Increasing studies focus on depolymerization of chondroitin sulfate (CS) to enhance its biological activities. In the present study, low-molecular-weight chondroitin sulfate (LMWCS)‑iron complexes were obtained by photocatalysis-Fenton reaction. After degradation with the optimal condition of 0.25 % (w/v) TiO2, 10 mM FeSO4, and 400 mM H2O2 for 0, 15, and 60 min, the average relative molecular weights of CS were reduced to 4.77, 2.47, and 1.21 kDa, respectively. Electron paramagnetic resonance and free radical capture test identified •OH, •O2-, and h+ in the photocatalysis-Fenton system, among them h+ was the major contributor for CS degradation. The structures of degradation products were analyzed by UV, CD, XRD, SEM-EDS, and NMR, and the results indicated that CS chelated iron with its carboxyl and sulfate groups, leading to changes in conformation and microtopography. Then 10 oligosaccharides were identified in the degradation products using HPLC-MSn and the depolymerization mechanism was proposed. Furthermore, iron release was observed in simulated gastrointestinal digestion of LMWCS‑iron complexes. Notably, the everted gut sac experiment demonstrated that LMWCS‑iron complex possessed 3.75 times higher iron absorption than FeSO4 (p < 0.01) and 12.60 times higher CS absorption than original CS (p < 0.0001). In addition, LMWCS‑iron exhibited stronger in vitro antioxidant activity than CS.

Perineuronal nets (PNNs) are a condensed subtype of extracellular matrix that form a net-like coverings around certain neurons in the brain. PNNs are primarily composed of chondroitin sulfate (CS) proteoglycans from the lectican family that consist of CS-glycosaminoglycan side chains attached to a core protein. CS disaccharides can exist in various isoforms with different sulfation patterns. Literature suggests that CS disaccharide sulfation patterns can influence the function of PNNs as well as their labeling. This study was conducted to characterize such interregional CS disaccharide sulfation pattern differences in adult human (n = 81) and mouse (n = 19) brains. Liquid chromatography tandem mass spectrometry was used to quantify five different CS disaccharide sulfation patterns, which were then compared to immunolabeling of PNNs using Wisteria Floribunda Lectin (WFL) to identify CS-glycosaminoglycans and anti-aggrecan to identify CS proteoglycans. In healthy brains, significant regional and species-specific differences in CS disaccharide sulfation and single versus double-labeling pattern were identified. A secondary analysis to investigate how early-life stress impacts these PNN features discovered that although early-life stress increases WFL+ PNN density, the CS-glycosaminoglycan sulfation code and single versus double PNN-labeling distributions remained unaffected in both species. These results underscore PNN complexity in traditional research, emphasizing the need to consider their heterogeneity in future experiments.

Perineuronal nets (PNNs) are a condensed subtype of extracellular matrix that form a net-like coverings around certain neurons in the brain. PNNs are primarily composed of chondroitin sulfate (CS) proteoglycans from the lectican family that consist of CS-glycosaminoglycan (CS-GAG) side chains attached to a core protein. CS disaccharides can exist in various isoforms with different sulfation patterns. Literature suggests that CS disaccharide sulfation patterns can influence the function of PNNs as well as their labeling. This study was conducted to characterize such interregional CS disaccharide sulfation pattern differences in adult human (N = 81) and mouse (N = 19) brains. Liquid chromatography tandem mass spectrometry was used to quantify five different CS disaccharide sulfation patterns, which were then compared to immunolabeling of PNNs using Wisteria Floribunda Lectin (WFL) to identify CS-GAGs and anti-aggrecan to identify CS proteoglycans. In healthy brains, significant regional and species-specific differences in CS disaccharide sulfation and single versus double-labeling pattern were identified. A secondary analysis to investigate how early-life stress (ELS) impacts these PNN features discovered that although ELS increases WFL+ PNN density, the CS-GAG sulfation code and single versus double PNN-labeling distributions remained unaffected in both species. These results underscore PNN complexity in traditional research, emphasizing the need to consider their heterogeneity in future experiments.

