Classifications

• • A—HUMAN NECESSITIES
  • A21—BAKING; EDIBLE DOUGHS
  • A21D—TREATMENT OF FLOUR OR DOUGH FOR BAKING, e.g. BY ADDITION OF MATERIALS; BAKING; BAKERY PRODUCTS
  • A21D2/00—Treatment of flour or dough by adding materials thereto before or during baking
• • A—HUMAN NECESSITIES
  • A21—BAKING; EDIBLE DOUGHS
  • A21D—TREATMENT OF FLOUR OR DOUGH FOR BAKING, e.g. BY ADDITION OF MATERIALS; BAKING; BAKERY PRODUCTS
  • A21D8/00—Methods for preparing or baking dough
  • A21D8/02—Methods for preparing dough; Treating dough prior to baking
  • A21D8/04—Methods for preparing dough; Treating dough prior to baking treating dough with microorganisms or enzymes
  • A21D8/042—Methods for preparing dough; Treating dough prior to baking treating dough with microorganisms or enzymes with enzymes
• • A—HUMAN NECESSITIES
  • A21—BAKING; EDIBLE DOUGHS
  • A21D—TREATMENT OF FLOUR OR DOUGH FOR BAKING, e.g. BY ADDITION OF MATERIALS; BAKING; BAKERY PRODUCTS
  • A21D13/00—Finished or partly finished bakery products
  • A21D13/40—Products characterised by the type, form or use
  • A21D13/42—Tortillas
• • A—HUMAN NECESSITIES
  • A21—BAKING; EDIBLE DOUGHS
  • A21D—TREATMENT OF FLOUR OR DOUGH FOR BAKING, e.g. BY ADDITION OF MATERIALS; BAKING; BAKERY PRODUCTS
  • A21D13/00—Finished or partly finished bakery products
  • A21D13/60—Deep-fried products, e.g. doughnuts
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23D—EDIBLE OILS OR FATS, e.g. MARGARINES, SHORTENINGS OR COOKING OILS
  • A23D9/00—Other edible oils or fats, e.g. shortenings or cooking oils
  • A23D9/007—Other edible oils or fats, e.g. shortenings or cooking oils characterised by ingredients other than fatty acid triglycerides
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
  • A23L19/00—Products from fruits or vegetables; Preparation or treatment thereof
  • A23L19/10—Products from fruits or vegetables; Preparation or treatment thereof of tuberous or like starch containing root crops
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
  • A23L19/00—Products from fruits or vegetables; Preparation or treatment thereof
  • A23L19/10—Products from fruits or vegetables; Preparation or treatment thereof of tuberous or like starch containing root crops
  • A23L19/12—Products from fruits or vegetables; Preparation or treatment thereof of tuberous or like starch containing root crops of potatoes
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
  • A23L19/00—Products from fruits or vegetables; Preparation or treatment thereof
  • A23L19/10—Products from fruits or vegetables; Preparation or treatment thereof of tuberous or like starch containing root crops
  • A23L19/12—Products from fruits or vegetables; Preparation or treatment thereof of tuberous or like starch containing root crops of potatoes
  • A23L19/18—Roasted or fried products, e.g. snacks or chips
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
  • A23L5/00—Preparation or treatment of foods or foodstuffs, in general; Food or foodstuffs obtained thereby; Materials therefor
  • A23L5/20—Removal of unwanted matter, e.g. deodorisation or detoxification
  • A23L5/27—Removal of unwanted matter, e.g. deodorisation or detoxification by chemical treatment, by adsorption or by absorption
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
  • A23L7/00—Cereal-derived products; Malt products; Preparation or treatment thereof
  • A23L7/10—Cereal-derived products
  • A23L7/117—Flakes or other shapes of ready-to-eat type; Semi-finished or partly-finished products therefor
  • A23L7/13—Snacks or the like obtained by oil frying of a formed cereal dough
• • C—CHEMISTRY; METALLURGY
  • C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
  • C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
  • C11B5/00—Preserving by using additives, e.g. anti-oxidants
• • C—CHEMISTRY; METALLURGY
  • C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
  • C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
  • C11B5/00—Preserving by using additives, e.g. anti-oxidants
  • C11B5/0042—Preserving by using additives, e.g. anti-oxidants containing nitrogen
  • C11B5/005—Amines or imines
• • C—CHEMISTRY; METALLURGY
  • C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
  • C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
  • C11B5/00—Preserving by using additives, e.g. anti-oxidants
  • C11B5/0085—Substances of natural origin of unknown constitution, f.i. plant extracts

Definitions

• the present invention relates to a method for reducing the amount of acrylamide in thermally processed foods and permits the production of foods having significantly reduced
• the invention more specifically relates to: a) adding a combination of two or more acrylamide-reducing agents when making a fabricated food product and b) the use of various acrylamide-reducing agents during the production of potato flakes or other intermediate products used in making a fabricated food product.
• acrylamide has long been used in its polymer form in industrial applications for water treatment, enhanced oil recovery, papermaking, flocculants, thickeners, ore processing and permanent press fabrics.
• Acrylamide participates as a white crystalline solid, is odorless, and is highly soluble in water (2155 g/L at 30°C).
• Synonyms for acrylamide include 2-propenamide, ethylene carboxamide, acrylic acid amide, vinyl amide, and propenoic acid amide.
• Acrylamide has a molecular mass of 71.08, a melting point of 84.5°C, and a boiling point of 125 0 C at 25 mmHg.
