Food additives are substances that are used in foods by being added, mixed or infiltrated into food, or by other methods in the process of food production, for the purpose of processing or preserving food, or for the purpose of performing certain specific roles for technological purposes in its manufacture, processing, preparation, treatment, packaging, transport or storage. Examples of the roles that may be played by food additives include preservation (to extend the shelf‐life of foods by protecting them against microbes), coloring (to add or restore or sharpen color in a food), sweetening (to sharpen the taste and add flavor), antioxidation (to extend the shelf‐life of foods by protecting them against oxidation (i.e. fat rancidity and color changes)), as flour treatment agents (to improve the baking properties), stabilizers (to stabilize food during its production, packaging or storage), flavorings and flavoring enhancers (to impart the desired aroma/odor or taste to food), etc. Therefore food additives ultimately become a component of the food. The term “food improvement agents” encompasses food additives, food enzymes and food flavorings.
According to Article 3 of the EU scope of legislation on food additives as controlled by the EU Regulation 1333/2008, the definition of food additives or food improvement agents is:
Any substance not normally consumed as a food in itself and not normally used as a characteristic ingredient of food, whether or not it has nutritive value, the intentional addition of which to food for a technological purpose in the manufacture, processing, preparation, treatment, packaging, transport or storage of such food results, or may be reasonably expected to result, in it or its by‐products becoming directly or indirectly a component of such foods (Food Additives Legislation, Guidance Notes).
There are certain items that are not part of this regulation and they include normal food/food ingredients, even if they are added to perform a controlled function and therefore by this definition, they are not considered as food additives. Other items that are excluded from this regulation include processing aids, including filtration aids and release agents. Substances will fall under the definition of processing aid, if:
Food additives fall into four main categories and they include:
In order to enforce the regulation and monitor strict adherence to the legislation, the European Union has established a number or coding system for all substances used as food additives (E numbers). The E numbers are codes for substances that are permitted to be used as food additives and these codes have to be found in the labeling information for all foods containing such additives.
For example, E number/codes that range from 100–199 are given to food coloring agents such that E100–E109 are for yellow food colors; E110–E119 (oranges); E120–E129 (reds); E130–E139 (blues and violets); E140–E149 (greens); E150–E159 (browns and blacks); and E160–E199 (gold and others).
Food additives that fall under the group of preservatives have E numbers between 200 and 299, such that E200–E209 (sorbates); E21 E219 (benzoates); E220–E229 (sulfites); E230–E239 (phenols and formates/methanoates); E240–E259 (nitrates); E260–E269 (acetates/ethanoates); E270–E279 (lactates); E280–E289 (propionates/propanoates); and E290–E299 (others).
Antioxidants and acidity regulators have their E numbering between E300 and E399, such that E300–E305 (ascorbates/vitamin C); E306–E309 (tocopherols/vitamin E); E310–E319 (gallates and erythorbates); E320–E329 (lactates); E330–E339 (citrate and tartrates); E340–E349 (phosphates); E350–E359 (malates and adipates); E360–E369 (succinates and fumarates); and E370–E399 (others).
Thickeners, stabilizers and emulsifiers are classified with E numbers ranging between 400 and 499 as follows: E400–E409 (alginates); E410–E419 (natural gums); E420–E429 (other natural agents); E430–E439 (polyoxyethene compounds); E440–E449 (natural emulsifiers); E450–E459 (phosphates); E460–E469 (cellulose compounds); E470–E489 (fatty acids and related compounds); and E490–E499 (others).
Acid (pH) regulators and anti‐caking agents have their E numbers ranging between 500 and 599 in the following pattern: E500–E509 (mineral acids and bases); E510–E519 (chlorides and sulfates); E520–E529 (sulfates and hydroxides); E530–E549 (alkali metal compounds); E550–E559 (silicates); E570–E579 (stearates and gluconates); and E580–E599 (others).
Flavor and flavor enhancer E numbering is between 600 and 699, such that E620–E629 (glutamates and guanylates); E630–E639 (inosinates); and E640–E649 (others).
Antibiotics have their E numbers ranging between E700 and E799, while other additives have their E numbers ranging between E900 and E999, such that E900–E909 (waxes); E910–E919 (synthetic glazes); E920–E929 (improving agents); E930–E949 (packaging gases); E950–E969 (sweeteners); and E990–E999 (foaming agents).
New additives that do not find a proper place within these classifications are classes between E1100 and E1599.
