-derived bioactive peptides detected in preclinical in vivo models to pharmacodynamic read-outs relevant for humans. Scope and approach In this review, we describe in vitro assays of digestion, permeation, and metabolism as indirect predictors of the potential oral BA of hydrolysates and their constituent bioactive peptides. We discuss the relationship between industrial hydrolysis processes and the oral BA of hydrolysates and their peptide by-products. Key findings Hydrolysates are challenging for analytical detection methods due to capacity for enzymatic generation of peptides with novel sequences and also new modifications of these peptides during digestion. Mass spectrometry and peptidomics can improve the capacity to detect individual peptides released from complex hydrolysates in biological milieu.
Article
Background Functional and healthier foods has been seen as important elements in a healthier lifestyle. In order to obtain these products, reformulation is the main strategy to develop health-oriented solutions. In addition to the health benefits, increasing evidence have been suggesting that functional foods can also influence satiety, a major factor related to the frequency of food consumption and, consequently, the health. Scope and approach The aim of this review is to discuss the current evidence associating the consumption of functional foods with satiety. The studies with healthier and functional food products disclose the relation between protein, dietary fiber, fat, sugar, and salt with satiety. Key findings and conclusions Satiety is a complex and dynamic process that can be modified, at some extent, with food ingredients. Increasing dietary fiber and modifying fat and sugar composition are the main strategies to obtain healthier and functional foods with increased satiating effect. Major advances are still necessary to optimize the satiety of products with high protein content and clarify the relation between salt and satiety.
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Biodegradable polymers obtained from renewable resources, such as chitosan and collagen, are sustainable alternatives to develop environmentally friendly materials. Due to their abundance, biocompatibility and antimicrobial properties, chitosan and collagen could become a suitable source for food and biomedical applications. In particular, chitosan formulations are used for food packaging purposes to develop intelligent packaging with the aim of providing information about the quality of the packaged product or to prepare active packaging and extend food shelf life. In this regard, chitosan nanoparticles can be used to provide a sustained release of active substances. Regarding collagen, denatured collagen or gelatin is prevalently used in food industry as a food additive, microencapsulating agent or biodegradable packaging material due to its rheological properties and physical versatility. In turn, collagen-derived peptides have revealed antioxidant and antihypertensive activity, among other health beneficial effects for cosmetic and nutraceutical applications. Additionally, collagen is widely used in tissue engineering, also combined with chitosan, to achieve the functional properties required for specific applications in the biomedical field. In this sense, collagen/chitosan scaffolds have been used for bone, cartilage and skin regeneration. This research in the design and processing of materials based on proteins and polysaccharides is leading to great advances in food and biomedical fields.
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Collagen hydrolysates are dietary supplements used for nutritional and medical purposes. They are complex mixtures of low‐molecular‐weight peptides obtained from the enzymatic hydrolysis of collagen, which provide intrinsic batch‐to‐batch heterogeneity. In consequence, the quality of these products, which is related to the reproducibility of their mass distribution pattern, should be addressed. Here, we propose an analytical approach to determine the peptide pattern as a quality attribute of Colagenart®, a product containing collagen hydrolysate. In addition, we evaluated the safety by measuring the viability of two cell lines exposed to the product. The consistency of peptide distribution was determined using Size Exclusion Chromatography (SEC), Mass Spectrometry coupled to a reversed phase UPLC system (MS‐RP‐UPLC), and Shaped‐pulse off‐resonance water‐presaturation proton nuclear magnetic resonance spectrometry [1Hwater_presat NMR]. The mass distribution pattern determined by SEC was in a range from 1.35 to 17 kDa, and from 2 to 14 kDa by MS‐RP‐UPLC. [1Hwater_presat NMR] showed the detailed spin‐systems of the collagen hydrolysates components by global assignment of backbone Hα and NH, as well as side‐chain proton resonances. Additionally, short‐range intraresidue connectivity pathways of identified spin‐regions were obtained by a 2D homonuclear shift correlation Shaped‐pulse solvent suppression COSY scheme. Safety analysis of Colagenart® was evaluated in CaCo‐2 and HepG2 cells at 2.5 and 25 μg/mL and no negative effects were observed. The results demonstrated batch‐to‐batch reproducibility, which evinces the utility of this approach to establish the consistency of the quality attributes of collagen hydrolysates. We propose state‐of‐the art analytical methodologies (SEC, MS, and NMR) to evaluate peptide profile and composition of collagen hydrolysates as quality attributes. These methodologies are suitable to be implemented for quality control purposes.