• acrylamide monomer In very recent times, a wide variety of foods have tested positive for the presence of acrylamide monomer. Acrylatnide has especially been found primarily in carbohydrate food products that have been heated or processed at high temperatures. Examples of foods that have tested positive for acrylamide include coffee, cereals, cookies, potato chips, crackers, french-fried potatoes, breads and rolls, and fried breaded meats, hi general, relatively low contents of acrylamide have been found in heated protein-rich foods, while relatively high contents of acrylamide have been found in carbohydrate-rich foods, compared to non- detectable levels in unheated and boiled foods.
• Reported levels of acrylamide found in various similarly processed foods include a range of 330 - 2,300 ( ⁇ g/kg) in potato chips, a range of 300 - 1100 ( ⁇ g/kg) in french fries, a range 120 - 180 ( ⁇ g/kg) in corn chips, and levels ranging from not detectable up to 1400 ( ⁇ g/kg) in various breakfast cereals.
• acrylamide is formed from the presence of amino acids and reducing sugars.
• a reaction between free asparagine, an amino acid commonly found in raw vegetables, and free reducing sugars accounts for the majority of acrylamide found in fried food products.
• Asparagine accounts for approximately 40% of the total free amino acids found in raw potatoes, approximately 18% of the total free amino acids found in high protein rye, and approximately 14% of the total free amino acids found in wheat.
• acrylamide from amino acids other than asparagine is possible, but it has not yet been confirmed to any degree of certainty. For example, some acrylamide formation has been reported from testing glutamine, methionine, cysteine, and aspartic acid as precursors. These findings are difficult to confirm, however, due to potential asparagine impurities in stock amino acids. Nonetheless, asparagine has been identified as the amino acid precursor most responsible for the formation of acrylamide.
• the Maillard reaction requires heat, moisture, reducing sugars, and amino acids.
• the Maillard reaction involves a series of complex reactions with numerous intermediates, but can be generally described as involving three steps.
• the first step of the Maillard reaction involves the combination of a free amino group (from free amino acids and/or proteins) with a reducing sugar (such as glucose) to form Amadori or Heyns rearrangement products.
• the second step involves degradation of the Amadori or Heyns rearrangement products via different alternative routes involving deoxyosones, fission, or Strecker degradation.
• the third step of the Maillard reaction is characterized by the formation of brown nitrogenous polymers and co-polymers.
• Figure 1 illustrates a simplification of suspected pathways for the formation of acrylamide starting with asparagine and glucose.
• fried potato chips from raw potato stock.
• the raw potatoes which contain about 80% or more water by weight, first proceed to a peeling step 21. After the skins are peeled from the raw potatoes, the potatoes are then transported to a slicing step 22.
• the thickness of each potato slice at the slicing step 22 is dependent on the desired the thickness of the final product. An example in the prior art involves slicing the potatoes to about 0.053 inches in thickness. These slices are then transported to a washing step 23, wherein the surface starch
• the washed potato slices are then transported to a cooking step 24.
• This cooking step 24 typically involves frying the slices in a continuous fryer at, for example, 177 0 C for approximately 2.5 minutes.
• the cooking step generally reduces the moisture level of the chip to less than 2% by weight. For example, a typical fried potato chip exits the fryer at approximately 1.4% moisture by weight.
• the cooked potato chips are then transported to a seasoning step 25, where seasonings are applied in a rotation drum.
• seasoning step 25 seasonings are applied in a rotation drum.
• the seasoned chips proceed to a packaging step 26.
• This packaging step 26 usually involves feeding the seasoned chips to one or more weighing devices that then direct chips to one or more vertical form, fill, and seal machines for packaging in a flexible package. Once packaged, the product goes into distribution and is purchased by a consumer.
• Minor adjustments in a number of the potato chip processing steps described above can result in significant changes in the characteristics of the final product.
• an extended residence time of the slices in water at the washing step 23 can result in leaching compounds from the slices that provide the end product with its potato flavor, color and texture.
• Increased residence times or heating temperatures at the cooking step 24 can result in an increase in the Maillard browning levels in the chip, as well as a lower moisture
• ingredients may be necessary to establish mechanisms that provide for the absorption of the added ingredients into the interior portions of the slices without disrupting the cellular structure of the chip or leaching beneficial compounds from the slice.
• snacks can also be made from a dough.
• fabricated snack means a snack food that uses as its starting ingredient something
• fabricated snacks include fabricated potato chips that use a dehydrated potato product as a starting material and corn chips that use masa flour as its starting material.
• the dehydrated potato product can be potato flour, potato flakes, potato granules, or other forms in which dehydrated potatoes exist. When any of these terms are used in this application, it is understood that all of these variations are included.
• a fabricated potato chip does not require the peeling step 21, the slicing step 22, or the washing step 23. Instead, fabricated potato chips start with, for example, potato flakes, which are mixed with water and other minor ingredients to form a dough. This dough is then sheeted and cut before proceeding to a cooking step. The cooking step may involve frying or baking. The chips then proceed to a seasoning step and a packaging step.
• the mixing of the potato dough generally lends itself to the easy addition of other ingredients.
• the addition of such ingredients to a raw food product, such as potato slices requires that a mechanism be found to allow for the penetration of ingredients into the cellular structure of the product.
• mixing step must be done with the consideration that the ingredients may adversely affect the sheeting characteristics of the dough as well as the final chip characteristics.
• the proposed invention involves the reduction of acrylamide in food products.