In this classification, some of the additives are permitted with specified maximum limits, while other have been banned and are not supposed to be used (zero maximum limit). For example, within food colors, there are a number that are known to be associated with hypereactivity reactions. For example, tatrazine (E102); alkanet, alkannin (E103), Quinoline Yellow (E104), Sunset Yellow FCF. range Yellow S (E110); Cochineal/Carminic Acid (E120); and Cochineal/Carminic Acid (E122) have been banned as they have been implicated in causing hypereactivity in humans, especially children and those who suffer from asthma. Others include, for example, amaranth (E123), which has been labeled as a dangerous food color additive, while others including Ponceau 4R/Cochineal Red A (E124); Erythrosine BS (E127); Patent Blue V (E131); and Indigo Carmine/Idigotine (E132), are also associated with increasing hypereactivity and triggering of allergy in certain individuals. Green S/Acid Brilliant Green BS (E142) has been reported to be carcinogenic. Black PN/Brilliant Black BN (E151) and Carbon Black/Vegetable Carbon (Charcoal) (E153) fall in the same category (Magnuson et al., 2013).
In the case of preservatives, there are also many that cause hypereactivity and certain disorders; for example, sorbic acid (E200) and its salts, mainly sodium sorbate (E201), potassium sorbate (E202); calcium sorbate (E203); benzoic acid (E210) and its various salts (normally added in dairy products) mainly (E211–E219), i.e. Sodium Benzoate; Potassium Benzoate; Calcium Benzoate; Ethyl 4‐hydroxybenzoate; Ethyl 4‐hydroxybenzoate, Sodium Salt; Propyl 4‐hydroxybenzoate; Propyl 4‐hydroxybenzoate, Sodium Salt; Methyl 4‐hydroxybenzoate and Methyl 4‐hydroxybenzoate, and Sodium Salt, respectively. The sorbic and benzoic acids together with their salts are known to cause hyperactivity types of reactions in certain individuals.
The biphenyls and diphenyls (E230), also used as preservatives together with their derivatives, mainly 2‐hydroxybiphenyl (E231); sodium biphenyl‐2‐yl oxide (E232); 2‐(thiazol‐4‐yl) benzimidazole (E233) and hexamine (E239), are known to cause headaches as well as dermatological disorders. Other preservatives that are normally used in meat and meat products (nitrates and nitrites such as potassium nitrate (E249); sodium nitrite (E250); sodium nitrate (E251) and potassium nitrate (saltpetre) (E252)), as well as acetic acid (E260), and a few of its salt derivatives, mainly potassium acetate (E261) and potassium hydrogen diacetate (E262) and also carbon dioxide (E290), also cause problems in the intestines, skin disorders and headaches.
Antioxidants are the food additives that are in most cases safer, with the exception of Butylated Hydroxyanisole (BHA) (E320) and Butylated Hydroxytoluene (BHT) (E321), which are associated with hypereactivity reactions.
However, citric acid (E330) and its salt derivatives such as sodium citrates (E331); potassium citrates (E332) and calcium citrates (E333); tartaric acid (E334) and its salts, mainly sodium tartrate (E335); potassium tartrate (cream of Tartar) (E336); potassium sodium tartrate (E337) and orthophosphoric acid (E338), cause constipation, intestinal problems and headaches.
The majority of emulsifiers and stabilizers are known to be safe, except for Tragacanth (E413), which is suspected to cause problems in individuals who suffer from allergies and intolerance reactions. Other food additives, such as Monosodium Glutamate (MSG) (E621), are known to cause skin disorders, headaches and intestinal problems.
Generally, some of the food additives have been declared safe, some have been declared dangerous and therefore banned/forbidden from their inclusion into foods, some are carcinogenic and therefore banned and others are suspicious. Labeling of all the ingredients in terms of their presence, grade and quantity is mandatory and monitored by guidelines and regulations.
Different types of food additives have different analytical methods and approaches of analysis, depending on their chemistry, the aim of analysis, and the detection levels required, matrices, etc.
Food colors as food additives are normally added to either make up for color loss due to exposure to light, air, moisture and variations in temperature; to enhance naturally occurring colors; or to add color to foods that would otherwise be colorless or colored differently. However, certain food coloring agents have been reported to trigger adverse reactions with respect to allergies, etc. Therefore food colors are regulated to ensure that they adhere to safety standards.
