Analytical Methods for Dietary Supplements

Dietary supplement manufacturers have spent nearly 13 years in limbo, waiting for final publication of the good manufacturing practices (GMPs) for supplements that were mandated by the Dietary Supplement Health and Education Act of 1994 (DSHEA). The absence of an official set of GMPs for supplements, however, hasn’t stopped many companies from implementing strict quality control procedures. Because the proposed GMPs call for manufacturers to ensure that what is in the bottle matches what is on the label, interest in analytical methods for supplements is high. Several well-publicized disputes between the industry and testing organizations in recent months have underlined the importance of analytical testing.

Two key aspects of analytical testing are accuracy and precision. Accuracy generally refers to how close a measurement is to something’s real value. Precision refers to how likely a test is to obtain the same value when the test is repeated. When it comes to quality control, manufacturers want to use analytical methods that are both accurate and precise because those methods are most likely to lead to safe, effective, and consistent products.

Validated methods provide an extra layer of assurance that an analytical test is everything it needs to be. While analytical methods are procedures for determining the physical or chemical properties of a material or ingredient-for instance, its melting point or the amount of a particular chemical it contains-validated methods are analytical methods that have been shown to be accurate, reproducible, and appropriate for measuring a particular material. “Many people use the terms validated method or validation very freely without really understanding what they mean,” says Mark Roman, PhD, president of Tampa Bay Analytical Research Inc. (Largo, FL). “Strictly speaking, method validation is a systematic and well-defined process of demonstrating or confirming the performance characteristics of an analytical method. The key performance characteristics are accuracy and precision. Simply put, validation is demonstrating that the method is suitable for its intended purpose.”

According to Tatania Emmick, associate scientist at Kemin Health (Des Moines, IA), accuracy and precision are critical. “It is not uncommon for two laboratories to test the same product and yield different results when testing for the same compound,” Emmick says. “This is often due to differences in sample preparation, method selectivity, and scope of the method. The intent of a method validation is to measure all of these parameters. A validation is a series of experiments designed to measure the accuracy and reproducibility of an analytical method as well as evaluate its selectivity and scope.”

The fact that a method is validated for one material does not mean that it is valid for other materials, adds Roman, who also chairs analytical method provider AOAC International’s (Gaithersburg, MD) dietary supplements methods committee. “The responsibility of proving that a method works with a particular material or product lies squarely with the user of the method,” Roman says.

DIFFERENT LEVELS OF VALIDATION

Although people often refer to validated methods as one category of analytical testing, there are actually several different levels of validation. The most basic level, according to Roman, is single-laboratory validation (SLV). “An SLV is a series of experiments that demonstrate the performance characteristics of a method in a particular laboratory, but it gives no information about how reproducible the results of the method will be when it is used in another laboratory,” Roman explains. “The SLV study is also the means by which a laboratory may prove that a method is suitable for the analysis of a particular product or ingredient in its own laboratory.”

In the next level, peer-verified validation, several different laboratories may participate, providing more information about how reproducible the method’s results are. If at least eight labs participate on a validation study, the method is said to be collaboratively studied, says Roman, who adds that all AOAC Official Methods of Analyses must successfully complete a collaborative study.

Testing Conflicts:
Is There a Validated Method to the Madness?

In January, ConsumerLab.com (White Plains, NY) and The Vitamin Shoppe (North Bergen, NJ) found themselves at loggerheads. ConsumerLab.com claimed it found a significant amount of lead in a women’s supplement produced by The Vitamin Shoppe. The Vitamin Shoppe recalled the supplements but also criticized ConsumerLab.com for not revealing its methodology.

According to Vitamin Shoppe CEO Tom Tolworthy, the company constantly tests for material identity and the presence of microbial and heavy metal contamination. Tolworthy says The Vitamin Shoppe’s ability to look into the matter was “hampered by the unwillingness of the company making the allegations to share its complete results and testing methodology.” ConsumerLab.com’s Web site states that its lead analyses are performed using an atomic absorption/graphite furnace method or inductively coupled plasma–mass spectroscopy (ICP-MS). In a January 24 press release, ConsumerLab.com CEO Tod Cooperman, MD, said ConsumerLab.com shared its methods and results and did “nothing to hamper and everything to assist The Vitamin Shoppe.” Cooperman did not return an e-mail seeking comment for the article.

But Tolworthy says The Vitamin Shoppe needed more information to complete its investigation. “We simply asked for the test methodology used and the lab and scientists conducting the tests,” Tolworthy says, adding that the company wanted to see “if there is something in the way the results were obtained or how they were read” that could explain the differences in test results. Tolworthy notes that “using a scientifically valid testing methodology is absolutely necessary to achieve reliable results.”

“We tested the same lot at the time of manufacture and after the press release, with different results,” Tolworthy says. “We have also tested retained samples of the raw materials used and aggregately obtained a different result.” Tolworthy maintains that The Vitamin Shoppe doesn’t have enough information to agree or disagree with ConsumerLab.com’s findings.