• a reducing agent is used to magnify the effect of an acrylamide-reducing
• cysteine is used as an acrylamide- reducing agent in conjunction with a reducing agent such as ascorbic acid, stannous chloride, sodium sulfite, or sodium meta-bisulfite.
• a reducing agent such as ascorbic acid, stannous chloride, sodium sulfite, or sodium meta-bisulfite.
• the reducing agent can magnify the effectiveness of an acrylamide-reducing agent having a free thiol, thereby minimizing off-flavors that can be apparent with higher levels of acrylamide reducing agents.
• the present invention provides a means for enhancing the quality and characteristics of the end product. Further, such a method of acrylamide reduction is generally easy to implement.
• Figure 1 illustrates a simplification of suspected pathways for the formation of acrylamide starting with asparagine and glucose.
• Figure 2 illustrates well-known prior art methods for making fried potato chips from raw potato stock.
• Figures 3A and 3B illustrate methods of making a fabricated snack food according to two separate embodiments of the invention.
• Figure 4 graphically illustrates the acrylamide levels found in a series of tests in which cysteine and lysine were added.
• Figure 5 graphically illustrates the acrylamide levels found in a series of tests in which CaCl 2 was combined with phosphoric acid or citric acid.
• Figure 6 graphically illustrates the acrylamide levels found in a series of tests in which CaCl 2 and phosphoric acid were added to potato flakes having various levels of reducing sugars.
• Figure 7 graphically illustrates the acrylamide levels found in a series of tests in which CaCl 2 and phosphoric acid were added to potato flakes.
• Figure 8 graphically illustrates the acrylamide levels found in a series of tests in which CaCl 2 and citric Acid were added to the mix for corn chips.
• Figure 9 graphically illustrates the acrylamide levels found in potato chips fabricated with cysteine, calcium chloride, and either phosphoric acid or citric acid.
• Figure 10 graphically illustrates the acrylamide levels found in potato chips when calcium chloride and phosphoric acid are added at either the flakes making step or the chip fabrication step.
• Figure 11 graphically illustrates the effect of asparaginase and buffering on acrylamide level in potato chips.
• Figure 12 graphically illustrates the acrylamide levels found in potato chips fried in oil
• Figure 13 graphically illustrates the effect of the addition of an oxidizing agent or reducing agent to an acrylamide-reducing agent having a free thiol.
• a protein source or amino acid source is provided by a protein source or amino acid source.
• Many plant-derived food ingredients such as rice, wheat, corn, barley, soy, potato and oats contain asparagine and are primarily carbohydrates having minor amino acid components.
• such food ingredients have a small amino acid pool, which contains other amino acids in addition to asparagine.
• thermally processed is meant food or food ingredients wherein components of the food, such as a mixture of food ingredients, are heated at temperatures of at least 8O 0 C.
• the thermal processing of the food or food ingredients takes place at temperatures between about 100 0 C and 205 0 C.
• the food ingredient may be separately processed at elevated temperature prior to the formation of the final food product.
• An example of a thermally processed food ingredient is potato flakes, which is formed from raw potatoes in a process that exposes the potato to temperatures as high as 170 0 C.
• thermally processed food ingredients include processed oats, par-boiled and dried rice, cooked soy products, corn masa, roasted coffee beans and roasted cacao beans.
• raw food ingredients can be used in the preparation of the final food product wherein the production of the final food product includes a thermal heating step.
• raw material processing wherein the final food product results from a thermal heating step is the manufacture of potato chips from raw potato slices by the step of frying at a temperature of from about 100 0 C to about 205 0 C or
• acrylamide a significant formation of acrylamide has been found to occur when the amino acid asparagine is heated in the presence of a reducing sugar. Heating other amino acids such as lysine and alanine in the presence of a reducing sugar such as glucose does not lead to the formation of acrylamide. But, surprisingly, the addition of other amino acids to the asparagine-sugar mixture can increase or decrease the amount of acrylamide formed.
• a reduction of acrylamide in thermally processed foods can be achieved by inactivating the asparagine.
• inactivating is meant removing asparagine from the food or rendering asparagine non-reactive along the acrylamide formation route by means of conversion or binding to another chemical that interferes with the formation of acrylamide from asparagine.
• Lysine and glycine reduced acrylamide formation but not as much as cysteine. All treatments with lysine and/or glycine but without glutamine and cysteine had less than 220 ppb acrylamide (a 85% reduction).
• cysteine, lysine, and glycine demonstrate the effectiveness of cysteine, lysine, and glycine in reducing acrylamide formation.
• glutamine results demonstrate that not all amino acids are effective at reducing acrylamide formation.
• the combination of cysteine, lysine, or glycine with an amino acid that alone can accelerate the formation of acrylamide (such as glutamine) can likewise reduce the acrylamide formation.
• a solution of asparagine (0.176 %) and glucose (0.4%) was prepared in pH 7.0 sodium phosphate buffer. Two concentrations of amino acid (cysteine (CYS), lysine (LYS), glutamine (GLN), or methionine (MET)) were added. The two concentrations were 0.2 and 1.0 moles of amino acid per mole of glucose. In half of the tests, two ml of the solutions were heated at 12O 0 C for 40 minutes; in the other half, two ml were heated at 150 0 C for 15 minutes. After heating, acrylamide was measured by GC-MS, with the results shown in Table 2. The control was asparagine and glucose solution without an added amino acid.
