2.1. Effects of Oral Administration of CNFs and SDACNFs on Plasma Metabolites
In this study, capillary electrophoresis-time-of-flight mass spectrometry (CE-TOFMS) revealed 224 peaks (129 cations and 95 anions). Liquid chromatography (LC)-TOFMS detected 90 peaks (49 positive and 41 negative). The results of the hierarchical cluster analysis (HCA) are shown in
Figure 1. Relative areas of metabolites after oral administration of CNFs, SDACNFs, chitin, and cellulose nanofibers (CLNFs) were compared with those of the control group. All
p-values were determined using Welch’s
t-test. The ratios of the metabolite concentrations to that of the control group are shown in
Table 1.
Figure 1.
Results of hierarchical cluster analysis of plasma metabolites. An ordinate and an abscissa indicate peaks and names of experimental groups, respectively. Hierarchical cluster analysis (HCA) was performed for peaks, and distance between peaks are shown as tree diagrams. Denser green and red indicate shorter and longer distances than those of the control group, respectively. CNF, chitin nanofiber; SDACNF, surface-deacetylated CNF; CLNF, cellulose nanofiber.
Figure 1.
Results of hierarchical cluster analysis of plasma metabolites. An ordinate and an abscissa indicate peaks and names of experimental groups, respectively. Hierarchical cluster analysis (HCA) was performed for peaks, and distance between peaks are shown as tree diagrams. Denser green and red indicate shorter and longer distances than those of the control group, respectively. CNF, chitin nanofiber; SDACNF, surface-deacetylated CNF; CLNF, cellulose nanofiber.
Table 1.
Ratios of relative areas of nanofiber metabolites to control group.
Table 1.
Ratios of relative areas of nanofiber metabolites to control group.
Compound | Comparative Analysis |
---|
CNF/Control | SDACNF/Control | Chitin/Control | CLNF/Control |
---|
Ratio | p-Value | Ratio | p-Value | Ratio | p-Value | Ratio | p-Value |
---|
ATP | 2.9 | 0.003 | 1.2 | 0.516 | 1.5 | 0.070 | 1.5 | 0.122 |
2,3-Diphosphoglyceric acid | 2.7 | 0.003 | 1.1 | 0.805 | 1.5 | 0.238 | 1.5 | 0.277 |
ADP | 2.2 | 0.003 | 1.1 | 0.678 | 1.4 | 0.201 | 1.3 | 0.346 |
5-Hydroxytryptophan | 2.0 | 0.003 | 1.8 | 0.151 | 2.1 | 0.065 | 2.0 | 0.051 |
ADP-ribose | 1.8 | 0.003 | 1.2 | 0.429 | 1.1 | 0.689 | 1.3 | 0.447 |
UTP | 1.7 | 0.028 | 1.2 | 0.161 | 1.0 | 0.986 | 1.4 | N.A. |
Cystine | 1.7 | 0.003 | 1.6 | 0.009 | 3.4 | 1.1 × 10−6 | 2.3 | 5.2 × 10−4 |
Serotonin | 1.6 | 0.014 | 1.4 | 0.238 | 1.6 | 0.056 | 1.3 | 0.240 |
Urocanic acid | 1.6 | 0.058 | 0.8 | 0.013 | 1.0 | 0.750 | 1.2 | 0.202 |
2,6-Diaminopimelic acid | 1.5 | 0.043 | 1.1 | N.A. | 1.7 | 0.040 | 2.4 | 0.253 |
1-Methylhistamine | 1.5 | 0.031 | 1.1 | 0.301 | 1.0 | 0.764 | 1.2 | 0.610 |
2-Aminoisobutyric acid | 1.4 | 0.023 | 0.9 | 0.470 | 1.0 | 0.723 | 1.3 | 0.041 |
2-Hydroxyisobutyric acid | 1.4 | 0.017 | 1.0 | 0.618 | 1.1 | 0.208 | 1.2 | 0.353 |
N5-Ethylglutamine | 1.3 | 0.041 | 0.9 | 0.566 | 0.8 | 0.104 | 0.7 | 0.080 |
Thiamine | 1.3 | 0.018 | 1.0 | 0.950 | 0.7 | 0.095 | 0.9 | 0.617 |
Putrescine | 1.2 | 0.002 | 1.0 | 0.707 | 1.1 | 0.469 | 1.3 | 0.305 |
Trimethylamine
N-oxide | 1.1 | 0.696 | 1.6 | 0.013 | 1.4 | 0.238 | 2.2 | 0.109 |