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The study investigated the acute effects of meals containing either salmon or veal in combination with carbohydrates with high or low glycemic index (GI) on diet-induced thermogenesis (DIT) (primary endpoint), appetite sensations, and energy intake (EI). Twenty-five overweight men and women ingested four iso-caloric test meals: salmon with mashed potatoes (high GI) (SM), salmon with wholegrain pasta (low GI) (SP), veal with mashed potatoes (VM) and veal with wholegrain pasta (VP). Energy expenditure was measured in the fasting state and six times postprandially for 25 min with 5-min breaks between each measurement. Appetite sensations were measured every 30 min. Blood samples, from arterialized venous blood, were drawn every 20 min until an ad libitum buffet-style lunch was served 3.5 h later. DIT was 40% higher after the SM meal compared to the SP meal (p = 0.002). Prospective food consumption was lower after the SM meal compared with the VP meal (p = 0.01). There were no differences in satiety, hunger, fullness, or ad libitum EI between the test meals (all p > 0.05). In conclusion, salmon with high GI carbohydrates increased DIT compared to salmon with low GI carbohydrates. This indicates that DIT is sensitive to the GI of the carbohydrates after intake of salmon but not veal.
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The current pilot study investigates whether oral supplementation of specific collagen peptides improves symptoms and tendon vascularisation in patients with chronic mid-portion Achilles tendinopathy in combination with structured exercise. Participants were given a placebo or specific collagen peptides (TENDOFORTE®) in combination with a bi-daily calf-strengthening program for 6 months. Group AB received specific collagen peptides for the first 3 months before crossing over to placebo. Group BA received placebo first before crossing over to specific collagen peptides. At baseline (T1), 3 (T2) and 6 (T3) months, Victorian Institute of Sports Assessment–Achilles (VISA-A) questionnaires and microvascularity measurements through contrast-enhanced ultrasound were obtained in 20 patients. Linear mixed modeling statistics showed that after 3 months, VISA-A increased significantly for group AB with 12.6 (9.7; 15.5), while in group BA VISA-A increased only by 5.3 (2.3; 8.3) points. After crossing over group AB and BA showed subsequently a significant increase in VISA-A of, respectively, 5.9 (2.8; 9.0) and 17.7 (14.6; 20.7). No adverse advents were reported. Microvascularity decreased in both groups to a similar extent and was moderately associated with VISA-A (Rc2:0.68). We conclude that oral supplementation of specific collagen peptides may accelerate the clinical benefits of a well-structured calf-strengthening and return-to-running program in Achilles tendinopathy patients.
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Background: Increased amino acid availability stimulates muscle protein synthesis (MPS), which is critical for maintaining or increasing muscle mass when combined with training. Previous research suggests that whey protein is superior to soy protein in regard to stimulating MPS and muscle mass. Nevertheless, with respect to a future lack of dietary protein and an increasing need for using eco-friendly protein sources it is of great interest to investigate the quality of alternative protein sources, like insect protein. Objective: Our aim was to compare the postprandial amino acid (AA) availability and AA profile in the blood after ingestion of protein isolate from the lesser mealworm, whey isolate, and soy isolate. Design: Six healthy young men participated in a randomized cross-over study and received three different protein supplementations (25 g of crude protein from whey, soy, insect or placebo (water)) on four separate days. Blood samples were collected at pre, 0 min, 20 min, 40 min, 60 min, 90 min, and 120 min. Physical activity and dietary intake were standardized before each trial, and participants were instructed to be fasting from the night before. AA concentrations in blood samples were determined using ¹H NMR spectroscopy. Results: A significant rise in blood concentration of essential amino acids (EAA), branched-chain amino acids (BCAA) and leucine was detected over the 120 min period for all protein supplements. Nevertheless, the change in AA profile was significantly greater after ingestion of whey than soy and insect protein (p < 0.05). Area under the curve (AUC) analysis and AA profile revealed comparable AA concentrations for soy and insect protein, whereas whey promoted a ~97% and ~140% greater AUC value than soy and insect protein, respectively. A tendency towards higher AA concentrations beyond the 120 min period was observed for insect protein. Conclusion: We report that ingestion of whey, soy, and insect protein isolate increases blood concentrations of EAA, BCAA, and leucine over a 120 min period (whey > insect = soy). Insect protein induced blood AA concentrations similar to soy protein. However, a tendency towards higher blood AA concentrations at the end of the 120 min period post ingestion was observed for insect protein, which indicates that it can be considered a "slow" digestible protein source.
