With the exception of the required addition of trans fat in 2006, the Nutrition Facts label has remained largely unchanged for the last 20 years. In 2016, the Food and Drug Administration (FDA) published final rules for the new label based on new scientific information and the link between diet and chronic diseases. These changes aim to help consumers make more informed choices, and manufacturers must switch to the new label by January 1, 2020. Different analytical techniques are used to determine the composition of food products, and to generate the necessary data to create accurate Nutrition Facts labels. New developments in instrumentation and technology challenge established techniques, offering greater specificity and sensitivity.
Changing Requirements and Established Methods
Current Nutrition Facts labels have 13 key requirements including: serving size and servings per package; calories and calories from fat; total, saturated, and trans fat; cholesterol; sodium; carbohydrate; fiber; sugar; protein; vitamins A and C; calcium; and iron. The new requirements will necessitate design changes to the serving size, servings per container, and calories text. The amount of added sugar in grams and %DV (percent of daily value) will be required, and the amount of vitamin D, potassium, calcium, and iron will replace vitamins A and C, and must be presented as %DV in micrograms or milligrams. According to the FDA, the new label is aimed at making it easier for consumers to make informed choices, and is based on updated nutrition science.
There are many analytical laboratories that perform nutrient analysis; however, the FDA does not approve or recommend specific laboratories. Therefore, it is the manufacturer’s responsibility to work with a laboratory that employs appropriate methods including those published by The Association of Analytical Chemists (AOAC), a globally recognized, independent, third party, not-for-profit association and voluntary consensus standards developing organization. Analytical Food Laboratories (AFL) in Grand Prairie, TX is one such laboratory: “We use methodologies that have been proven by other agencies (AOAC, USDA, AOCS),” says Stephen Wuller, PhD, CFS, chemistry manager at AFL.
Quantifying Macro- and Micro-Nutrients
Different vitamins, minerals, macro-, and micro-nutrients are quantified with various analytical methods. The amount of crude protein in a sample is often determined from nitrogen content using the Kjeldahl method, which relies on digestion in concentrated sulfuric acid followed by the conversion of organic nitrogen to ammonium sulfate. Ammonia is distilled into boric acid solution and the borate ions are titrated to calculate nitrogen content. Alternatively, the Dumas combustion method may be used to quantify nitrogen, and subsequently protein in a sample. The method relies on sample combustion at high temperature in an oxygen atmosphere, and oxidation and reduction tubes to convert nitrogen to N2. Nitrogen gas is measured via thermal conductivity detector. “Our combustion analyzer determines the nitrogen gas content in combusted foods and from that value determines the protein content,” says Robert Abad, principal food chemist at Pacific Coast Analytical Services.
Total fat determination can be achieved with gas chromatography coupled to a flame ionization detector (GC-FID). “We analyze fatty acids and cholesterol with GCFID,” explains Abad. Prior to analysis, all lipid compound classes must be extracted, and bonds and interactions with non-fat compounds must be broken. This is often achieved with non-polar organic solvents like chloroform, n-hexane, and polar organic solvents such as methanol. The Soxhlet method is used to extract neutral lipid, such as triacylglycerol from various foods, and involves transferring partially soluble components of a solid to a liquid phase with a Soxhlet extractor. While this is an effective technique, new efficient methods are emerging. “We examine new technologies that either can improve or speed up analysis,” says Wuller. “Adoption of a pressurized heated fat extractor essentially halved the total analysis time vs. traditional Soxhlet methodology.”
In recent years, the demonization of fats by food manufacturers and the media has shifted, and sugar is now considered the primary dietary culprit of obesity and cardiometabolic disease. Nutritional labeling requirements for sugar encompass all monosaccharides (fructose, glucose, and galactose) and disaccharides (lactose, maltose, and sucrose) in the food product. These are referred to as free sugars, which are naturally present in various food products, and may also be added by the manufacturer. Current analytical methods for sugar quantification include LC-RI (liquid chromatography with refractive index detection), and LC-ELSI (liquid chromatography with evaporative light scattering detection).
Under the new regulations, the amount of added sugar, in grams and %DV, will be required. However, differentiating between naturally occurring and added sugars poses challenges for analytical labs. “For added sugars, there’s still no way analytically to differentiate the sugar (e.g. sucrose) coming from additional added or the original product,” explains Wuller. The inability to distinguish naturally occurring or added sugars creates an opportunity for manufacturers to modify products with added sugars to improve flavor or reduce cost. He notes, however, that “column and/or instrument manufacturers provide technical notes/applications that have proved useful for new testing requests.”
The upcoming changes will also include the replacement of vitamins A and C content with vitamin D. The rationale for the change is that vitamin A and C deficiencies are now very rare, while many Americans do not meet the requirements for vitamin D. “We use LC-MS/MS for vitamin D,” says Wuller. Indeed, LC-MS (liquid chromatography coupled to mass spectrometry) is the AOAC validated method for analysis of vitamin D in food. While methods differ slightly depending on the nature of the sample, they all involve four essential steps: digestion to break down the matrix; extraction of vitamin D from the matrix; clean-up; and quantitative detection by HPLC. However, as vitamin D exists in D2 (ergocalciferol) and D3 (cholecalciferol) forms, it poses challenges for chromatographic resolution. New stationary phases, and LC-MS and LC-MS/MS with stable isotope-labeled internal standards aim to improve accuracy.
In addition to calcium and iron, the %DV and milligrams or micrograms of potassium must be included to meet the new label requirements. ICP-MS (inductively coupled plasma mass spectrometry) enables simultaneous detection of these elements, has a wide linear dynamic range, high sensitivity, and facilitates high sample throughput.
With various guidelines and well-established methods, nutritional analyses may initially seem like routine processes. Certainly, established methods are valuable; however, the challenge lies in the complexity of the samples. As Wuller explains, “mixed matrix samples (e.g. beef enchilada or stews with vegetables and meats) or when the method is not amenable to the typical food analysis (e.g. dietary fiber on meat samples)” are examples of circumstances that require special consideration. According to Abad, “A wide range of sample matrices come in and out of the lab on a daily basis ranging from dog snacks to potato chips to edible insects and CBD oils. Since we are a food testing lab, we see all sorts of challenging and laborintensive products with challenging matrices. One of the main goals with any sample is to ensure a homogeneous matrix. Having this allows the best representation of uniformly distributed macro and micronutrient content in any particular sampling.”
A homogeneous matrix is critical for accurate analyses, but how is this achieved? “We have developed many useful techniques working with blenders, grinders, mortar and pestles, and other mechanical means to ensure that the sample is homogeneous and fits the criteria for the particular analyte being tested,” explains Abad. “To do this efficiently and effectively is a difficult task that evolves with the samples that continue to come into the lab.”
The upcoming changes to the Nutrition Facts labels represent a hurdle for food product manufacturers. Fortunately, analytical food labs are equipped with reliable methodology and new technologies for accurate analyses.
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