Glucosamine sulfate (GS) and chondroitin sulfate (CS) are widely marketed as joint-protective supplements, and have been extensively studied for the management of osteoarthritis, with mixed results [1
]. GS is a ubiquitous sulfated monosaccharide found in shellfish exoskeletons and in fungi; it can also be produced from plants by fermentation. GS is a key building block for glycosaminoglycans in the extracellular matrix of cartilage and other connective tissues. CS is a complex polysaccharide (glycosaminoglycan) composed of repeating disaccharide chains (glucuronic acid and N-acetylgalactosamine) with sulfate groups in various locations, depending on the source of CS. CS is a structural component of cartilage, providing resistance to compression. CS supplements are produced from bovine, porcine, and marine cartilage.
Oral GS and CS supplements are thought to have anti-inflammatory and antiapoptotic effects on articular cartilage and bone [5
]. However, only 10–12% of GS and 5–15% of CS are absorbed from the gut [7
]. Absorption of CS from the small intestine is so low that it has been studied as a promising coating agent for drug delivery to the colon [10
]. Once chondroitin reaches the cecum, it must be degraded by the gut bacteria to disaccharides in order to be absorbed [11
]. GS, as a monosaccharide, does not require bacterial processing for absorption; however, gut bacteria consume more than 50% of orally administered GS before it can be absorbed [7
]. Further, the absorbed fraction varies with antibiotic use, suggesting that gut microbiome plays an important role in the bioavailability of GS and CS to the host.
Since GS and CS are used by gut bacteria [12
], their therapeutic effects may be exerted through gut bacterial pathways. For example, GS and CS are substrates for sulfate-reducing bacteria, which are implicated in the synthesis of anti-inflammatory compounds and are currently under active investigation for prevention and treatment of several inflammatory and metabolic diseases [13
]. Glucosamine and chondroitin are also important components of intestinal mucin, acting as a barrier between gut flora and the intestinal wall, and potentially affecting gut permeability and intestinal immune mediation [16
]. Understanding the effects of GS and CS on gut microbiota might provide insight into their mechanisms of action and help explain their varied effectiveness in osteoarthritis studies. Hence, we sought to systematically review current evidence of glucosamine and chondroitin sulfate effects on the gut microbiome composition.
2. Materials and Methods
Search strategy: This review followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. We searched Web of Science, MEDLINE, EMBASE, and Scopus databases for articles in English published in peer-reviewed journals and indexed up until July 2018. We used multiple search terms capturing microbiome, microflora, intestinal microbiota/flora, gut microbiota/flora and glucosamine or chondroitin concepts (Appendix A
). Search terms were reviewed with an experienced librarian.
Population, interventions, comparisons: We included original studies involving adult humans (age 18 years or older) or other adult mammals, and reported effects of chondroitin sulfate or glucosamine sulfate on the gut microbiome in vivo or in vitro. Any comparator was permitted. We excluded studies that evaluated n-acetyl glucosamine, as it is not typically used as a supplement. We also excluded studies of mixed interventions, such as combined prebiotic and probiotic formulations of CS or GS with other starches, bacteria, or bacterial products.
Outcomes: Key outcomes of interest were differences in the total gut microbial diversity, and absolute or relative abundance of individual microbial species after exposure to CS or GS, when compared with baseline value or control. We formatted results to universal taxonomy from phylum level to lowest available taxonomic level using the NCBI Taxonomy browser (https://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi
Study selection and data extraction: Two authors (AS and RD) screened titles and abstracts for inclusion. Articles selected for full-text review were discussed by all co-authors for final inclusion. Data were extracted into pre-specified structured tables. Missing data points were marked “unknown” or “not reported”.
Summary measures: We performed a qualitative synthesis of findings from included studies. Overall gut bacterial diversity and relative abundance of genera were summarized as “increased,” “decreased,” or “unchanged”. For studies that reported changes in relative abundances of gut bacterial genera, but not statistical significance, we included those genera only if their relative abundance changed at least two-fold. We did not request or analyze data not included in the published reports.
Risk of bias and quality of evidence assessment: The SYRCLE risk of bias tool from the Cochrane collaboration was used for animal studies, and the standard Cochrane tool was used for human studies. The SYRCLE tool evaluates risk of bias using the same criteria as the Cochrane tool for human studies, but adds additional criteria specific to animal studies [19
]. Quality of evidence was assessed using CERQual methodology [20
]. The CERQual tool has been developed by the Cochrane collaboration for reviews of qualitative evidence and topics with limited knowledge. Assessments were performed by a single author (AS) with team consensus by all authors.