There are a diverse number of methods that have been devised to determine coloring agents used in foodstuffs, with the majority of them centering on the determination of numerous water‐soluble foodstuffs. The methods include sample preparation methods based on solvent extraction, solid phase extraction, as well as separation and detection using chromatographic‐based techniques such as paper chromatography, TLC and HPLC (Dennis et al., 1977, 1998; Chen et al., 1998; Ren et al., 1990; Wu and Zhang, 1992; Zhou and Li, 1990); and spectrophotometric‐based methods (Berzas Nevado et al., 1998; Capitan et al., 1996, 1988; Garcia Penalver et al., 1999; Lau et al., 1995; Ni and Gong, 1997; Sayar and Ozdemir, 1998; Valencia et al., 2000). Electrochemical‐based methods include voltammetric methods (Berzas Nevado et al., 1997; Ni and Bai, 1997); and electrical migration driven methods such as capillary electrophoresis (Berzas Nevado et al., 1999; Kuo et al., 1998; Thompson and Trenerry, 1995; Wang et al., 1998), etc.
Organic acids used as food additives play several roles, including influencing taste, flavor, stability and the role of preservation because of their effects on bacteria, whereby non‐dissociated or non‐ionized organic acids possess the capability to penetrate the bacteria cell wall and thereby disrupt their normal physiologic functioning (Van Immerseel et al., 2006). However, the food industry and suppliers are obligated to meet the labeling requirements as enforced by the regulations.
There are numerous methods published for the determination of benzoic acid in foodstuffs. The majority of these methods are separation‐based methods and they include gas chromatography (AOAC Official Method 983.16., 2000; Choong et al., 1995; De Luca et al., 1995; González et al., 1998, 1999; Lin et al., 1999; Ochiai et al., 1996); high pressure liquid chromatography (HPLC) (Andrade et al., 1999; AOAC Official Method 994.11, 2000; Castellari et al., 1997; Chen et al., 2001; Hannisdal, 1992; Heinanen and Barbas, 2001; Kantasubrata and Imamkhasani, 1991; Mihyar et al., 1999; Pylypiw and Grether, 2000; Silva et al., 2000; Willetts et al., 1996); Validation of Enforcement Methods Service (VEMS Method 0290); micellar electrokinetic chromatography (MEKC) (Boyce, 1999); the use of lanthanide‐sensitized luminescence (Aguilar‐Caballos et al., 1999); spectrophotometric (Hamano et al., 1997); high performance thin layer chromatography (HPTLC) (Khan et al., 1994); and potentiometric (Pezza et al., 2001).
The sulfites used in foods and beverages as sulfiting agents encompass a group of compounds that includes sulfur dioxide and sulfite salts such as sodium sulfite, sodium bisulfite and sodium metabisulfite, where they are used as preservative agents to prevent the action of microbes that cause food spoilage as well as browning reactions in food and beverage products (Wedzicha, 1984). In addition to the added sulfites, many other adduct species are generated because the sulfiting agents tend to undergo many different types of reactions in food/beverage matrices, which generate a variety of derivative species such as sulfite, bisulfite, metabisulfite, etc. (Adachi et al., 1979; Sullivan et al., 1990).
Despite their roles as preservatives in foods, sulfites have been implicated as the cause of allergic reactions as well as some asthmatic responses in certain individuals (Vally et al., 2000) and therefore regulations and legislation require that there must be proper labeling of any food or beverage that contains any sulfite above 10 mg/L or mg/kg (US Food and Drug Administration, 1986).
In order to enforce the regulatory compliance, a reliable analytical method with high selectivity and sensitivity needs to be devised in order to monitor low concentrations of sulfites in complex food/beverage matrices.
Methods for the determination of sulfites/sulfur dioxide in foodstuffs include both direct methods such as titrimetric, electrochemical techniques, mainly polarographic‐based techniques, electrometric and colorimetric‐based methods as well as indirect methods such as those which utilize distillation principles coupled to absorption of the sulfur dioxide in an oxidizing agent, mainly iodine or hydrogen peroxide and the quantitation is normally performed using techniques including volumetric, gravimetric, colorimetric or electrochemical techniques.
Of the many methods that have been devised for the analysis of sulfites in various food/beverage products, an optimized Monier‐Williams Method (AOAC Method 990.28) has been used more frequently despite the fact that it is time‐consuming, labor‐intensive, and has been implicated to produce false positive responses under certain circumstances (AOAC Official Method 990.28, 2000; Kim, 1989). Another method, such as the AOAC Method 990.31, which involves the alkaline extraction of sulfites in foods and beverages followed by separation using ion‐exclusion chromatography with direct current amperometric detection, has been reported to generate more accurate results (AOAC Official Method 990.31, 2000; Kim, 1990; Kim and Kim, 1986).