N-Acetylglycine | 1.1 | 0.324 | 0.9 | 0.040 | 1.0 | 0.721 | 1.2 | 0.054 |
FA(17:1) | 0.9 | 0.544 | 0.7 | 0.022 | 0.7 | 0.013 | 0.9 | 0.253 |
Cystathionine | 0.9 | 0.223 | 0.8 | 0.028 | 1.1 | 0.485 | 0.9 | 0.633 |
Stearic acid | 0.9 | 0.349 | 0.8 | 0.033 | 0.8 | 0.016 | 0.9 | 0.353 |
Glycolic acid | 0.8 | 0.026 | 1.0 | 0.864 | 1.0 | 0.826 | 1.0 | 0.934 |
Glu | 0.8 | 0.038 | 0.8 | 0.101 | 0.8 | 0.031 | 0.8 | 0.042 |
Betaine | 0.8 | 0.046 | 0.9 | 0.220 | 1.1 | 0.372 | 0.9 | 0.184 |
Argininosuccinic acid | 0.7 | 0.027 | 0.7 | 0.024 | 0.6 | 0.005 | 0.8 | 0.429 |
AC(18:0) | 0.7 | 0.007 | 0.6 | 0.002 | 0.5 | 4.9 × 10−4 | 0.8 | 0.140 |
Palmitoylcarnitine | 0.7 | 0.138 | 0.4 | 0.019 | 0.4 | 0.020 | 0.7 | 0.221 |
AC(18:1) | 0.6 | 0.080 | 0.4 | 0.019 | 0.5 | 0.029 | 0.8 | 0.525 |
AC(18:2) | 0.5 | 0.015 | 0.4 | 0.008 | 0.5 | 0.013 | 0.8 | 0.431 |
Taurodeoxycholic acid | 0.4 | 0.028 | 0.4 | 0.025 | 0.5 | 0.056 | 0.5 | 0.083 |
20α-Hydroxyprogesterone | 0.4 | 0.013 | 0.7 | 0.176 | 0.8 | 0.375 | 0.9 | 0.734 |
Taurocholic acid | 0.3 | 0.028 | 0.9 | 0.872 | 0.4 | 0.042 | 0.6 | 0.122 |
The results of the determination of metabolite concentrations are shown in
Table 2. The results of relative areas are shown in
Table 3. In the CNFs group, plasma levels of adenosine triphosphate (ATP) and adenosine diphosphate (ADP) were significantly higher than they were in the control group. On the other hand, no increase in plasma levels of ATP and ADP were observed in the SDACNF, chitin, and CLNF groups. ATP is the source of energy in living cells and acts as an allosteric effector of numerous cell processes such as active transport, nucleic acid synthesis, muscle activity, and movement [
25,
26,
27,
28,
29]. Endogenous ATP and ADP can be released into the extracellular milieu as a consequence of several mechanisms including cell lysis, opening channel-like pathways, and exocytosis of secretory vesicles [
30,
31]. In addition to their roles as intracellular energy transporters, the purine nucleotides ATP and ADP are important extracellular signaling molecules [
32].
Figure 2 shows the results of plasma metabolite levels related to the tricarboxylic acid cycle (TCA) cycle. Although there was no significant difference, plasma levels of metabolites related the TCA cycle were higher in the treated groups than they were in the control group. These results might indicate that oral administration of CNFs increase plasma ATP level by affecting the TCA cycle. The results revealed that only the CNF group showed plasma serotonin (5-hydroxytryptamine, 5-HT) and 5-hydroxytryptophan (5-HTP) levels that were significantly higher than those of the control group (
Table 1). Tryptophan is converted to 5-HT via 5-HTP [
33]. It is well known that 5-HT act as a brain neurotransmitter. Moreover, it is an important regulatory factor in the gastrointestinal tract (GIT) and other organ systems. More than 90% of the body’s 5-HT is synthesized in the gut [
34]. More recently, it was reported that some gut microbiota synthesize 5-HT [
35] while some others secrete ATP [
29]. The results of the present study might indicate that CNFs stimulate the functions of gut microbiota.