• Table 2 Effect of Temperature and Concentration of Amino Acids on Acrylamide Level
• a control formed 1332 ppb of acrylamide after 40 minutes at 120 0 C, and 3127 ppb of acrylamide after 15 minutes at 150 0 C.
• Cysteine and lysine reduced acrylamide formation at 12O 0 C and 15O 0 C, with the acrylamide reduction being roughly proportional to the concentration of added cysteine or lysine.
• Test potato flakes were manufactured with 750 ppm (parts per million) of added L- cysteine. The control potato flakes did not contain added L-cysteine. Three grams of potato flakes were weighed into a glass vial. After tightly capping, the vials were heated for 15 minutes or 40 minutes at 12O 0 C. Acrylamide was measured by GC-MS in parts per billion (ppb).
• a sheeting step 31 the dough is run through a sheeter, which flattens the dough, and is then cut into discrete pieces.
• a cooking step 32 the cut pieces are baked until they reach a specified color and water content.
• the resulting chips are then seasoned in a seasoning step 33 and placed in packages in a packaging step 34.
• Table 6 Effect of Lysine and Various Levels of Cysteine on Acrylamide Level
• the dry ingredients were first mixed together, then oil was added to each dry blend and mixed.
• the cysteine or lysine was dissolved in the water prior to adding to the dough.
• the moisture level of the dough prior to sheeting was 40% to 45% by weight.
• the dough was sheeted to produce a thickness of between 0.020 and 0.030 inches, cut into chip-sized pieces, and baked.
• FIG. 4 shows the resulting acrylamide levels in graphical form. In this drawing, the level of acrylamide detected in each sample is shown by a shaded bar 402. Each bar has a label listing the appropriate test immediately below and is calibrated to the scale for
• an acrylamide-reducing agent is an additive that reduces acylamide content.
• cysteine or lysine to the dough significantly lowers the level of acrylamide present in the finished product.
• the cysteine samples show that the level of acrylamide is lowered in roughly a direct proportion to the amount of cysteine added. Consideration must be made, however, for the collateral effects on the characteristics (such as color, taste, and texture) of the final product from the addition of an amino acid to the manufacturing process. Additional tests were also run, using added cysteine, lysine, and combinations of each of the two amino acids with CaCl 2 . These tests used the same procedure as described in the tests above, but used potato flakes having varying levels of reducing sugars and varying amounts of amino acids and CaCl 2 added. In Table 7 below, lot 1 of potato flakes had 0.81% reducing sugars (this portion of the table reproduces the results from the test shown above), lot 2 had 1.0% and lot 3 had 1.8% reducing sugars.
• the desired amino acid cannot be simply mixed with the potato slices, as with the embodiments illustrated above, since this would destroy the integrity of the slices, hi one embodiment, the potato slices are immersed in an aqueous solution containing the desired amino acid additive for a period of time sufficient to allow the amino acid to migrate into the cellular structure of the potato slices. This can be done, for example, during the washing step 23 illustrated in Figure 2.
• Table 8 shows the result of adding one weight percent of cysteine to the wash treatment that was described in step 23 of Figure 2 above. All washes were at room temperature for the time indicated; the control treatments had nothing added to the water. The chips were fried in cottonseed oil at 178 0 C for the indicated time.
• Table 8 Effect of Cysteine in Wash Water of Potato Slices on Acrylamide As shown in this table, immersing potato slices of .053 inch thickness for 15 minutes in an aqueous solution containing a concentration of one weight percent of cysteine is sufficient to reduce the acrylamide level of the final product on the order of 100-200 ppb.
• the invention has also been demonstrated by adding cysteine to the corn dough (or masa) for tortilla chips.
• Dissolved L-cysteine was added to cooked corn during the milling process so that cysteine was uniformly distributed in the masa produced during milling.
• the addition of 600 ppm of L-cysteine reduced acrylamide from 190 ppb in the control product to 75 ppb in the L-cysteine treated product.
• Any number of amino acids can be used with the invention disclosed herein, as long as adjustments are made for the collateral effects of the additional ingredient(s), such as changes to the color, taste, and texture of the food.
• ⁇ - amino acids where the -NH 2 group is attached to the alpha carbon atom
• the preferred embodiment of this invention uses cysteine, lysine, and/or glycine.
• amino acids such as histidine, alanine, methionine, glutamic acid, aspartic acid, proline, phenylalanine, valine, and arginine may also be used.
• amino acids and in particular cysteine, lysine, and glycine, are relatively inexpensive and commonly used as food additives. These preferred amino acids can be used alone or in combination in order to reduce the amount of acrylamide in the final
• the amino acid can be added to a food product prior to heating by way of either adding the commercially available amino acid to the starting material of the food product or adding another food ingredient that contains a high concentration level of the free amino acid.
• casein contains free lysine and gelatin contains free glycine.
• the amino acid may be added as a commercially available amino acid or as a food having a concentration of the free amino acid(s) that is greater than the naturally occurring level of asparagine in the food.
• the amount of amino acid that should be added to the food in order to reduce the acrylamide levels to an acceptable level can be expressed in several ways. In order to be commercially acceptable, the amount of amino acid added should be enough to reduce the final level of acrylamide production by at least twenty percent (20%) as compared to a product that is not so treated. More preferably, the level of acrylamide production should be reduced by an amount in the range of thirty-five to ninety-five percent (35-95%). Even more preferably, the level of acrylamide production should be reduced by an amount in the range of fifty to ninety-five percent (50-95%). In a preferred embodiment using cysteine, it has been determined that the addition of at least 100 ppm can be effective in reducing acrylamide.
