Spectrophotometry part 2 determination of food dye content
Advances in Agrophysical Research. Food is a complex system comprised predominantly of water, fat, proteins and carbohydrates together with numerous minor components. The functional properties of these components, which are governed by their molecular structure and intra- and intermolecular interactions within food system, and the amounts present define the characteristics of food products.
Improving Food Dye Analysis in Commercial Products with Spectrophotometry
Quality of food products refers to the minimum standards for substances to qualify as fit for human consumption or permitted to come in contact with food. Appearance, color, flavor and texture are critical aspects for the sensory quality of food. The food quality includes also chemical, biological and microbial factors, e.
Recently, public interest in food quality and production has increased, probably related to changes in eating habits, consumer behavior, and the development and increased industrialization of the food supplying chains.
The demand for high quality and safety in food production obviously calls for high standards for quality and process control, which in turns requires appropriate analytical tools to investigate food. Spectroscopic methods have been historically very successful at evaluating the quality of agricultural products, especially food.
These methods are highly desirable for analysis of food components because they often require minimal or no sample preparation, provide rapid and on-line analysis, and have the potential to run multiple tests on a single sample. The latter technique is routinely used as a quality assurance tool to determine compositional and functional analysis of food ingredients, process intermediates, and finished products [ 1 ].
The aim of this paper is to demonstrate applicability of four spectroscopic techniques, e. UV—VIS spectroscopy, fluorescence, infrared and Raman spectroscopy, as rapid analysis methods to determine the quality of cereals, cereals products and oils. Additionally, physical foundations of the aforementioned methods are described.
The transmission of the light by the sample is shown in figure 1. Absorption of the studied sample depends on the length of the radiation wave, the thickness of the sample and the characteristic extinction coefficient at a given wavelength. The UV-VIS spectroscopy is mainly used to examine the quality of edible oils regarding a number of parameters including the anisidine value.
Anisidine value is a measurement of the level of fats oxidation, and is used for the assessment of poorer quality oils. Precisely, it is the measure of aldehyde production during oxidation of fats. The anisidine value AV is defined as one hundred-fold value of absorbance of a solution of a fat sample containing aldehydes which have reacted with p-anisidine.Two simple spectrophotometric methods have been proposed for simultaneous determination of two colorants Indigotin and Brilliant Blue and two sweeteners Acesulfame-K and Aspartame in synthetic mixtures and chewing gums without any prior separation or purification.
The first method, derivative spectrophotometry ZCDSis based on recording the first derivative curves for Indigotin, Brillant Blue, and Acesulfame-K and third-derivative curve for Aspartame and determining each component using the zero-crossing technique. The other method, ratio derivative spectrophotometry RDSdepends on application ratio spectra of first- and third-derivative spectrophotometry to resolve the interference due to spectral overlapping.
Both colorants and sweeteners showed good linearity, with regression coefficients of 0. The accuracy and precision of the methods have been determined, and the methods have been validated by analyzing synthetic mixtures containing colorants and sweeteners. Two methods were applied for the above combination, and satisfactory results were obtained.
Colorants are added to foods to make them more attractive, replacing their natural color that can be lost during the industrial process or to avoid variations in the color of the final product. The trouble is that some synthetic azo dyes can be toxic to the human health and when in contact with some drugs can cause allergic and asthmatic reactions to some people, induced the development of cancer and others diseases [ 1 ]. In this way, in the last years, efforts have been made to control and to limit the amount of synthetic colorants that are added in foods, whereas the more toxic dyes have been banned.
Thus, it is necessary to have efficient methodologies to control the amount of colorants in foods. The synthetic indigotin dye, indigotin indigo carmine, EINDand the synthetic azo dye, brilliant blue EBB are among the colorants used in common foods such as sweets, drinks, ice cream, and chewing gum. As with many other food additives, the analytical control of these colorants is of considerable importance in the food industry because of their toxic and carcinogenic potential [ 23 ].
Several methods have been proposed for the codetermination of colorants in mixtures. Artificial sweeteners are also widely used in food, beverage, confectionary, and pharmaceutical industries throughout the world. They are the modern non-caloric alternatives to sugars as additives in foods and drinks.
