Chondroitin Testing for Sports Nutrition and Joint Health Supplements
Author: Dr. Hong You
Email: hongyou@eurofinsus.com
New Testing Service Now Available at our Eurofins Brea Lab - Chondroitin Testing
Chondroitin is a compound that is abundant in the connective tissues of people and animals. European Medicines Agency indicates chondroitin sulfate as an anti-osteoarthritis prescription drug, but the US FDA regulates chondroitin sulfate as a dietary supplement, giving U.S. consumers the flexibility to obtain and use chondroitin more freely. In the U.S., Chondroitin sulfate is one of the most popular joint health supplements because it is clinically proven effective and safe in treating osteoarthritis. In sports and fitness nutrition, Chondroitin sulfate is often used to prevent joint pain or treat sports injuries.
Chondroitin sulfate is a sulfated glycosaminoglycan composed of a chain of repeating disaccharide carbohydrates. Animal-derived natural chondroitin sulfate is found as a part of a proteoglycan that consists of a protein core and glycosaminoglycan side chains. From the product quality control standpoint, chondroitin sulfate is challenging to analyze. Depending on the sources, chondroitin sulfate glycosaminoglycan polymer can have wide-range molecular weight, making it difficult to be quantified intact by liquid chromatography with mass spectrometry due to the lack of appropriate reference material. Liquid chromatography with a UV-VIS detector cannot test the intact chondroitin sulfate as well because the polymer has virtually no chromophores. Also, the structure of chondroitin sulfate is similar to other nonfunctional glycosaminoglycans such as keratin sulfate and heparin sulfate, which may present as adulterants and/or interferences that lead to false-positive results. Therefore, one of the keys of chondroitin analysis is to use a specific method that targets chondroitin sulfate.
Eurofins Brea lab (formerly known as Analytical Laboratories in Anaheim, Inc.) is a market leader for Chondroitin testing for 20 years and is the original method developer/author of the AOAC official method 2015.11 Determination of Chondroitin Sulfate Content in Raw Materials and Dietary Supplements. There are several techniques that are used to characterize and quantify chondroitin sulfate in dietary supplements. The selection of the testing technique is dependent on sample matrix, formulation, and methods used when establishing product specification
We offer the below validated or scientifically-valid methods to tackle different matrices and scenarios for the chondroitin sulfate analysis.
Enzymatic hydrolysis with liquid-chromatography analysis
Target Matrices: Dietary supplement (complex formulation) and processed raw ingredient
Method description: This technique quantifies chondroitin sulfate sodium in different matrices described above. During sample preparation, Chondroitinase is used to break the glycosaminoglycan into chondroitin disaccharide units. This method then uses reversed-phase liquid-chromatography equipped with PDA detector to determine the total amount of chondroitin sulfate sodium by using corresponding external chondroitin disaccharide calibration standards (ΔDi-0S, ΔDi-4S, and ΔDi-6S). Because Chondroitinase will only hydrolyze chondroitin sulfate, this technique can differentiate chondroitin sulfate from other glycosaminoglycans such as keratin sulfate and heparin sulfate. This technique was adopted as AOAC official method 2015.11 and sometimes recognized as the “AOAC method.” The current Brea method is optimized from the original AOAC method, and it returns equivalent results as the AOAC method.
Enzymatic hydrolysis with liquid-chromatography analysis for shark products
Target Matrices: Dietary supplements and ingredients containing processed shark products
Method description: This method is an extension of the regular enzymatic hydrolysis technique and utilizes a separate column separation and data processing process. In addition to the chondroitin disaccharides (ΔDi-0S, ΔDi-4S, and ΔDi-6S) that are included in the regular enzymatic hydrolysis method, this method reports and calculates several additional chondroitin disaccharides ΔDi-di(2,6)S, ΔDi-di(4,6)S, ΔDi-tri(2,4,6)S that are unique to the marine/shark products. This method does not have an additional protease pretreatment step.
Photometric Titration
Target Matrices: Dietary supplement (simple formulation) and high-purity raw ingredient (USP grade)
Method description: Cetyl pyridinium chloride (CPC) is a positively charged polymer that can form an ion-pair with chondroitin sulfate during sample preparation. This ion-pair is insoluble in water and cause turbidity. The concentration of chondroitin sulfate can be determined by titrating a solution of chondroitin sulfate and measuring the resulting turbidity. This technique is adopted by the United States Pharmacopeia as its monograph method and sometimes recognized as the “USP method.” This technique can be performed relatively easily at a low cost. However, it has several drawbacks because it cannot differentiate chondroitin sulfate from other glycosaminoglycans and other anionic large-molecules, leading to possible false-positive results. This technique also can not break proteoglycan containing chondroitin sulfate, making it not appropriate to be used for analyzing unprocessed crude animal-derived raw material.