• cysteine addition is between 100 ppm and 10,000 ppm, with the most preferred range in the amount of about 1,000 ppm.
• a mole ratio of the added amino acid to the reducing sugar present in the product of at least 0.1 mole of amino acid to one mole of reducing sugars (0.1:1) has been found to be effective in reducing acrylamide formation. More preferably the molar ratio of added amino acid to reducing sugars should be between
• glucose is consumed by lysine and glycine, there will be less glucose to react with asparagine to form acrylamide.
• the amino group of amino acids can react with the double bond of acrylamide, a Michael addition.
• the free thiol of cysteine can also react with the double bond of acrylamide.
• Another embodiment of the invention involves reducing the production of acrylamide by the addition of a divalent or trivalent cation to a formula for a snack food prior to the cooking or thermal processing of that snack food.
• a divalent or trivalent cation to a formula for a snack food prior to the cooking or thermal processing of that snack food.
• Chemists will understand that cations do not exist in isolation, but are found in the presence of an anion having the same valence.
• the salt containing the divalent or trivalent cation it is the cation present in the salt that is believed to provide a reduction in acrylamide formation by reducing the solubility of asparagine in water.
• These cations are also referred to herein as a cation with a valence of at least two.
• cations of a single valence are not effective in use with the present invention. In choosing an appropriate compound containing
• the cation having a valence of at least two in combination with an anion the relevant factors are water solubility, food safety, and least alteration to the characteristics of the particular food.
• Combinations of various salts can be used, even though they are discussed herein only as individuals salts.
• Chemists speak of the valence of an atom as a measure of its ability to combine with other elements. Specifically, a divalent atom has the ability to form two ionic bonds with other atoms, while a trivalent atom can form three ionic bonds with other atoms.
• a cation is a positively charged ion, that is, an atom that has lost one or more electrons, giving it a positive charge.
• a divalent or trivalent cation then, is a positively charged ion that has availability for two or three ionic bonds, respectively.
• Simple model systems can be used to test the effects of divalent or trivalent cations on acrylamide formation. Heating asparagine and glucose in 1:1 mole proportions can generate acrylamide. Quantitative comparisons of acrylamide content with and without an added salt measures the ability of the salt to promote or inhibit acrylamide formation. Two sample preparation and heating methods were used. One method involved mixing the dry
• a 20 mL (milliliter) glass vial containing L-asparagine monohydrate (0.15 g, 1 mmole), glucose (0.2 g, 1 mmole) and water (0.4 mL) was covered with aluminum foil and heated in a gas chromatography (GC) oven programmed to heat from 40° to 22O 0 C at 20°/minute, hold two minutes at 22O 0 C, and cool from 220° to 4O 0 C at 20°/min.
• GC-MS gas chromatography-mass spectroscopy
• the process for making baked fabricated potato chips consists of the steps shown in Figure 3B.
• the dough preparation step 35 combines potato flakes with water, the cation/anion pair (which in this case is calcium chloride) and other minor ingredients, which are thoroughly mixed to form a dough.
• the dough is run through a sheeter, which flattens the dough, and then is cut into individual pieces, hi the cooking step 37, the formed pieces are cooked to a specified color and water content. The resultant chips are then seasoned in seasoning step 38 and packaged in packaging step 39.
• the amount of cation added should be enough to reduce the final level of acrylamide production by at least twenty percent (20%). More preferably, the level of acrylamide production should be reduced by an amount in the range of thirty- five to ninety-five percent (35-95%). Even more preferably, the level of acrylamide production should be reduced by an amount in the range of fifty to ninety-five percent (50-95%).
• the amount of divalent or trivalent cation to be added can be given as a ratio between the moles of cation to the moles of free asparagine present in the food product. The ratio of the moles of divalent or trivalent cation to moles of free asparagine should be at least one to five (1:5). More preferably, the ratio is at least one to three (1 :3), and more preferably still,
• moles of asparagine is between about 1 :2 and 1 : 1.
• the molar ratio of cation to asparagine can be as high
• Table 14 Effect of CaCl 2 Across Varying Levels of Reducing Sugars & Cation Levels As seen in this table, the addition OfCaCl 2 consistently reduces the level of acrylamide in the final product, even when the weight ratio of added CaCl 2 to potato flakes is lower than 1:250.
• any number of salts that form a divalent or trivalent cation can be used with the invention disclosed herein, as long as adjustments are made for the collateral effects of this additional ingredient.
• the effect of lowering the acrylamide level appears to derive from the divalent or trivalent cation, rather than from the anion that is paired with it.
• Limitations to the cation/anion pair, other than valence are related to their acceptability in foods, such as safety, solubility, and their effect on taste, odor, appearance, and texture. For example, the cation's effectiveness can be directly related to its solubility.
• Highly soluble salts such as those salts comprising acetate or chloride anions, are most preferred additives.
• Less soluble salts, such as those salts comprising carbonate or hydroxide anions can be made more soluble by addition of phosphoric or citric acids or by disrupting the cellular structure of the starch based food. Suggested cations include calcium, magnesium, aluminum, iron, copper, and zinc.