Consumers select low-calorie foods added with artificial sweeteners to decrease or to control calorie intake and thus body mass and to aid control of certain health or medical conditions such as diabetes and hypoglycemia.
In the past few years, micellar electrokinetic chromatography MEKC and capillary zone electrophoresis CZE have been applied to the simultaneous determination of several kinds of sweeteners [ 26 ]. Other methods less commonly used for the Acesulfame-K and Aspartame determination are Fourier transform Raman spectrometry [ 27 ], chemometry [ 28 — 30 ], and the others [ 31 ] Scheme 1.
Chromatography and capillary electrophoresis are very suitable when the sample contains several colorants or sweeteners, and these methods need special equipment that may not be available in certain quality laboratories.
All multivariate calibration methods, including partial least squares PLSrequire the data processing with powerful software as well as the manipulations of the abstract vector space, and its application to the regression analysis and various chemometric methods are often used for more complex mixtures.
But the foods, beverages, and pharmaceuticals contain only one, two, or rarely three colorants and sweeteners. Thus, analytical methods for alternative technique are always useful, especially if the methods are simple, cheap, and comparatively fast. Spectrophotometry, as an alternative methodology, is suitable for routine laboratories especially for developing countries, and sometimes, serious analytical problems can be resolved by this common technique.
One of the classic analytical problems of spectrophotometric multicomponent analysis is that the analyte of interest is often accompanied by other compounds absorbing in the same spectral region. Under computer-controlled instrumentation, derivative spectrophotometry is playing a very important role in the resolution of band overlapping in quantitative analysis [ 32 ].
However, for binary mixtures, the classical zero-crossing method requires often to use a wavelength with low sensitivity in the measurements. Salinas et al.
The main advantage of the ratio derivative spectrophotometry is the chance of performing easy measurements in correspondence of peaks so it permits the use of the wavelength of highest value of analytical signals a maximum or a minimumand moreover, the presence of a lot of maxima and minima is another advantage by the fact that these wavelengths give an opportunity for the determination of active compounds in the presence of other compounds and ingredients which possibly interfere with the assay.
Although a great variety of methods have been applied to the analysis of the aforementioned compounds in foods, there is no report about simultaneous estimation of this combination in synthetic mixtures or in food samples. The aim of this work is to develop easy, sensitive, and fast spectrophotometric methods that can be applied for the routine analysis of the colorants BB and IND and sweeteners ASP and ACE-K simultaneously in the quaternary laboratory mixtures, foods, and drinks without previous separation.
In the present study, measurements were assayed zero-crossing and ratio spectra derivative techniques [ 34 — 36 ]. Two methods were successfully applied for the above combination, and satisfactory results were obtained.
No extraction, no evaporation step, no complexation agent, and no harmful chemicals are involved in the suggested methods, in that connection decreasing time and the error in quantitation and therefore can be used for routine analysis of both colorants in quality control and routine laboratories.
A double-beam Shimadzu UV-VIS spectrophotometer, connected to personal computer compatible with a laser printer, was used. The bundle software, version 2. All chemicals were analytical grade, and double distilled water was used throughout the experiments.In this experiment you were given a concentrated known stock solution of a colored fabric dye, from which you prepared five diluted solutions using a buret and a volumetric flask whose concentrations you knew exactly.
This method of analysis is perhaps the most common method of determining the concentration of colored species in solution and is an important tool for the modern chemical analysis laboratory. Here is some sample data:.
It is the absorbance of a colored solution which is directly related to the concentration of the colored species in the solution. Be very careful how you use your calculator for this calculation. Some calculators require different keystrokes for using the logarithm of a number in a calculation. Absorbance values should come out to be between 0 and 2 why? The absorbances for all the solutions in my sample data are summarized below. Solution 1 Solution 2 Solution 3 Solution 4 Solution 5 0.
For Solution 1, we took 2. The concentration of the original stock solution was ppm parts per millionso Solution 1 has a concentration of. The concentrations of the diluted samples of stock solution are indicated below:. Solution 1 Solution 2 Solution 3 Solution 4 Solution 5 2. Looking back at the absorbances of standard Solutionsthe absorbance of the unknown lies between those of Solution 3 and Solution 4, which means the unknown must have a concentration somewhere between You need to determine the exact concentration of your unknown, however, and to do this you will have to make a calibration graph showing the exact relationship between absorbance and concentration.