Figure 3 and
Figure 4 show the effects of oral administration of CNFs and SDACNFs on plasma acyl-carnitine (AC) and fatty acid (FA) levels. The SDACNFs group showed plasma levels of FA(17:1), FA(22:4), AC(18:0), AC(18:1), and AC(18:2) that were significantly lower than those of the control group (
Table 1). In particular, long-chain ACs perform important carrier functions during FA β-oxidation. First, carnitine palmitoyltransferase I (CPT I) catalyzes the transfer of acyl groups from acyl-coenzyme A to free carnitine to form ACs outside the mitochondrial membrane. Next, the ACs are transported to the mitochondrial matrix where the acyl groups are transferred to coenzyme A to form acyl-coenzyme A, which can participate in β-oxidation to supply energy [
36]. When incomplete β-oxidation of fatty acids occurs within the mitochondria, ACs accumulate in the plasma [
36]. Plasma AC concentrations have been shown to be elevated in individuals who are obese with either impaired glucose tolerance or diabetes [
37,
38]. Some reports described that intake of high fat diet increase blood AC levels in both experimental animals and humans [
39,
40,
41]. It is indicated that plasma free FA levels are elevated in obese individuals [
42], most likely due to increased free FA release associated with an expansion in fat mass [
43,
44]. In this study, significant decreases of plasma levels of FA(17:1), FA(22:4), AC(18:0), AC(18:1), and AC(18:2) were observed. The results of the present study indicate that one possible mechanism of the anti-obesity effects of orally administered SDACNFs might involve the tuning of lipid metabolism. To the best of our knowledge, there are few reports that describe the relationship between oral administration of chitin derivatives and changes in plasma ACs or FAs. Further study must be perform to this point.
Figure 3 and
Figure 4 show the effects of oral administration of CNFs and SDACNFs on plasma acyl-carnitine (AC) and fatty acid (FA) levels. The SDACNFs group showed plasma levels of FA(17:1), FA(22:4), AC(18:0), AC(18:1), and AC(18:2) that were significantly lower than those of the control group (
Table 1). In particular, long-chain ACs perform important carrier functions during FA β-oxidation. First, carnitine palmitoyltransferase I (CPT I) catalyzes the transfer of acyl groups from acyl-coenzyme A to free carnitine to form ACs outside the mitochondrial membrane. Next, the ACs are transported to the mitochondrial matrix where the acyl groups are transferred to coenzyme A to form acyl-coenzyme A, which can participate in β-oxidation to supply energy [
36]. When incomplete β-oxidation of fatty acids occurs within the mitochondria, ACs accumulate in the plasma [
36]. Plasma AC concentrations have been shown to be elevated in individuals who are obese with either impaired glucose tolerance or diabetes [
37,
38]. Some reports described that intake of high fat diet increase blood AC levels in both experimental animals and humans [
39,
40,
41]. It is indicated that plasma free FA levels are elevated in obese individuals [
42], most likely due to increased free FA release associated with an expansion in fat mass [
43,
44]. In this study, significant decreases of plasma levels of FA(17:1), FA(22:4), AC(18:0), AC(18:1), and AC(18:2) were observed. The results of the present study indicate that one possible mechanism of the anti-obesity effects of orally administered SDACNFs might involve the tuning of lipid metabolism. To the best of our knowledge, there are few reports that describe the relationship between oral administration of chitin derivatives and changes in plasma ACs or FAs. Further study must be perform to this point.
Table 2.
Relative areas of plasma metabolite among experimental groups.
Table 2.
Relative areas of plasma metabolite among experimental groups.