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Collagen synthesis is severely diminished in osteoarthritis; thus, enhancing it may help the regeneration of cartilage. This requires large amounts of glycine, proline and lysine. Previous works of our group have shown that glycine is an essential amino acid, which must be present in the diet in large amounts to satisfy the demands for collagen synthesis. Other authors have shown that proline is conditionally essential. In this work we studied the effect of these amino acids on type II collagen synthesis. Bovine articular chondrocytes were cultured under a wide range of different concentrations of glycine, proline and lysine. Chondrocytes were characterized by type II collagen immunocytochemistry of confluence monolayer cultures. Cell growth and viability were assayed by trypan blue dye exclusion method. Type II collagen was measured in the monolayer, every 48 hours for 15 days by ELISA. Increase in concentrations of proline and lysine in the culture medium enhances the synthesis of type II collagen at low concentrations, but these effects decay before 1.0 mM. Increase of glycine as of 1.0 mM exceeds these effects and this increase continues more persistently by 60-75%. Since the large effects produced by proline and lysine are within the physiological range, while the effect of glycine corresponds to a much higher range, these results demonstrated a severe glycine deficiency for collagen synthesis. Thus, increasing glycine in the diet may well be a strategy for helping cartilage regeneration by enhancing collagen synthesis, which could contribute to the treatment and prevention of osteoarthritis.
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Following an initial ankle sprain it is not unlikely that chronic ankle instability (CAI) will develop. CAI is associated with impaired perceived functional and mechanical properties of the ligaments. Nutritional supplementation with collagen peptides has been shown to improve the functional and mechanical properties of the connective tissue. The purpose of this study was to investigate the effectiveness of specific collagen peptide supplementation (SCP) to improve ankle stability in athletes with CAI. 50 male and female athletes with CAI completed a randomized, double-blinded and placebo-controlled study with a daily oral administration of either 5 g SCP or 5 g placebo (Maltodextrin) over a period of six months. Both, the Cumberland Ankle Instability Tool (CAIT) and the German version of the Foot and Ankle Ability Measure (FAAM-G) were used to measure the subjective perceived function of the ankle. Additionally, the mechanical stability was determined by measuring the ankle stiffness by an ankle arthrometer. Finally, a three-month follow-up was performed. ANOVA analysis indicated that the subjective ankle stability was improved in both the CAIT (p < 0.001) and the FAAM-G (p < 0.001) following SCP supplementation compared with placebo. No significant changes between the groups were detected in the results of the ankle arthrometer. After six month the subjective report of the ankle stability function significantly improved and the three month follow-up revealed a significant decline in the number of ankle joint injuries (p < 0.05). These data support the concept that specific collagen peptide supplementation in athletes with chronic ankle instability results in significant improvements in subjective perceived ankle stability. The reduction in the re-injury rate of ankle sprains in the follow-up period suggests that these findings have clinical relevance.