• Suitable salts of these cations include calcium chloride, calcium citrate, calcium lactate, calcium malate, calcium gluconate, calcium phosphate, calcium acetate, calcium sodium EDTA, calcium glycerophosphate, calcium hydroxide, calcium lactobionate, calcium oxide, calcium propionate, calcium carbonate, calcium stearoyl lactate, magnesium chloride, magnesium citrate, magnesium lactate, magnesium malate, magnesium gluconate, magnesium phosphate, magnesium hydroxide, magnesium carbonate, magnesium sulfate, aluminum chloride hexahydrate, aluminum chloride, aluminum hydroxide, ammonium alum, potassium alum, sodium alum, aluminum sulfate, ferric chloride, ferrous gluconate, ferric ammonium citrate, ferric pyrophosphate, ferrous fumarate, ferrous lactate, ferrous sulfate, cupric chloride, cupric gluconate, cupric sulfate, zinc gluconate, zinc oxide, and zinc
• the presently preferred embodiment of this invention uses calcium chloride, although it is believed that the requirements may be best met by a combination of salts of one or more of the appropriate cations.
• a number of the salts, such as calcium salts, and in particular calcium chloride are relatively inexpensive and commonly used as food.
• Calcium chloride can be used in combination with calcium citrate, thereby reducing the collateral taste effects of CaCl 2 .
• any number of calcium salts can be used in combination with one or more magnesium salts.
• One skilled in the art will understand that the specific formulation of salts required can be adjusted depending on the food product in question and the desired end-
• the inventors have found that calcium ions more effectively reduce acrylamide content at acidic pH.
• the addition of calcium chloride in the presence of an acid was studied and compared to a sample with just the acid.
• results are presented in three separate tables (16A, 16B, and 16C) with each table showing the results for one of the levels of sugar in the potato flakes. Additionally, the tests are arranged so that the controls, with no calcium chloride or phosphoric acid, are on the left- hand side. Within the table, each level of calcium chloride (CC) is grouped together, with variations in the phosphoric acid (PA) following.
• CC calcium chloride
• PA phosphoric acid
• Table 16C CaCl 2 /Phosphoric Acid Effect on Acrylamide Level - 2.07% Reducing Sugars
• Figure 6 shows a graph corresponding to the three tables
• results are again grouped by the level of reducing sugar available from the potato; within each group there is a general movement downward as first one and then several acrylamide-reducing agents are used to lower the acrylamide level.
• the amount of calcium chloride and phosphoric acid necessary to bring the level of acrylamide to a desired level produced objectionable flavors.
• the following test was designed to reveal if the addition to the potato dough of cysteine - which has been shown to lower the levels of acrylamide in the chips - would allow the levels of calcium chloride and acid to be lowered to acceptable taste levels while keeping the level of acrylamide low.
• the three agents were added to the masa (dough) at a ratio of (i.) 0.106% Ca/Cl 2 , 0.084% citric acid, and 0.005% L.
• cysteine in a first experiment (ii) 0.106% Ca/Cl 2 and 0.084% citric acid, but no cysteine in a second experiment, and 0.053% CaZCl 2 , 0.042% citric acid with 0.005% L. cysteine as a third experiment.
• the masa is about 50% moisture, so the concentrations would approximately double if one translates these ratios to solids only. Additionally, in each test, part of the run was flavored with a nacho cheese seasoning at about 10% of the base chip weight.
• Results of this test are shown in Table 18 below, hi this table, for each category of chip, e.g., plain chip, control, the results of the first-run experiment are given in acrylamide #1; the results of the second experiment are given as acrylamide #2, and the average of the two given as acrylamide average. Only one moisture level was taken, in the first experiment; that value is shown.
• category of chip e.g., plain chip, control
• Figure 8 graphically presents the same data as the table above.
• two bars 802 show the acrylamide results.
• Acrylamide results 802a from the first experiment are shown on the left for each type chip, with the acrylamide results 802b from the second experiment shown on the right. Both acrylamide results are calibrated to the markings on the left of the graph.
• the single moisture level is shown as a point 804 overlying the acrylamide graph and is calibrated to the markings on the right of the graph.
• Figure 9 demonstrates graphically the results of this experiment. Results are shown grouped first by the level of reducing sugars, then by the amount of acrylamide-reducing agents added. As in the previous graphs, bars 902 representing the level of acrylamide are calibrated according to the markings on the left-hand side of the graph, while the points 904 representing the moisture level are calibrated according to the markings to the right-hand side of the graph.
• Potato flakes can be made either with a series of water and steam cooks (conventional) or with a steam cook only (which leaches less from the exposed surfaces of the potato). The cooked potatoes are then mashed and drum dried. Analysis of flakes has revealed very low acrylamide levels in flakes (less than 100 ppb), although the products made from these flakes can attain much higher levels of acrylamide.
• Asparaginase is known to decompose asparagine to aspartic acid and ammonia. Although it is not possible to utilize this enzyme in making potato chips from
• the process of making flakes by cooking and mashing potatoes breaks down the cell walls and provides an opportunity for asparaginase to work.
• the asparaginase is added to the food ingredient in a pure form as food grade asparaginase.
• the inventors designed the following sets of experiments to study the effectiveness of various agents added during the production of the potato flakes in reducing the level of acrylamide in products made with the potato flakes.
• the potatoes comprised 20% solids and 1% reducing sugar.
• the potatoes were cooked for 16 minutes and mashed with added ingredients. All batches received 13.7 gm of an emulsifler and 0.4 gm of citric acid. Four of the six batches had phosphoric acid added at one of two levels (0.2% and 0.4% of potato solids) and three of the four batches received CaCl 2 at one of two levels (0.45% and 0.90% of the weight of potato solids).