This systematic review evaluated the evidence for the effects of CS and GS on the gut microbiome. Overall, few high-quality studies were available, and the confidence in the evidence was low for CS and insufficient for GS. However, some concordant results emerged. In several studies, CS supplementation did not alter the overall gut microbial diversity, but affected the abundance of individual bacterial genera. The most consistent finding between heterogeneous animal and human studies was an increase in the abundance of genus Bacteroides following exposure to CS in vivo and in vitro.
is the most abundant bacterial genus in the human gut, and is enriched in people consuming a “Western” diet high in meat and fat [29
]. Members of the Bacteroides
genus are known to digest a wide variety of animal and plant glycans as their primary energy source [31
]. In the absence of dietary glycans, Bacteroides
can digest intestinal mucin [33
], which can lead to inflammation at the intestinal wall, and downstream inflammatory effects on the host [35
]. Hence, supplemental CS might serve as an exogenous substrate for Bacteroides
, and protect intestinal mucin from degradation [37
]. For example, the Liu et al. [21
] study, which demonstrated an increase in Bacteroides
following CS supplementation, described lower blood lipopolysaccharide (LPS) levels in mice that received CS during exhaustive exercise. This illustrates a possible gut barrier protective effect of CS under exhaustive exercise conditions. Liu et al. [21
] further described an increase in fecal short-chain fatty acid production associated with CS supplementation - another possible mechanism of anti-inflammatory and gut-protective effect of CS supplementation in mice. It appears that different species within the Bacteroides
genus may exhibit different responses to CS [38
]. Thus, further studies at species level are needed to fully understand the effects of CS supplementation on gut microflora.
Several members of genus Bacteroides
secrete sulfatases, and are capable of cleaving sulfate groups from chondroitin sulfate, and other sulfated glycans [39
]. This not only increases the bioavailability of complex glycans to the host and other gut bacteria, but also releases sulfate for utilization by sulfate reducing bacteria (SRB). Of note, one SRB species, Desulfovibrio piger
, increased in abundance following CS feeding in two mouse studies. SRB have been of increasing interest in human health, due to their positive effects on weight loss and insulin sensitivity [41
], as well as possible detrimental effects in inflammatory bowel disease due to increased H2
S production [42
]. Among studies evaluated in this review, Pichette et al. [23
] and Rey et al. both showed that CS supplementation increases the abundance of D. piger and colonic H2
S levels in mice. Pichette et al. [23
] went on to show that these changes are associated with increased GLP1 and insulin secretion, as well as improved oral glucose tolerance. Rey et al. demonstrated that increases in D. piger abundance and colonic H2
S following CS supplementation did not compromise the gut barrier. These findings suggest beneficial functional effects of D. piger in a setting of CS supplementation.
Other concordant review findings included an increase in the abundance of Bacteroidales S24-7
family following exposure to CS (two studies), and a decrease in the abundance of Lactobacillus
(two studies). Bacteroidales S24-7
is a recently discovered and less characterized family of gut bacteria [43
]. Members of this family are known to process plant and host glycans, and are also known to increase in murine models of colitis during treatment-induced remission [44
]. The decrease in Lactobacillus
may be related to anti-adhesion properties of CS, as was suggested in one vaginal microbiome study, where CS isomers A and C markedly reduced adhesion of Lactobacillus
to epithelial cells [45
]. While intriguing, these findings require further validation.
Low to very low-quality evidence showed variation in response to CS depending on its source, isoform, and host characteristics. Different sources of CS are associated with different locations of sulfate groups on the CS molecule; this situation presents a challenge for gut bacteria, as different enzymes are needed to digest different isoforms [46
]. Bacteria may adapt and expand their enzyme repertoire, however this process may take time, and may not always make biological sense [47
]. For example, in a setting of diverse high-carbohydrate diet, other carbohydrates, like mannose and xylan, may be used preferentially instead of CS [27
]. Therefore, both host diet and gut microbial composition likely determine the rate of CS degradation and its subsequent therapeutic effects.
It is less clear why CS effects on gut microbiota differed by sex in one mouse study. Previous mouse studies have reported strain-specific sex differences in gut microbial composition among different strains of mice [49
]. Sex differences were also observed in human studies, including lower relative abundance of phylum Bacteroidetes
in women [50
]. Interestingly, intra-articular CS concentrations are also reportedly lower in women than in men [51
]. The possible sex differences in CS metabolism necessitate further investigation, and inclusion of animals of both sexes in future studies.
Evidence for GS effects on gut microbiome was limited to one low-quality human study. Many similar mechanisms related to degradation of CS apply to GS, including mucin protection [52
], sulfate donation, and varied digestion depending on other sources of carbohydrate in the diet and gut microbial composition [53
]. However, differences are likely many, since glucosamine is a monosaccharide, and its digestion is likely metabolically “easier” than that of CS. Further studies are needed to understand gut utilization of GS, its local effects, and absorption.
Our review and the included studies had several limitations. Search was limited to peer-reviewed articles in the English language, and therefore may be subject to publication bias. The quality, methodology, and reporting of results were highly variable among included studies, and calculated pooled estimates of results were not obtained. It should also be noted that all mouse studies of CS supplementation used high-dose controlled feeding protocols, which may not be representative of typically lower relative doses of supplemental CS used by humans. All human studies that reported the effects of CS and GS on the gut microbiome were of low quality, which underscores the lack of direct evidence to address this research question in humans.