In this work, the interaction of chondroitin sulfate (CS) and dermatan sulfate (DS) with plant lectins was studied by affinity capillary electrophoresis (ACE), surface plasmon resonance (SPR) technology, molecular docking simulation, and circular dichroism spectroscopy. The ACE method was used for the first time to study the interaction of Ricinus Communis Agglutinin I (RCA I), Wisteria Floribunda Lectin (WFA), and Soybean Agglutinin (SBA) with CS and DS, and the results were in good agreement with those of the SPR method. The results of experiments indicate that RCA I has a strong binding affinity with CS, and the sulfated position does not affect the relationship, but the degree of sulfation can affect the combination of RCA I with CS to some extent. However, the binding affinity with DS is very weak. This study lays the foundation for developing more specialized analysis methods for CS and DS based on RCA I.

With the prevalence of glucosamine- and chondroitin-containing dietary supplements for people with osteoarthritis in the marketplace, it is important to have an accurate and reproducible analytical method for the quantitation of these compounds in finished products. NMR spectroscopic method based both on low- (80 MHz) and high- (500-600 MHz) field NMR instrumentation was established, compared and validated for the determination of chondroitin sulfate and glucosamine in dietary supplements. The proposed method was applied for analysis of 20 different dietary supplements. In the majority of cases, quantification results obtained on the low-field NMR spectrometer are similar to those obtained with high-field 500-600 MHz NMR devices. Validation results in terms of accuracy, precision, reproducibility, limit of detection and recovery demonstrated that the developed method is fit for purpose for the marketed products. The NMR method was extended to the analysis of methylsulfonylmethane, adulterant maltodextrin, acetate and inorganic ions. Low-field NMR can be a quicker and cheaper alternative to more expensive high-field NMR measurements for quality control of the investigated dietary supplements. High-field NMR instrumentation can be more favorable for samples with complex composition due to better resolution, simultaneously giving the possibility of analysis of inorganic species such as potassium and chloride.

BACKGROUND: Aging is frequently associated with impairments of the musculoskeletal system and many elderly people experience joint discomfort or pain which might reduce their ability to move and consequently alter their quality of life. A beneficial effect of fish cartilage hydrolysate (FCH) on pain and joint function has recently been shown in an ACLT/pMMx osteoarthritis rat model.
METHODS: We therefore performed an exploratory, non-comparative, multi-centric clinical trial including 33 subjects with moderate knee joint discomfort and loss of functionality to investigate the efficacy of FCH on their algo-functional status. We further determined the potential health benefit of FCH in an original clinical ex vivo study investigating the role of FCH human metabolites on primary human chondrocytes.
RESULTS: FCH significantly improved knee pain and function, as assessed by the Knee injury and Osteoarthritis Outcome Score (KOOS). Moreover, FCH significantly reduced pain at rest and while walking, and patient global assessment (PGA), as assessed by the Visual Analogue Scale (VAS), and improved patients\' quality of life (SF-36). FCH metabolites decreased the synthesis of catabolic factors (MMP-13) and pro-inflammatory mediators (NO, PGE2) and limited the inhibitory effect of IL-1β on the synthesis of cartilage matrix components (GAG and collagen).
CONCLUSIONS: Thus, these data provide insights on the mode of action of FCH in humans and contribute to explain how FCH may relieve pain and improve joint function in subjects with knee discomfort. Although these preliminary data need to be confirmed in a randomized controlled trial, they strongly support the potential health benefit of such an active ingredient.
BACKGROUND: The study was registered on clinicaltrials.gov with the identifier NCT04420091 (09/06/2020).