Compound Name | Control | CNF | SDACNF | Chitin | CLNF |
---|
Mean | SD | Mean | SD | Mean | SD | Mean | SD | Mean | SD |
---|
ATP | 8.0 × 10−4 | 3.1 × 10−4 | 2.3 × 10−3 | 7.5 × 10−4 | 9.6 × 10−4 | 4.3 × 10−4 | 1.2 × 10−3 | 3.0 × 10−4 | 1.2 × 10−3 | 4.5 × 10−4 |
2,3-Diphosphoglyceric acid | 6.4 × 10−4 | 1.5 × 10−4 | 1.7 × 10−3 | 5.2 × 10−4 | 6.7 × 10−4 | 2.6 × 10−4 | 9.6 × 10−4 | 5.2 × 10−4 | 9.8 × 10−4 | 5.9 × 10−4 |
ADP | 7.8 × 10−4 | 3.0 × 10−4 | 1.7 × 10−3 | 4.4 × 10−4 | 8.8 × 10−4 | 4.3 × 10−4 | 1.1 × 10−3 | 3.5 × 10−4 | 1.1 × 10−3 | 5.1 × 10−4 |
5-Hydroxytryptophan | 8.4 × 10−6 | 1.9 × 10−6 | 1.7 × 10−5 | 4.0 × 10−6 | 1.5 × 10−5 | 6.9 × 10−6 | 1.8 × 10−5 | 8.5 × 10−6 | 1.7 × 10−5 | 5.6 × 10−6 |
ADP-ribose | 1.2 × 10−4 | 4.0 × 10−5 | 2.2 × 10−4 | 3.6 × 10−5 | 1.5 × 10−4 | 5.9 × 10−5 | 1.3 × 10−4 | 2.9 × 10−5 | 1.7 × 10−4 | 1.1 × 10−4 |
UTP | 9.4 × 10−5 | 1.8 × 10−5 | 1.6 × 10−4 | 5.3 × 10−5 | 1.2 × 10−4 | 1.5 × 10−6 | 9.4 × 10−5 | 2.1 × 10−5 | 1.3 × 10−4 | N.A. |
Cystine | 7.4 × 10−4 | 2.1× 10−4 | 1.2 × 10−4 | 1.2 × 10−4 | 1.2 × 10−4 | 1.4 × 10−4 | 2.5 × 10−3 | 2.1 × 10−4 | 1.7 × 10−3 | 2.9 × 10−4 |
Serotonin | 1.3 × 10−4 | 4.7 × 10−5 | 2.1 × 10−4 | 3.1 × 10−5 | 1.8 × 10−4 | 7.5 × 10−5 | 2.1 × 10−4 | 6.3 × 10−5 | 1.7 × 10−4 | 5.2 × 10−5 |
Urocanic acid | 2.1 × 10−4 | 2.3 × 10−5 | 3.3 × 10−4 | 1.2 × 10−4 | 1.7 × 10−4 | 1.7 × 10−5 | 2.1 × 10−4 | 3.4 × 10−5 | 2.5 × 10−4 | 6.5 × 10−5 |
32,6-Diaminopimelic acid | 5.8 × 10−5 | 8.4 × 10−6 | 8.6 × 10−5 | 1.2 × 10−5 | 6.6 × 10−5 | N.A. | 9.8 × 10−5 | 8.2 × 10−6 | 1.4 × 10−4 | 8.6 × 10−5 |
1-Methylhistamine | 1.2 × 10−4 | 1.1 × 10−5 | 1.8 × 10−4 | 4.8 × 10−5 | 1.4 × 10−4 | 2.4 × 10−5 | 1.2 × 10−4 | 1.6 × 10−5 | 1.4 × 10−4 | 7.8 × 10−5 |
2-Aminoisobutyric acid | 2.4 × 10−3 | 5.2 × 10−4 | 3.4 × 10−3 | 6.7 × 10−4 | 2.2 × 10−3 | 4.5 × 10−4 | 2.3 × 10−3 | 3.2 × 10−4 | 3.2 × 10−3 | 4.8 × 10−4 |
2-Hydroxyisobutyric acid | 5.2 × 10−4 | 9.8 × 10−5 | 7.3 × 10−4 | 1.3 × 10−4 | 5.0 × 10−4 | 5.0 × 10−4 | 6.0 × 10−4 | 7.4 × 10−5 | 6.0 × 10−4 | 1.5 × 10−4 |