• the dough used 4629 gm of potato flakes and potato starch, 56 gm of emulsifler, 162 ml of liquid sucrose and 2300 ml of water.
• both batches received these additives at the given levels as the dough was made.
• the dough was rolled to a thickness of 0.64 mm, cut into pieces, and fried at 350 0 F for 20 seconds. Table 20 below shows the results of the tests for these various batches.
• Table 20 Effect OfCaCl 2 / Phosphoric Acid added to Flakes or Dough on Acrylamide Level As seen in the results above and in the accompanying graph of Figure 10, the acrylamide level was the highest in Test C when only phosphoric acid was added to the flake preparation and was the lowest when calcium chloride and phosphoric acid were used in combination.
• Asparaginase is an enzyme that decomposes asparagine to aspartic acid and ammonia.
• aspartic acid does not form acrylamide
• the inventors reasoned that asparaginase treatment should reduce acrylamide formation when the potato flakes are heated.
• the following test was performed. Two grams of standard potato flakes was mixed with 35 ml of water in a metal drying pan. The pan was covered and heated at 100 0 C for 60 minutes. After cooling, 250 units of asparaginase in 5 ml water were added, an amount of asparaginase that is significantly more than the calculated amount necessary. For control, potato flakes and 5 ml of water without enzyme was mixed. The potato flakes with asparaginase were held at room temperature for 1 hour. After enzyme treatment, the potato flake slurry was dried at 6O 0 C overnight.
• the pans with dried potato flakes were covered and heated at 12O 0 C for 40 minutes.
• Acrylamide was measured by gas chromatograph, mass spectrometry of brominated derivative.
• the control flakes contained 11,036 ppb of acrylamide, while the asparaginase-treated flakes contained 117 ppb of acrylamide, a reduction of more than 98%.
• Potato flakes were pretreated in one of four ways. In each of the four groups, 2 grams of potato flakes were mixed with 35 milliliters of water. In the control pre-treatment group (a), the potato flakes and water were mixed to form a paste. In group (b), the potato flakes were homogenized with 25 ml of water in a Bio Homogenizer M 133/1281-0 at high speed and mixed with an additional 10 ml of deionized water. In group (c), the potato flakes and water were mixed, covered, and heated at 6O 0 C for 60 minutes.
• group (d) the potato flakes and water were mixed, covered, and heated at 100 0 C for 60 minutes.
• pre-treatment group (a), (b), (c), and (d) the flakes were divided, with half of the pre-treatment group being treated with asparaginase while the other half served as controls, with no added asparaginase.
• a solution of asparaginase was prepared by dissolving 1000 units in 40 milliliters of deionized water. The asparaginase was from Erwinia chrysanthemi, Sigma A-2925 EC 3.5.1.1. Five milliliters of asparaginase solution (5ml) was added to each of the test potato flake slurries (a), (b), (c), and (d).
• asparaginase treatment reduced acrylamide formation by more than 98% for all pretreatments.
• Neither homogenizing nor heating the potato flakes before adding the enzyme increased the effectiveness of asparaginase.
• asparagine is accessible to asparaginase without treatments to further damage cell structure.
• the amount of asparaginase used to treat the potato flakes was in large excess. If potato flakes contain 1% asparagine, adding 125 units of asparaginase to 2 grams of potato flakes for 1 hour is approximately a 50-fold excess of enzyme.
• the buffering was done with a solution of sodium hydroxide, made with four grams of sodium hydroxide added to one liter of water to form a tenth molar solution. Two batches of potato flakes were made as controls, one buffered and one un- buffered.
• Asparaginase was added to two additional batches of potato flakes; again one was buffered while the other was not.
• the asparaginase was obtained from Sigma Chemical and was mixed with water in a ratio of 8 to 1 water to enzyme.
• the mash was held for 40 minutes after adding the enzyme, in a covered container to minimize dehydration and held at approximately 36°C.
• the mash was then processed on a drum dryer to produce the flakes.
• the potato flakes were used to make potato dough according to the previously shown protocols, with the results shown in Table 22 below.
• Table 22 Effect of Asparaginase and Buffering on Acrylamide Level in Potato Chips
• the addition of asparaginase without a buffer reduced the production of acrylamide in the finished chips from 768 to 54 ppb, a reduction of 93%.
• the use of a buffer did not appear to have the desired effect on the formation of acrylamide; rather the use of the buffered solution allowed a greater amount of acrylamide to form in both the control and the asparaginase experiments.
• the asparaginase reduced the level of acrylamide from 1199 to 111 , a reduction of 91 %.
• Figure 11 shows the results from Table 22 in a graphical manner.
• bars 1102 represent the level of acrylamide for each experiment, calibrated according to the markings on the left-hand side of the graph, while points 1104 represent the moisture level in the chips a, calibrated according to the markings on the right-hand side of the graph. Tests were also run on the samples to check for free asparagine to determine if the enzyme was active. The results are shown below in Table 23.
• Table 23 Test for Free Asparagine in Enzyme Treated Flakes
• the addition of asparaginase reduced the free asparagine from 1.71 to 0.061, a reduction of 96.5%.
• the addition of asparaginase reduced the free asparagine from 2.55 to 0.027, a reduction of 98.9%.
• sample flakes from each group were evaluated in a model system. In this model system, a small amount of flakes from each sample was mixed with water to form an approximate 50% solution of flakes to water. This solution was heated in a test tube for 40 minutes at 12O 0 C. The sample was then analyzed for acrylamide formation, with the results shown in Table 24.
