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Flour Analysis

The production of uniform bakery products require control over the raw materials used in their formation. Flour is a biological material and when obtained from different sources can vary considerably in its protein quality, protein quantity, ash, moisture, enzymatic activity, color, and physical properties. It is essential for the baker to be aware of any variations in these characteristics from one flour shipment to the next. The purpose of flour testing is to measure specific properties or characteristics of a flour.

Ideally the results of these tests can be related to the flour’s performance in the bakery.

The American Association of Cereal Chemists (AACC) publishes approved methods for determining various properties of flour and bakery products.


The simple air-oven method is sufficiently accurate for the routine analysis of flour moisture at the flour mill or bakery. The procedure involves heating a small sample of flour (~2g) for 1 hr at 266˚F (130˚C + 1˚C) and taking the loss in weight as the moisture content.

The moisture content of the flour is important for two reasons. First, the higher the moisture content, the lower the amount of dry solids in the flour. Flour specifications usually limit the flour moisture to 14% or less. It is in the miller’s interest to hold the moisture as close to 14% as possible. Secondly, flour with greater than 14% moisture is not stable at room temperature. Organisms naturally present in the flour will start to grow at high moistures, producing off odors and flavors.


Ash is the mineral material in flour. The ash content of any given flour is affected primarily by the ash content of the wheat from which it was milled and its milling extraction. The test for determining the ash content involves incinerating a known weight of flour under controlled conditions, weighing the residue, and calculating the percentage of ash based upon the original sample weight.

The ash content of wheat varies from about 1.50 to about 2.00%. The pure endosperm contains about 0.35% ash. Considering that the wheat kernel contains about 80% endosperm, it becomes clear that the non-endosperm parts of the kernel (pericarp, aleurone, and germ) are very high in ash when compared to the endosperm. Thus, the ash content is a sensitive measure of the amount of non-endosperm material that is in the flour.

The goal of milling is to separate the endosperm from the non-endosperm parts of the wheat kernel. This separating is difficult and never clean. Thus, there is always contamination of endosperm with non-endosperm and visa versa. As flour yield is increased, the amount of contamination with non-endosperm increases and the ash content increases. Thus, the ash content is a good and sensitive measure of the contamination of the endosperm.

Millers will often comment that the ash does not affect the baking performance of flour. This is probably true. However, the non-endosperm parts of the wheat kernel are known to decrease baking quality and as the ash content increases so does the level of non-endosperm material.

The ash content of white pan bread flour has increased over the years from 0.45% in the 1950s to the current level of 0.50-0.55%. This has undoubtedly resulted from negotiations where the miller has agreed to the flour buyer’s price but only if he can raise the ash content of the flour a couple of points (0.02%).


The amount of protein in a food material is usually determined by measuring the nitrogen content of the material and multiplying that value by a factor. The nitrogen content of a given protein varies depending on its source. For milk products a factor of 6.38 is used, for most cereal grains the factor is 6.25, and in wheat products the factor is 5.70. These factors depend on the percentage of nitrogen in the respective proteins.

The flour protein content is an important parameter for bread flour. Flours containing higher protein contents are more expensive than flours of lower protein content. Likewise, flours with very low proteins for cakes are also more expensive. There is usually, but not always, a good correlation between protein content and bakery performance of a flour.

The classic procedure to determine the nitrogen was the Kjeldahl procedure. This involved digesting the sample in concentrated sulfuric acid, then neutralizing the acid with concentrated sodium hydroxide, followed by distillation of the ammonia (derived from the nitrogen in the protein) into a standard acid. The procedure worked well, however it was an environmental nightmare. In addition to the strong acid and base, the catalysts used to speed the digestion included such materials as mercury and selenium. It should surprise no one that the procedure is seldom used today.

The Kjeldahl procedure has been replaced by the Dumas combustion procedure. In the original Dumas procedure the sample is mixed with cupric oxide and heated in a stream of carbon dioxide in a combustion tube packed with cupric oxide and copper metal. The organic material is converted to carbon dioxide, water and nitrogen. The gas stream is led into 50% potassium hydroxide. This absorbs the carbon dioxide and any oxides of sulfur, leaving only nitrogen as a gas. The volume of nitrogen is then determined. Various machines have been developed to carry out the analysis automatically. The percent nitrogen is then converted to protein using the appropriate factor. Both the Dumas combustion and the Kjeldahl procedures estimate the quantity (total amount) of protein and not the protein quality. As discussed elsewhere, the quantity of protein is extremely important in the baking performance of a flour.