N5-Ethylglutamine | 1.3 × 10−3 | 3.4 × 10−4 | 1.7 × 10−3 | 2.2 × 10−4 | 1.2 × 10−3 | 2.1 × 10−4 | 9.8 × 10−4 | 1.2 × 10−4 | 9.4 × 10−4 | 1.7 × 10−4 |
Thiamine | 1.2 × 10−4 | 2.1 × 10−5 | 1.6 × 10−4 | 1.6 × 10−5 | 1.3 × 10−4 | 7.9 × 10−5 | 9.3 × 10−5 | 2.9 × 10−5 | 1.2 × 10−4 | 2.9 × 10−5 |
Putrescine | 1.5 × 10−4 | 1.1 × 10−5 | 1.8 × 10−4 | 1.0 × 10−5 | 1.5 × 10−4 | 2.4 × 10−5 | 1.6 × 10−4 | 1.8 × 10−5 | 1.9 × 10−4 | 7.5 × 10−5 |
Trimethylamine N-oxide | 4.6 × 10−3 | 7.7 × 10−4 | 5.0 × 10−3 | 2.6 × 10−3 | 7.4 × 10−3 | 1.6 × 10−3 | 6.6 × 10−3 | 3.2 × 10−3 | 9.9 × 10−3 | 5.8 × 10−3 |
N-Acetylglycine | 1.7 × 10−4 | 1.5 × 10−5 | 1.8 × 10−4 | 1.7 × 10−5 | 1.5 × 10−4 | 1.5 × 10−5 | 1.7 × 10−4 | 2.7 × 10−5 | 2.2 × 10−4 | 3.5 × 10−5 |
FA(17:1) | 3.2 × 10−5 | 4.6 × 10−6 | 3.0 × 10−5 | 7.8 × 10−6 | 2.3 × 10−5 | 5.1 × 10−6 | 2.2 × 10−5 | 5.2 × 10−6 | 2.8 × 10−5 | 5.4 × 10−6 |
Ser | 4.3 × 10−2 | 9.7 × 10−3 | 4.0 × 10−2 | 5.6 × 10−3 | 4.0 × 10−2 | 4.3 × 10−3 | 3.7 × 10−2 | 4.6 × 10−3 | 3.7 × 10−2 | 6.1 × 10−3 |
Cytidine | 5.9 × 10−4 | 1.1 × 10−4 | 5.4 × 10−4 | 1.6 × 10−4 | 5.4 × 10−4 | 9.3 × 10−5 | 6.2 × 10−4 | 5.3 × 10−5 | 7.0 × 10−4 | 1.3 × 10−4 |
Cystathionine | 2.6 × 10−4 | 2.1 × 10−5 | 2.4 × 10−4 | 2.7 × 10−5 | 2.2 × 10−4 | 2.6 × 10−4 | 2.8 × 10−4 | 5.7 × 10−5 | 2.5 × 10−4 | 4.0 × 10−5 |
Stearic acid | 1.1 × 10−3 | 1.2 × 10−4 | 9.7 × 10−4 | 2.0 × 10−4 | 8.7 × 10−4 | 1.2 × 10−4 | 8.1 × 10−4 | 1.4 × 10−4 | 9.7 × 10−4 | 1.9 × 10−4 |
Glyoxylic acid | 1.4 × 10−4 | 4.0 × 10−4 | 1.3 × 10−4 | 2.0 × 10−5 | 1.4 × 10−4 | 2.4 × 10−4 | 1.4 × 10−4 | 2.2 × 10−5 | 1.9 × 10−4 | 5.8 × 10−5 |
Glu | 1.4 × 10−2 | 2.4 × 10−3 | 1.1 × 10−2 | 8.0 × 10−4 | 1.1 × 10−2 | 2.3 × 10−3 | 1.1 × 10−2 | 1.1 × 10−3 | 1.1 × 10−2 | 9.1 × 10−4 |
Betaine | 5.1 × 10−2 | 9.3 × 10−3 | 3.9 × 10−2 | 5.9 × 10−3 | 4.4 × 10−2 | 4.7 × 10−3 | 5.6 × 10−2 | 1.0 × 10−2 | 4.4 × 10−2 | 4.5 × 10−3 |
Argininosuccinic acid | 1.7 × 10−3 | 2.9 × 10−5 | 1.3 × 10−4 | 2.5 × 10−5 | 1.3 × 10−4 | 2.7 × 10−5 | 1.1 × 10−4 | 2.6 × 10−5 | 1.4 × 10−4 | 7.5 × 10−5 |