The graph should be a straight linesince the absorbance of a colored solution is directly proportional to the concentration of the colored species in the solution.
Be sure to follow the directions for graphing found in the Appendix to the lab manual. Here is a crude approximation of the graph for this experiment. Your graph must be much more precise and more carefully drawn for credit. To find the concentration of the unknown from the graph, look up its absorbance on the vertical axis, draw a line across to the plotted data line, and then drop a line to the concentration axis and read off the concentration.
Experiment Spectrophotometric Determination of Dyes.Muszynskiego 1, Lodz, Poland. Selected synthetic food dyes tartrazine, Ponceau 4R, Brilliant Blue, orange yellow, and azorubine were isolated from liquid preparations mouthwashes and beverages by Solid Phase Extraction on aminopropyl-bonded silica with diluted aqueous sodium hydroxide as an eluent.
Synthetic food dyes are still common food additives despite the growing awareness of their negative influence on the human organism.
The particularly harmful food colorants are azo dyes that exhibit carcinogenic and potentially genotoxic activity [ 1 ]. The list of dyes permitted in the European Union contains over 30 substances of which 12 are synthetic colorants [ 2 ]. Legal requirements and limitations regarding the application of food dyes have led to the development of several analytical techniques that enable the detection and quantification of these food additives.
Liquid chromatography, including thin layer chromatography TLC in normal NP and reversed RP phase mode, has been successfully used to quantify food dyes in different matrices.
Selected references on TLC methods proposed to separate and quantify permitted and illegal synthetic food dyes are listed in Table 1. The actual determination of colorants in different food matrices was often preceded by the sample pretreatment stage including Solid Phase Extraction SPE on sorbents such as cellulose cotton wool [ 3 ], octadecyl-bonded silica RP [ 4 — 7 ], polyamide powder [ 8 ], alumina [ 9 ], polyurethane foam [ 10 ], or amino-modified silica NH 2 [ 5 ].
Quantification of dyes, separated by TLC, was achieved by sorbent scraping, extraction, and spectrophotometric analysis of extracts [ 1112 ], mass spectroscopy [ 13 ], densitometry [ 451214 ], or software processing of images scanned with flatbed scanners [ 15 — 18 ]. The purpose of this study was to develop a new, easy, and rapid methodology that could be used to quantify the most common synthetic dyestuffs in liquid matrices such as beverages or mouthwashes using the cheapest possible chromatographic plates and a simple sample clean-up step.
Their purity was assessed spectrophotometrically according to [ 19 ]. The preparations analyzed in this study were purchased locally mouthwashes or prepared by spiking a commercial, colorless isotonic drink with appropriate dyes at concentrations 0.
The solutions for the standard addition VIS spectrophotometric determinations of E and E dyes in mouthwashes were prepared by aqueous blue mouthwash or ethanolic red mouthwash dilution of 5. The volumes of standard solutions added to the subsequent flasks were 0, 0. The adsorbed dyes were desorbed by washing the sorbent with NaOH 0.
Standard solutions prepared according to Section 2. Samples of mouthwashes prepared according to Section 2. Absorbances for both series of mouthwash solutions were plotted against the concentrations of the added standards and the dye concentrations without the standard addition were obtained by extrapolation of linear plots:. Liquid matrices such as beverages, drops, mouthwashes, and pharmaceutical preparations often have physicochemical properties that make their direct analysis by chromatographic or spectroscopic techniques impossible even after dilution.
Their relatively high viscosity, opacity, and complex composition are the reasons why the analysis of food dyes in such samples is often a two-step process involving some isolation process prior to the actual analysis.
Synthetic food dyes including those analyzed in our study have been isolated from drinks and drops by SPE on the supports such as RP [ 4 — 7 ], cotton wool [ 3 ], polyurethane foam [ 10 ], or aminopropyl-bonded silica [ 5 ].
We have decided to use the commercial SPE columns filled with NH 2 -modified silica, recommended for normal phase extraction of polar compounds and as a weak anion exchanger WAX for organic anions to which the food colorants quantified in our study also belong. The of the NH 2 functional group is around 9.