Other methods for the analysis of sulfites in foods and beverages are enzymatic‐based methods where sulfite in liquid foods or extracts of solid foods is determined utilizing enzymatic principles which involve the reduction of NADH to nicotinamide adenine dinucleotide, and the decrease in NADH is related to the concentration of sulfite using spectrophotometric techniques (NMKL, 1993). Another approach in the measurements of sulfites in food involves the application of differential pulse polarography whereby sulfur dioxide is measured from acidified samples, and then determined by differential pulse polarography. Ion exclusion chromatography has been used to measure sulfites in food by measuring sulfur dioxide released from food matrices by direct alkali extraction. Sulfites in foods have also been analyzed using capillary electrophoresis, where anionic forms of the sulfiting adducts are analyzed (Hirata et al., 2000; Trenerry, 1996).
Nitrates and nitrites are normally added to foods such as cured sandwich meats, bacon, salami or sausages, in order to give them color and also extend their shelf life. However, nitrates and nitrites tend to form nitrosamines in the body, which has the potential to increase the risk of developing gastrointestinal cancer and, in infants, methemoglobinemia (Hord et al., 2009). The acceptable daily intake (ADI) for nitrites, expressed as sodium nitrite, is 0.1 mg/kg body weight. For this reason, there are regulations that govern the use of these additives in foods and therefore reliable methods have been devised and some are continually being added for the analysis of these species in food matrices
Methods of analysis for nitrates and nitrites in foods include spectroscopic techniques after enzymatic reduction (Haman et al., 1998; Korkmaz et al., 1993); ion‐exchange chromatography (Bosch et al., 1995; Merino et al., 2000; Radisavljevic et al., 1996); differential pulse voltammetry (Mesaros et al., 1998); and capillary electrophoresis (Jimidar et al., 1995) among others.
Gallates are artificial food additives that possess antioxidant properties and therefore help stop oxygen molecules from mixing with the oil in food, which would cause the food to go rancid. However, gallates such as propyl gallate, etc. have been implicated to have carcinogenic effects, stomach and skin irritability, as well as allergic reactions that impact breathing. They have also been linked to kidney and liver disorders. For these reasons, gallates as food additives are regulated and labeling requirements have been enforced.
There are a number of methods that have been reported for the determination of gallates (propyl, octyl and dodecyl) in foodstuffs and they include HPLC (Aparicio et al., 2000; Noguera‐Orti et al., 2000; Yamada et al., 1993); micellar electrokinetic chromatography (Boyce, 1999; Hall et al., 1994); spectrophotometric‐based methods (Aguilar‐Caballos et al., 1997, 2000); voltammetric electrochemical methods (Ni et al., 2000); and colorimetric methods (AOAC Official Method 952.09, 2000).
Polysorbates are non‐ionic emulsifying agents that are normally produced by copolymerizing sorbitan anhydride and ethylene oxide such that a fatty acid is esterified to one terminal hydroxyl group of the polyoxyethylene‐oxide side chain. The type of the attached fatty acid used determines the numbers in the names of polysorbates. For example, if monolaurate is used then we have polyoxyethylene sorbitan monolaurate (polysorbate 20); polyoxyethylene sorbitan monooleate (polysorbate 80); polyoxyethylene sorbitan monopalmitate (polysorbate 40); polyoxyethylene sorbitan monostearate (polysorbate 60); and polyoxyethylene sorbitan tristearate (polysorbate 65).
Polysorbate 80 and other food additives that belong to emulsifiers (e.g. emulsifiers carboxymethylcellulose, often referred to as cellulose gum, and polysorbate 80 also known as Tween 80), which are used in processed foods, have been reported to promote inflammatory bowel disease and a cluster of obesity‐related diseases known as metabolic syndrome.
Methods of analysis for the determination of polysorbates in foodstuffs include those which are based on gravimetry followed by colorimetry and gas chromatography (Daniels et al., 1982; Kato et al., 1989; Smullin et al., 1971; Tonogai et al., 1987) and high performance liquid chromatographic methods (Takeda et al., 2001).
Sweeteners, such as sucrose acetate isobutyrate, are sometimes used in certain drinks to a certain maximum permitted as stipulated in the appropriate guidelines. Therefore labeling requirements have been enforced to ensure that the values for recommended acceptable daily intakes, toxicological data and limits are listed. Gas chromatographic based methods are normally among those used to analyze certain sweeteners in foods.
There are regulations in place for various additives. However, new ones are being introduced in the market. Adulteration due to food additives and food improving agents has been gaining popularity among food vendors, manufacturers, food processing industries and business entities. This stems from the whole idea of making economic gains more than for other reasons. Food forensic laboratories ought to have method development schemes in place to continually detect adulteration due to disallowed additives, or new additives which are yet to receive certification.