“The real objective of using AOAC Official Methods of Analyses is to help ensure that dietary supplement products contain the labeled amount of their ingredients over their shelf life,” Roman says. “These methods could be used by raw material suppliers, finished product manufacturers, third-party contract analytical laboratories, or regulatory laboratories.

“There is a misconception that manufacturers are required to use AOAC methods to test their products, but an AOAC method is considered a reference method,” Roman says. “ It is viewed as ‘official’ by FDA, and results generated using it can stand up in a court of law.” Roman adds, however, that there is no reason why manufacturers can’t use their own in-house methods if they have demonstrated that the methods are equivalent or better than AOAC methods.

Validated methods could also help level the industry playing field, Roman says. “Disputes between laboratories and manufacturers over analytical results would be reduced, and sensational news articles stating that a particular supplement did not meet a label claim based on a report from a testing organization that uses unvalidated, inappropriate methods would hopefully stop. For this to occur, however, there needs to be consensus over what analytical methods are to be used, and these methods need to be validated and shown to be fit for their purpose.”

INDUSTRY INVOLVEMENT

The subject of method validation has received more attention recently because the proposed GMPs call for support of label claims, says Emmick, who also chairs AOAC’s working group on lutein and zeaxanthin. “The rapid growth of the supplement industry coupled with the overwhelming number of nutraceutical ingredients on the market has made it challenging for scientists to develop and validate test methods for all the active compounds,” Emmick says. “To support this daunting effort, AOAC received funding in 2001 from FDA, NIH, and the Office of Dietary Supplements to find and validate 20 methods within five years. Recently, a significant portion of this funding was cut. Now, in order to achieve this target, industry involvement is absolutely essential.”

Although AOAC has a permanent staff, they primarily serve an administrative rather than a technical role. “The actual association is composed of the volunteer technical experts that come from industry, government, and academia,” Roman says. “If any methods are to get through the AOAC process, they will have to be driven by industry, for it is industry that has the resources in terms of laboratories, money, and personnel, and industry that has the most to benefit from having official methods. Without industry participation, the AOAC dietary supplement program will die out.

Unfortunately, it seems to be the same companies that are volunteering their services over and over again, which puts a great burden on them. The more industry involvement there is, the more this burden gets spread out.”There are many opportunities for industry to get involved. AOAC has an organizational affiliate program to secure additional funding. But it also calls on the expertise of supplement and ingredient manufacturers to support validation and collaborative testing of analytical methods. Kemin Health, for instance, has been involved in efforts to identify and validate analytical methods for lutein and zeaxanthin, and has also offered to perform the SLV for the method, Emmick says.

“The most immediate need is for volunteer laboratories to participate in collaborative studies,” notes Roman. “There will hopefully be a number of these studies coming up within the next six months for which volunteer labs are needed. Another need is for study directors for these collaborative studies. The study director designs and oversees the collaborative study, and writes the final collaborative study manuscript. It requires some work, but in the end it is very rewarding, and AOAC has personnel in place to help guide new study directors through the process and make it as painless as possible.” Roman adds that companies can also volunteer to supply raw materials or finished products for use in an SLV or collaborative study.

THE BEST OFFENSE

Disputes between the industry and third-party testing organizations like ConsumerLab.com (White Plains, NY) or Consumer Reports (Yonkers, NY) happen more often than most dietary supplement manufacturers would like. In recent months, reports issued by testing organizations have questioned the quality of multivitamins, children’s supplements, and other products. Validated methods could be a useful tool for manufacturers that want to keep these conflicts to a minimum. As in athletics, the best offense is often a good defense.

“Validated methods would help companies defend themselves from these types of reports,” says Roman. “Because official methods do not exist for a large number of the ingredients on the market, these types of organizations are free to test the products using any analytical method they choose for the evaluation of a product. These organizations are rarely called upon to defend their reasoning for selecting a particular method, or to provide proof that the method was suitable for testing the selected products.”

Companies that use a validated method accepted by the scientific community to test their products would find themselves in a better position than critics that use a different method. “If a standard method existed, such as an AOAC Official Method, and the testing organization chose not to use this method, they would be in a position to explain why the official method was not used,” Roman says. “Unfortunately, no one can force these types of organizations to use an AOAC Official Method of Analysis, and rarely if ever will these testing organizations actually make sure that the methods they use work on the products they test.”

Can validated analytical methods help address or even preempt questions about product quality? Yes, but only if collaborative efforts continue. “The method development and validation activities are conducted by volunteer technical experts at their own cost and on their own time,” Roman says. “Many AOAC expert volunteers are government scientists within agencies such as FDA and USDA, and their ability to sit on volunteer committees and commit laboratory resources to the validation effort has been curtailed over the last few years and may continue to decline. As a result, it will be up to industry to take up the slack if the program is to move forward.”