AC(18:0) | 5.3 × 10−5 | 7.2 × 10−6 | 3.7 × 10−5 | 8.0 × 10−6 | 3.2 × 10−5 | 2.8 × 10−6 | 2.8 × 10−5 | 7.0 × 10−6 | 4.3 × 10−5 | 1.1 × 10−5 |
Palmitoylcarnitine | 3.5 × 10−4 | 1.3 × 10−4 | 2.4 × 10−4 | 8.7 × 10−5 | 1.3 × 10−4 | 1.9 × 10−5 | 1.5 × 10−4 | 4.7 × 10−5 | 2.4 × 10−4 | 1.2 × 10−4 |
AC(18:1) | 2.6 × 10−4 | 9.7 × 10−5 | 1.6 × 10−4 | 5.5 × 10−5 | 9.8 × 10−5 | 1.9 × 10−5 | 1.2 × 10−4 | 5.4 × 10−5 | 2.2 × 10−4 | 1.2 × 10−4 |
AC(18:2) | 1.2 × 10−4 | 3.5 × 10−5 | 6.3 × 10−5 | 2.1 × 10−5 | 5.0 × 10−5 | 1.1 × 10−5 | 5.9 × 10−5 | 2.7 × 10−5 | 9.9 × 10−5 | 5.5 × 10−5 |
Taurodeoxycholic acid | 6.2 × 10−5 | 2.5 × 10−5 | 2.5 × 10−5 | 6.2 × 10−6 | 2.5 × 10−5 | 9.1 × 10−6 | 3.1 × 10−5 | 1.7 × 10−5 | 3.4 × 10−5 | 1.9 × 10−5 |
20α-Hydroxyprogesterone | 1.3 × 10−5 | 4.8 × 10−5 | 4.9 × 10−5 | 2.2 × 10−5 | 9.6 × 10−5 | 2.9 × 10−5 | 1.1 × 10−4 | 3.1 × 10−5 | 1.2 × 10−4 | 8.6 × 10−5 |
Taurocholic acid | 4.0 × 10−4 | 1.8 × 10−4 | 1.4 × 10−4 | 3.1 × 10−5 | 3.7 × 10−4 | 3.3 × 10−4 | 1.7 × 10−4 | 1.1 × 10−4 | 2.4 × 10−4 | 7.6 × 10−5 |
Figure 2.
Levels of plasma metabolites related to tricarboxylic acid (TCA) cycle, nucleotides sugars, and pentose phosphate pathways. Each bar indicates median ± standard deviation (SD) of metabolite in each group. * p < 0.05, ** p < 0.01 indicate significant differences in metabolite concentrations compared to the control group. All the p-values were determined using Welch’s t-test. (blue, control; red, CNFs; green, SDACNFs; orange, chitin; navy, cellulose nanofiber (CLNF); G3P, d-glyceraldehyde 3-phosphate; 3-PG, 3-phosphoglycerate; 2-PG, 2-phosphoglycerate; ADP, adenosine diphosphate; ATP, adenosine triphosphate; 3-HBA, 3-hydroxybenzoate; PEP, phosphoenolpyruvate; NAD, nicotinamide adenine dinucleotide; NADP, nicotinamide adenine dinucleotide phosphate; NADPH, reduced nicotinamide adenine dinucleotide phosphate; Ala, alanine; Asp, aspartate; AcCoA, acetyl coenzyme A; Arg, arginine; SucCoA, succinyl coenzyme A; Glu, glucose; 2-OG, 2-oleoylglycerol).
Figure 2.