When this sorbent is supposed to be used as an anion exchanger, the sample must be applied at a pH at least 2 units below 9. The aminopropyl-bonded silica SPE isolation of food dyes reported in [ 5 ] was based on the approach i with anionic food dyes neutralized with ethanolic sulfuric acid. We have, however, decided to try approach ii or iii. According to our earlier research, the recovery of food colorants covered by our study from the aminopropyl-bonded silica is possible with aqueous sodium hydroxide or diluted solutions of organic basis including water soluble amines e.
Alternatively, we have used pH 7.Electromagnetic radiation is characterized by its frequency n or its wavelength l. These two are related by the velocity of light c.
The electromagnetic spectrum ranges from high-energy cosmic rays high frequency, short wavelength to very low-energy microwaves low frequency, long wavelength. Visible light represents a very narrow section of this range with wavelengths between nanometers nm for blue light to around nm for red light. Shorter wavelengths fall into the ultraviolet region and longer wavelengths are in the infrared region. White light is a mixture of all of the wavelengths in the visible range.
When light strikes an object, it may be reflectedabsorbedtransmittedor diffracted. A prism or a diffraction grating separates white light into its various colors. If some of the light is absorbed, the reflected or transmitted light has the complementary color of the absorbed light. A spectrophotometer uses an arrangement of prisms, mirrors, and slits to select light of a desired wavelength and to direct it toward a sample compartment and a detector.
The detector electronically measures the intensity of the light striking it.
Determination of Food Quality by Using Spectroscopic Methods
A sample is placed in the light path, and the instrument compares the intensity of the light going through the sample I to the intensity observed without the sample I o. The effect is measured either as Transmittance Tthe percentage of light that goes through the sample or as the Absorbance Absrepresenting the amount of light absorbed by the sample :. The Absorbance is seen to be proportional to the number of sheets of the colored material.
This is Lambert's Lawthe absorbance is directly proportional to the thickness or path length of the absorbing material. A spectrophotometer is often used to study solutions. A solution containing an absorbing material is compared to a reference solution of the same solvent and non-absorbing materials. However, the addition of the second drop to the first cell has exactly the same effect as adding it to the second cell.
In this case, the path length remains the same but the concentration of colored material is doubled, doubling the absorbance. This is Beer'sLaw : at constant path length, the absorbance is directly proportional to the concentration of absorbing material. The two laws are combined in the Beer-Lambert Law :. The constant a is called the extinction coefficient or the molar absorbtivity coefficient.
If there are several absorbing materials present, the effects are additive:. A graph of Absorbance vs Wavelength for a red dye shows a maximum at nm:. For a blue dye, the maximum occurs near nm:. If the both of these dyes are dissolved at the same concentrations to form a purple solution, the resulting graph shows both maxima. This can be seen clearly by looking at all three spectra at a wavelength of nm. The red dye shows an absorbance of 0.
The absorbtivity coefficients can be calculated for the two dyes at wavelengths where the other will not interfere:. We will calculate the absorbtivity coefficient for the red dye at nm to minimize the inflence of the blue dye.
There are mathematical methods to optimize these calculations in overlapping regions, but that is beyond the scope of this discussion. These values may be used to calculate the concentrations of these red and blue dyes in other mixtures:.A spectrophotometer is a specialized instrument that can be used to measure and quantify the reflectance and transmittance properties of a sample material. Among the many uses of spectrophotometers is in the food industry where many foods are colored using federally specified and approved food dyes.
Because these food dyes have been given rigorous chemical inspection, they have many properties that are well documented and that can be used to easily identify them. One of the ways that dyes can be quickly identified is through the use of spectrophotometry, which is an area of science that deals with how specific materials absorb and reflect light.
Spectrophotometers work by exposing a sample material to a polychromatic light source. The reflected light from the sample is then split to its various components, within the visible spectrum. The result will be what is known as a reflectance or spectral curve. The resulting data can now easily be analyzed to give a quantifiable measure of the sample's color.
This can be especially important in the food industry because foods have specific dyes that they are allowed to have in them, and any deviation can be the cause of a discarded batch of products, could turn consumers off to the product, or could induce fines from governmental agencies, under certain circumstances.