UNASSIGNED: To validate a quantitative high performance liquid chromatography (HPLC) assay for chondroitin sulfate (CS) and hyaluronic acid (HA) in synovial fluid, and to analyze glycan-patterns in patient samples.
UNASSIGNED: Synovial fluid from osteoarthritis (OA, n ​= ​25) and knee-injury (n ​= ​13) patients, a synovial fluid pool (SF-control) and purified aggrecan, were chondroitinase digested and together with CS- and HA-standards fluorophore labelled prior to quantitative HPLC analysis. N-glycan profiles of synovial fluid and aggrecan were assessed by mass spectrometry.
UNASSIGNED: Unsaturated uronic acid and sulfated-N-acetylgalactosamine (ΔUA-GalNAc4S and ΔUA-GalNAc6S) contributed to 95% of the total CS-signal in the SF-control sample. For HA and the CS variants in SF-control the intra- and inter-experiment coefficient of variation was between 3-12% and 11-19%, respectively; tenfold dilution gave recoveries between 74 and 122%, and biofluid stability test (room temperature storage and freeze-thaw cycles) showed recoveries between 81 and 140%. Synovial fluid concentrations of the CS variants ΔUA-GalNAc6S and ΔUA2S-GalNAc6S were three times higher in the recent injury group compared to the OA group, while HA was four times lower. Sixty-one different N-glycans were detected in the synovial fluid samples, but there were no differences in levels of N-glycan classes between patient groups. The CS-profile (levels of ΔUA-GalNAc4S and ΔUA-GalNAc6S) in synovial fluid resembled that of purified aggrecan from corresponding samples; the contribution to the N-glycan profile in synovial fluid from aggrecan was low.
UNASSIGNED: The HPLC-assay is suitable for analyzing CS variants and HA in synovial fluid samples, and the GAG-pattern differs between OA and recently knee injured subjects.

BACKGROUND: Oral treatment of glucosamine (GA) combined with chondroitin sulfate (CS) was reportedly effective for pain relief and function improvement in osteoarthritis patients with moderate to severe knee pain in clinical trials. While the effectiveness of GA and CS on both clinical and radiological findings has been demonstrated, only a few high-quality trials exist. Therefore, controversy regarding their effectiveness in real-world clinical practice remains.
OBJECTIVE: To investigate the impact of GA + CS on clinical outcomes of patients with knee and hip osteoarthritis in routine clinical practice.
METHODS: A multicenter prospective observational cohort study included 1102 patients of both genders with knee or hip osteoarthritis (Kellgren & Lawrence grades I-III) in 51 clinical centers in the Russian Federation from November 20, 2017, to March 20, 2020, who had started to receive oral capsules of glucosamine hydrochloride 500 mg and CS 400 mg according to the approved patient information leaflet starting from 3 capsules daily for 3 wk, followed by a reduced dosage of 2 capsules daily before study inclusion (minimal recommended treatment duration is 3-6 mo). Changes in subscale scores [Pain, Symptoms, Function, and Quality of Life (QOL)] of the Knee Injury and Osteoarthritis Outcome Score (KOOS)/Hip Disability and Osteoarthritis Outcome Score (HOOS) questionnaires during the observational period (up to 54-64 wk with a total of 4 visits). Patients\' treatment satisfaction, data on the combined oral use of glucosamine hydrochloride and CS, concomitant use of non-steroidal anti-inflammatory drugs (NSAIDs), and adverse events (AEs) were also evaluated.
RESULTS: A total of 1102 patients with knee and hip osteoarthritis were included in the study. The mean patient age was 60.4 years, most patients were women (87.8%), and their average body mass index was 29.49 kg/m2. All subscale scores (Pain, Symptoms, Function, and QOL) of the KOOS and HOOS demonstrated clinically and statistically significant improvements. In patients with knee osteoarthritis, the mean score increases from baseline to the end of Week 64 were 22.87, 20.78, 16.60, and 24.87 on Pain, Symptoms, Physical Function (KOOS-PS), and QOL subscales (P < 0.001 for all), respectively. In patients with hip osteoarthritis, the mean score increases were 22.81, 19.93, 18.77, and 22.71 on Pain, Symptoms, Physical Function (HOOS-PS), and QOL subscales (P < 0.001 for all), respectively. The number of patients using any NSAIDs decreased from 43.1% to 13.5% (P < 0.001) at the end of the observation period. Treatment-related AEs occurred in 2.8% of the patients and mainly included gastrointestinal disorders [25 AEs in 24 (2.2%) patients]. Most patients (78.1%) were satisfied with the treatment.
CONCLUSIONS: Long-term oral GA + CS was associated with decreased pain, reduced concomitant NSAID therapy, improved joint function and QOL in patients with knee and hip osteoarthritis in routine clinical practice.