Levels of plasma metabolites related to tricarboxylic acid (TCA) cycle, nucleotides sugars, and pentose phosphate pathways. Each bar indicates median ± standard deviation (SD) of metabolite in each group. * p < 0.05, ** p < 0.01 indicate significant differences in metabolite concentrations compared to the control group. All the p-values were determined using Welch’s t-test. (blue, control; red, CNFs; green, SDACNFs; orange, chitin; navy, cellulose nanofiber (CLNF); G3P, d-glyceraldehyde 3-phosphate; 3-PG, 3-phosphoglycerate; 2-PG, 2-phosphoglycerate; ADP, adenosine diphosphate; ATP, adenosine triphosphate; 3-HBA, 3-hydroxybenzoate; PEP, phosphoenolpyruvate; NAD, nicotinamide adenine dinucleotide; NADP, nicotinamide adenine dinucleotide phosphate; NADPH, reduced nicotinamide adenine dinucleotide phosphate; Ala, alanine; Asp, aspartate; AcCoA, acetyl coenzyme A; Arg, arginine; SucCoA, succinyl coenzyme A; Glu, glucose; 2-OG, 2-oleoylglycerol).
Figure 3.
Plasma acyl-carnitine (AC) levels. Each bar represents median ± standard deviation (SD) of metabolite in each group. * p < 0.05, ** p < 0.01 indicate significant differences in metabolite concentrations compared to the control group. All the p-values were determined using Welch’s t-test. (blue, control; red, chitin nanofibers (CNFs); green, surface-deacetylated (SDA) CNFs; orange, chitin; navy, cellulose nanofiber (CLNF); N.D.: not detected).
Figure 3.
Plasma acyl-carnitine (AC) levels. Each bar represents median ± standard deviation (SD) of metabolite in each group. * p < 0.05, ** p < 0.01 indicate significant differences in metabolite concentrations compared to the control group. All the p-values were determined using Welch’s t-test. (blue, control; red, chitin nanofibers (CNFs); green, surface-deacetylated (SDA) CNFs; orange, chitin; navy, cellulose nanofiber (CLNF); N.D.: not detected).
Figure 4.
Plasma fatty acid (FA) levels. Each bar represents median ± standard deviation (SD) of metabolite in each group. * p < 0.05, ** p < 0.01 indicate significant differences in metabolite concentrations compared to the control group. All the p-values were determined using Welch’s t-test. (blue, the control; red, chitin nanofibers (CNFs); green, surface-deacetylated (SDA) CNFs; orange, chitin; navy, cellulose nanofiber (CLNF); N.D., not detected).
Figure 4.
Plasma fatty acid (FA) levels. Each bar represents median ± standard deviation (SD) of metabolite in each group. * p < 0.05, ** p < 0.01 indicate significant differences in metabolite concentrations compared to the control group. All the p-values were determined using Welch’s t-test. (blue, the control; red, chitin nanofibers (CNFs); green, surface-deacetylated (SDA) CNFs; orange, chitin; navy, cellulose nanofiber (CLNF); N.D., not detected).
2.2. Effects of Oral Administrations of CNFs and SDACNFs on Gut Microbiota and Fecal Organic Acid Levels
The results of the gut microbiota analysis are shown in
Table 3. The SDACNF group showed a more significant increase in the content of
Bacteroidales than the control group did. On the other hand, no change in gut microbiota was observed in the control, CNF, chitin, and CLNF groups. The results of the determination of fecal short-chain FAs (SCFA) content are shown in
Table 4. The level of fecal lactic acid was significantly increased in the CNF group compared with that of the control group. Furthermore, the level of fecal acetic acid in the CNF group was significantly higher than that of the control group was. In the SDACNF group, the level of fecal propionic acid was significantly higher than that of the control group.
Previously, it was demonstrated that gut microbiota are involved in the maturation and regulation of host immunity and gut functions [
45]. The capsular antigen of the human commensal
Bacteroidales fragilis triggers T cell-dependent immune responses that can affect both the development and homeostasis of the host immune system [
46,
47,
48]. Some reports indicate that gut
Bacteroidales is associated with numerous diseases including inflammatory bowel disease [
49,
50], rheumatoid arthritis [
51], and obesity [
52]. The results of this study suggest that the anti-obesity effects of orally administered SDACNFs may be induced by changes in the population of gut
Bacteroidales.