• the average acrylamide level in the control chips was 1133.5 ppb. Adding 500 parts per million of rosemary to the frying oil reduced the acrylamide to 840, a reduction of 26%, while increasing the rosemary to 750 parts per million reduced the formation of acrylamide further, to 775, a reduction of 31.6%. However, increasing the rosemary to 1000 parts per
• Figure 12 demonstrates the results of the rosemary experiment graphically.
• the bars 1202 demonstrate the level of acrylamide and are calibrated to the divisions on the left-hand side of the graph, while the points 1204 demonstrate the amount of moisture in the chips and are calibrated to the divisions on the right-hand side of the graph.
• the disclosed test results have added to the knowledge of acrylamide-reducing agents that can be used in thermally processed, fabricated foods. Divalent and trivalent cations and amino acids have been shown to be effective in reducing the incidence of acrylamide in thermally processed, fabricated foods. These agents can be used individually, but can also be Used in combination with each other or with acids that increase their effectiveness.
• the combination of agents can be utilized to further drive down the incidence of acrylamide in thermally processed foods from that attainable by single agents or the combinations can be utilized to attain a low level of acrylamide without undue alterations in the taste and texture of the food product.
• Asparaginase has been tested as an effective acrylamide-reducing agent in fabricated foods. It has also been shown that these agents can be effective not only when
• agents can also be added to intermediate products, such as dried potato flakes or other dried potato products, during their manufacture.
• agents added to intermediate products can be as effective as those added to the dough.
• Another embodiment of the invention involves reducing the production of acrylamide by the addition of a reducing agent with a free thiol compound to a snack food dough prior to cooking or thermal processing.
• a free thiol compound is an acrylamide
• dithiothreitol has two thiol groups, acrylamide with dithiothreitol was similar to the compounds with one thiol group.
• the two thiol groups in dithiothreitol may react to from disulfides so dithiothreitol was less effective on an equal molar basis than the other thiol containing compounds.
• a test was then run with free thiol compounds and a reducing agent.
• a solution comprising 135 ppm of a free thiol compound (1.114 millimolar of cysteine), 2500 ppb acrylamide (0.0352 millimolars), and about 305 ppm reducing agent (1.35 millimolar of stannuous chloride dihydrate) was prepared in a 0.5 molar sodium phosphate buffer having a pH of 7.0. After heating at 12O 0 C for 40 minutes, the recovery of added acrylamide was measured to be less than 4%. Hence, the amount of acrylamide reduction with the sample containing a reducing agent was over 96%, an additional 17% over the free thiol alone, or control sample.
• a solution of 135 ppm of a free thiol (1.114 millimolar of cysteine), 2500 ppb of acrylamide (0.0352 millimolars), and a 235 ppm of an oxidizing agent (1.35 millimolars of dehydroascorbic acid) was prepared in a 0.5 molar solution of sodium phosphate buffer having a pH of 7.0. After heating at 120°C for 40 minutes, the recovery of added acrylamide was measured to be about 27%. Hence, the amount of acrylamide reduction with the sample containing the oxidizing agent was about 73%, which is less then the reduction achieved by the cysteine control sample.
• Figure 13 graphically illustrates the theorized effect of the addition of an oxidizing or reducing agent to an acrylamide-reducing agent.
• the reducing agents 1304 increase or magnify the effectiveness of cysteine by keeping cysteine in the reduced, thiol 1306 form.
• An oxidizing agent 1302, such as dehydroascorbic acid likely converts the cysteine thiol 1306 into an inactive cysteine disulfide (cystine) 1308.
• the reducing agent having a standard reduction potential (E°) of between about +0.2 and -2.0 volts is used.
• Casein was added to a vial at the 1% level.
• Sodium sulfite was added at 483 ppm (ug sulfur dioxide per g of potato flake) to the casein vial and one of the cysteine vials.
• the samples were each heated at 120°C for 40 minutes.
• the solutions were then measured for acrylamide concentrations. The results are shown in Table 29 below:
• Table 28 indicates that a 1% Casein addition failed to reduce acrylamide levels in potato flakes without a reducing agent.
• the thiol and reducing agent were less effective in reducing acrylamide levels in the potato flakes samples (Table 28 and 29) than in the non-potato flakes solutions.
• acrylamide was added in the non- potato flake samples but had to be formed in the potato flake samples.
• acrylamide formation was probably more important than decomposition.
• conditions were not optimized for potato flakes. The pH of the potato flakes was not adjusted to pH 7, which would increase the reactivity of cysteine with acrylamide.
• the free thiol compound 1306 is selected from the group consisting of cysteine, N-acetyl-L-cysteine, N-acetyl-cysteamine, glutathione reduced, dithiothreitol, casein, and combinations thereof.
• the reducing agent 1304 is selected from the group consisting of stannous chloride dihydrate, sodium sulfite, sodium meta-bisulfite, ascorbic acid, ascorbic acid derivatives, isoascorbic acid (erythorbic acid), salts of ascorbic acid derivatives, iron, zinc, ferrous ions, and combinations thereof.
• One advantage of the present invention is that the same reduction of acrylamide can he achieved by using less free thiol when the free thiol compound is mixed with a reducing agent. Thus, undesirable off-flavors can be reduced or eliminated.
• the acrylamide reduction can be achieved by using free thiol compound and reducing agent in any dough- based snack food.
• Another benefit of the present invention is the inherent nutritional benefit associated with some reducing agents. Ascorbic acid, for example, is also commonly known as vitamin C.

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