Article
Purpose Osteoarthritis (OA) is one of the most common causes of disability and a prevalent chronic disease. The use of collagen is growing due to the satisfactory results in the treatment of OA. However, the possible beneficial effects of collagen for the treatment of OA are currently controversial. The aim of the present meta-analysis was to evaluate the effect of collagen-based supplements on OA symptoms. Methods PubMed-Medline, Scopus, and Google Scholar databases were searched for randomized placebo-controlled trials evaluating the effect of orally administered collagen on OA symptoms using the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) scale and/or the Visual Analog Scale (VAS). Meta-analysis was conducted using a random-effects model and a generic inverse variance method. Heterogeneity was tested using the I² statistic index. Results Collagen treatment showed a significant reduction in the score of total WOMAC index (WMD − 8.00; 95% CI − 13.04, − 2.95; p = 0.002). After subgroup analysis of the WOMAC subscores, the collagen supplementation revealed a significant decrease in the stiffness subscore (WMD − 0.41; 95% CI − 0.74, − 0.08; p = 0.01), whereas the pain (WMD − 0.22; 95% CI − 1.58, 1.13; p = 0.75) and functional limitation (WMD − 0.62; 95% CI − 5.77, 4.52; p = 0.81) subscores did not have significant differences. Finally, a significant reduction was found in the VAS score after collagen administration (WMD − 16.57; 95% CI − 26.24, − 6.89; p < 0.001). Conclusion The results of this meta-analysis showed that collagen is effective in improving OA symptoms by the decrease of both total WOMAC index and VAS score.
Article
Patellar tendinopathy is one of the most common afflictions in jumping sports. This case study outlines the rehabilitation of a professional basketball player diagnosed by MRI with a central core patellar tendinopathy within the proximal enthesis. The player undertook a nutrition and strength based rehabilitation program combining gelatin ingestion and heavy isometric loading of the patella designed to produce significant stress relaxation as part of their competition schedule and a whole body training plan. On follow up one and a half years into the program an independent orthopedic surgeon declared the tendon normal on MRI. Importantly, the improved MRI results were associated with a decrease in pain and improved performance. This case study provides evidence that a nutritional intervention combined with a rehabilitation program that uses stress relaxation can improve clinical outcomes in elite athletes.
Article
Aging is a multifactorial and natural process that causes physiological changes in organs, tissues and cells over time. In the skin and cartilage, aging leads to a decrease in the synthesis and changes in the arrangement of proteoglycans and collagen, in addition to the loss of glycosaminoglycans, which are responsible for the integrity and health of these tissues. We hypothesized that daily oral supplementation with a liquid nutraceutical containing hydrolyzed fish collagen, vitamins, antioxidants and other active ingredients could improve skin texture and elasticity, and in addition have a protective effect on joint health. A double-blind, randomized, placebo-controlled clinical trial was conducted on 120 subjects who consumed either the test product or placebo on a daily basis for 90 days. Subjects consuming the test product had an overall significant increase in skin elasticity (+40%; P <.0001) when compared to placebo. Histological analysis of skin biopsies revealed positive changes in the skin architecture, with a reduction in solar elastosis and improvement in collagen fiber organization in the test product group. As reported in the self-perception questionnaires, these results were confirmed by the subjects’ own perceptions in that participants agreed their skin was more hydrated and more elastic. In addition, the consumption of the test product reduced joint pain by −43% and improved joint mobility by +39%. Oral supplementation with collagen bioactive peptides combined with chondroitin sulphate, glucosamine, L-carnitine, vitamins, and minerals significantly improved the clinical parameters related to skin aging and joint health, and therefore, might be an effective solution to slow down the hallmarks of aging.
Article
BACKGROUND Casein and whey proteins differ in amino acid composition and absorption rate; however the absorption rate of casein can be increased to mimic that of whey proteins by exogenous hydrolysis. In view of these compositional differences we studied the metabolic responses to intake of casein, hydrolyzed casein and whey proteins in overweight and moderately obese men and women by investigating select urinary and blood plasma metabolites. RESULTS A total of 21 urinary and 23 plasma metabolites were identified by NMR spectroscopy. The postprandial plasma metabolites revealed a significant diet‐time interaction for isoleucine (P = 0.001) and tyrosine (P = 0.001). The level of isoleucine and tyrosine peaked 90 min postprandial with a 1.4‐fold difference following intake of whey proteins compared to casein and hydrolyzed casein, respectively. A 1.2‐fold higher urinary level of lactate was observed after intake of whey proteins compared to intake of intact casein (P < 0.01). CONCLUSION The plasma metabolites revealed different amino acid profiles reflecting the amino acid composition of casein and whey proteins. Furthermore, the results support that casein hydrolysates do neither affect the postprandial amino acid absorption rate nor the amino acid level compared to that of intact casein. The urinary lactate increases following whey protein intake might indicate a higher metabolism of glucogenic amino acids.