The main products of intestinal bacterial fermentation of dietary fiber are SCFAs such as lactic acid, acetate, and propionate [
53]. SCFAs can be used for de novo synthesis of lipids and glucose, which are the main energy sources of the host [
54,
55]. Previous reports indicate that some microbiota promote 5-HT biosynthesis from colonic enterochromaffin (EC) cells, which supply 5-HT to the mucosa, lumen, and circulating platelets [
35]. Importantly, microbiota-dependent effects on gut 5-HT significantly affect the host physiology by modulating GIT motility and platelet functions. Reigsted
et al. [
56] in particular, reported that SCFAs promoted tryptophan hydroxylase 1 transcription in a human EC cell model [
56]. Furthermore, their results indicate that the actions of the gut microbiota mediated by SCFAs, are an important determinant of enteric 5-HT production and homeostasis. The results of the present study strongly indicate that the increase in plasma 5-HT level might be induced by the activation of gut microbiota functions by oral administration of CNFs.
Table 3.
Effects of oral administrations of chitin nanofibers (CNFs) and surface-deacetylated (SDA) CNFs on gut microbiota.
Table 3.
Effects of oral administrations of chitin nanofibers (CNFs) and surface-deacetylated (SDA) CNFs on gut microbiota.
% Peak Area | Control | CNF | SDACNF | Chitin | CLNF |
---|
Bacteroidales | 36.8 ± 3.5 | 38.1 ± 6.5 | 52.0 ± 6.6 * | 42.3 ± 3.4 | 41.1 ± 5.2 |
Lactobacillus | 28.9 ± 14.2 | 27.6 ± 5.2 | 18.1 ± 12.9 | 27.7 ± 2.8 | 21.4 ± 6.6 |
Clostridiales | 18.5 ± 12.4 | 14.0 ± 3.9 | 15.4 ± 12.6 | 15.1 ± 7.0 | 21.4 ± 6.1 |
Erysipelotrichaceae | 1.5 ± 0.7 | 1.6 ± 0.4 | 1.0 ± 0.3 | 2.1 ± 0.7 | 2.1 ± 0.5 |
Akkermansia | 0.3 ± 0.6 | 0.0 ± 0.0 | 0.0 ± 0.0 | 0.1 ± 0.2 | 0.0 ± 0.0 |
Anaeroplasma | 0.3 ± 0.2 | 0.4 ± 0.7 | 0.1 ± 0.1 | 0.1 ± 0.1 | 0.0 ± 0.0 |
Corinebacteriales | 0.6 ± 0.4 | 0.5 ± 0.1 | 0.5 ± 0.4 | 0.5 ± 0.3 | 0.7 ± 0.2 |
Mucispirillum | 2.5 ± 1.7 | 1.7 ± 0.2 | 1.4 ± 1.2 | 2.4 ± 0.3 | 1.3 ± 0.4 |
Parasutterella | 2.1 ± 1.5 | 4.1 ± 1.3 | 1.9 ± 1.0 | 2.0 ± 0.8 | 3.2 ± 1.2 |
Table 4.
Effects of oral administrations of chitin nanofibers (CNFs) and surface-deacetylated (SDA) CNFs on fecal short chain fatty acid (FA) levels.
Table 4.
Effects of oral administrations of chitin nanofibers (CNFs) and surface-deacetylated (SDA) CNFs on fecal short chain fatty acid (FA) levels.
mg/g | Control | CNF | SDACNF | Chitin | CLNF |
---|
Lactic acid | 2.1 ± 0.8 | 4.2 ± 1.2 * | 1.6 ± 1.3 | 1.6 ± 0.5 | 2.3 ± 0.6 |
Acetic acid | 3.3 ± 0.7 | 4.1 ± 0.7 † | 3.8 ± 0.8 | 3.3 ± 0.4 | 2.7 ± 0.3 |
Propionic acid | 0.3 ± 0.1 | 0.4 ± 0.1 | 0.5 ± 0.2 * | 0.5 ± 0.1 | 0.3 ± 0.1 |
n-Butyric acid | 0.1 ± 0.1 | 0.2 ± 0.1 | 0.2 ± 0.1 | 0.2 ± 0.1 | 0.2 ± 0.1 |