The manufacturer is required to strictly adhere to the product label with regards to the ingredients, including food dyes, that make up the recipe for each food product. Thus, food producers tend to be very careful when analyzing food colors, and there are three primary methods for doing so:.
Before production, food producers tend to collect resources and ingredients from many places before they produce the actual end product.
These ingredients, raw materials, can be analyzed using spectrophotometers to ensure that no nonstandard ingredients reach the production line, and this cuts down on waste and saves time by making sure that only approved ingredients even make it to production.
During production, the food manufacturing process can sometimes involve many steps and it's possible that food colorings could become mixed or diluted at any stage of this process.
Because of this, food producers often scan small test batches of products during the production process. This is done to ensure the quality of the end product and can identify any production-line level problems before they cause major issues down the line. The manufacturer can also look into checking the color of the product as it is being produced.
Color information can also be obtained, in real time, and can report to process control indicating whether the product color is within prescribed tolerances. If the color of the product does drift out of spec, the in-line system can send a signal to process control to affect a change in the color of the product. After production is completed. There are many factors that can cause food dyes to change color. If they are exposed to oxygen or light for too long, or if they are accidentally mixed with other dyes, even small deviations in dye color can cause big changes in the color of end products.
Because of this, food manufactures generally test a number of product samples before the finished product is shipped to retailers or distributors. This is done to ensure the quality of the product and to make sure that if any deviations have occurred, that they are caught early enough and meet quality and color specifications before they reach customers.
Regardless of the color being used, or the food that it's being used in, any federally approved dye will have a known quantifiable values. Because of this, food manufacturers are usually able to quickly find and identify color deviations before they can become a problem for retailers or consumers.
Konica Minolta acquires Instrument Systems, focuses on enlarging its illuminance meter and other measurement-technology markets. Identifying Food Dyes with Spectrophotometers. What Color is a Tennis Ball? Entering a New Era of Color Science. Can Babies Recognize Different Colors? What Determines Your Eye Color?Sitting at the kitchen table and trying to help my nine-year-old son to work through long division is like watching a squirrel trying to run from a pack of wild dogs.
As a responsible parent, I began looking for anything that might be contributing to his lack of focus and never ending energy-supply. I immediately went to my food pantry to identify any hidden culprits. The truth is that food color additives are a common ingredient in most processed foods. Many commercial food manufacturers rely heavily on colorants and dyes to improve the appearance of their products.
Industry leaders know that color can greatly influence both taste perception and consumer acceptability, so color additives take precedence when it comes to product formulation and overall quality. Monitoring the color intensity of food additives is one of the most important factors in the overall appearance of a food product, yet they must be carefully monitored in order to meet safety standards and regulations.
Spectrophotometers are widely used for food dye analysis in commercial products due to the efficient and accurate measurements they provide and the ability to store this data for careful monitoring and repeatability. Food product formulations often include natural or artificial coloring and careful monitoring of these additives is necessary to achieve desired results as well as meet industry standards.
Spectrophotometers provide the ability to accurately measure their color intensity and quantify this information for repeatability. Using the latest technology ensures that food products maintain the same color intensity that the consumer has come to expect.
Whether using synthetic or natural dyes. These artificial additives undergo strict compliance and safety regulations, and they are systematically reviewed for safety. Many manufacturers are making the change from artificial to natural food color additives, but with these changes comes new challenges in meeting consumer color expectations. Although many consumers are demanding natural alternatives, color quality is still the driving force behind commercial food product marketability.
As new color formulations arise, it will require FDA approval regardless of how it is derived.
Identifying Food Dyes with Spectrophotometers
Instrumental analysis is not only important for meeting the FDA regulations on color additives, but it is essential in ensuring that color standards continue to meet consumer expectations. The ability to maintain the color intensity of our food choices without the use of synthetic dyes requires the use of instrumental analysis to meet consumer demands. Color measurement instrumentation already plays an important role in food production and food dye analysis in commercial products.
Many regulatory agencies use spectrophotometry to ensure safety and quality in our food products. Spectrophotometers are highly accurate, affordable and take the guess work out of food production and manufacturing.