Lung cancer is one of the most commonly diagnosed cancer types. Studying the molecular changes that occur in lung cancer is important to understand tumor formation and identify new therapeutic targets and early markers of the disease to decrease mortality. Glycosaminoglycan chains play important roles in various signaling events in the tumor microenvironment. Therefore, we have determined the quantity and sulfation characteristics of chondroitin sulfate and heparan sulfate in formalin-fixed paraffin-embedded human lung tissue samples belonging to different lung cancer types as well as tumor adjacent normal areas. Glycosaminoglycan disaccharide analysis was performed using HPLC-MS following on-surface lyase digestion. Significant changes were identified predominantly in the case of chondroitin sulfate; for example, the total amount was higher in tumor tissue compared to the adjacent normal tissue. We also observed differences in the degree of sulfation and relative proportions of individual chondroitin sulfate disaccharides between lung cancer types and adjacent normal tissue. Furthermore, the differences in the 6-O-/4-O-sulfation ratio of chondroitin sulfate were different between the lung cancer types. Our pilot study revealed that further investigation of the role of chondroitin sulfate chains and enzymes involved in their biosynthesis is an important aspect of lung cancer research.

BACKGROUND: Joint discomfort is a widespread and growing problem in active adults. The rising interest in preventative nutrition has increased the demand for supplements reducing joint discomfort. Protocols assessing the effect of a nutritional intervention on health commonly involve a series of face-to-face meetings between participants and study staff that can weigh on resources, participant availabilities, and even increase dropout rates. Digital tools are increasingly added to protocols to facilitate study conduct, but fully digitally run studies are still scarce. With the increasing interest in real-world studies, the development of health apps for mobile devices to monitor study outcomes is of great importance.
OBJECTIVE: The purpose of this real-world study was to develop a specific mobile app, Ingredients for Life, to conduct a 100% digital study testing the effectiveness of a hydrolyzed cartilage matrix (HCM) supplement on joint discomfort in a heterogeneous group of healthy, active consumers.
METHODS: The Ingredients for Life mobile app using a visual analog scale was specifically developed to monitor the variation in joint pain after exercise by the study participants. A total of 201 healthy and physically active women and men (18-72 years old) with joint pain completed the study over a period of 16 weeks. Participants were randomly allocated to the study groups and did not receive any dietary or lifestyle advice. Each participant indicated one area of joint pain and logged the type and duration of their weekly activities. They received blinded study supplements and took a daily regimen of 1 g of HCM (HCM group) or 1 g of maltodextrin (placebo group) for 12 weeks while weekly logging joint pain scores in the app. This was followed by a 4-week washout period during which participants continued reporting their joint pain scores (until the end of week 16).
RESULTS: Joint pain was reduced within 3 weeks of taking a low dosage of HCM (1 g/day), regardless of gender, age group, and activity intensity when compared with the placebo group. After stopping supplementation, joint pain scores gradually increased but still remained significantly lower than those of the placebo group after 4 weeks of washout. The low dropout rate (<6% of participants, mainly in the placebo group) demonstrates that the digital study was well received by the study population.
CONCLUSIONS: The digital tool allowed us to measure a heterogeneous group of active adults in a real-world setting (without any lifestyle intervention), thus promoting inclusivity and diversity. With low dropout rates, it demonstrates that mobile apps can generate qualitative, quantifiable, real-world data showcasing supplement effectiveness. The study confirmed that the oral intake of a low dose (1 g/day) of HCM led to a significant reduction of joint pain from 3 weeks after starting supplementation.