Near Infrared Reflectance (NIR)

The rapid instrumental analysis of cereals and flours has considerable commercial appeal. Therefore, the near infrared reflectance (NIR) method of estimating protein and moisture contents has found ready acceptance in the milling and baking industries because it is capable of generating nearly instantaneous results. NIR instruments can be operated by non-technical personnel with good precision and reproducibility. The method’s accuracy is dependent upon its calibration.

Near infrared (NIR) methodology has been developed for the determination of protein, moisture, and starch of cereals and their milled products. The range of the electromagnetic spectrum extends from the very long radio waves to the very short gamma rays. The near infrared region is between 0.75 and 2.5 microns (μm).

The first commercial NIR instruments appeared in the 1970s and have been improved during the ensuing years by interfacing them with computers. This has led to the rapid evaluation of the spectral data whose numerical results are then shown on the readout screen.

In the NIR range, the absorption bands are broad and overlapping. Thus, measurements taken at any wavelength are affected by several components of the wheat or flour. Therefore, it is necessary to consider several bands of the spectrum to eliminate the interfering effects of other components. This approach necessitates measurements at several wavelengths and computations using multiple regression analysis, which requires computer facilities. The regression equations have to be developed for various cereal types and varieties to calibrate the instrument. They must be rechecked periodically with standard samples. While the equipment is expensive, it is also very efficient and worth the investment for laboratories that need rapid and accurate analyses.

The procedure for carrying out an analysis is quite simple. Essentially, it involves carefully filling a sample cup with the finely ground test material, e.g., flour or meal, and placing the cup in the drawer of the instrument. When the drawer is closed, the instrument automatically starts to analyze the sample by exposing its surface to radiation within a selected narrow band of wavelengths and measuring the reflectance. This reflectance is amplified and converted by the instrument’s microcomputer into numerical results that are displayed on a readout screen. Some newer instruments are transmitted radiation rather than reflected. The entire operation takes approximately one minute. Some of the newer instruments are designed to analyze whole grain samples.

Free Fatty Acids

The level of free fatty acids in flour milled from sound wheat is very low. However, if either the wheat or the flour is subjected to poor storage conditions (high moisture and/or high temperature), enzymes will degrade the native grain lipids and produce free fatty acids. Thus, the level of free fatty acids is a good measure of the storage conditions of either the grain or the flour. Flours with high levels of free fatty acids will be more subjected to rancidity than will sound flours. This is of little importance in bread but quite important in dry products (cookies, crackers, croutons, pretzels, etc.).

The procedure for determining free fatty acids is quite simple. The lipids are extracted with a suitable solvent such as petroleum ether. The petroleum ether is then evaporated off and the lipid is dispersed in a toluene-alcohol mixture and titrated with standard potassium hydroxide.

Damaged Starch

The starch in wheat occurs as partially crystalline granules. When placed in excess water, the granules will absorb about 30% of their weight. The crystallinity of the granules restricts it from absorbing additional water. During milling some of the granules are damaged. The damage results from the shear on the granule during roller milling. The shear shatters/ruptures some of the crystals. The damage may include the entire granule or just a part of it. This loss of crystals allows the granule to take up more water and swell more. Damaged starch will absorb as much as 300X its weight in water. Hard wheat flour contains a much higher level of damaged starch than does soft wheat. This apparently is because the soft wheat crushes easily during milling and does not subject the starch to as much shear.

Damaged starch is positive factor in bread flour because it increases the water absorption. High water absorption increases the yield of dough and bread from a flour, which has obvious positive effects on bakery profits. Damaged starch is a strong negative in flours for cookies and other dry finished products.

The damaged starch is highly susceptible to α- amylase attack. Much of the damaged starch is degraded to maltose and small dextrins by the combination of α- and β- amylase. This is the major reason that bread flours are malted (α- amylase added) at the mill. If the damaged starch is not removed during fermentation it interacts with the gluten and reduces bread volume.

Damaged starch is generally measured by enzymatic methods. The amount of reducing sugar produced in a certain time with excess enzyme is measured. The flour sample is subdivided into 2 subsamples, one of which is treated directly with the enzyme. The second subsample is autoclaved to gelatinize all the starch and then treated with the same enzyme system. The value obtained for the non-autoclaved sample is divided by the value for the autoclaved sample and the result is multiplied by 100. This gives the percentage of damaged starch. Most hard wheat flours will have from 6-9% damaged starch by the AACC procedure.

A second procedure used in an instrument that uses an electrode system to measure iodine. The amount of iodine bound is related to the amount of damaged starch. The procedure is accurate by requires that the electrode be properly maintained.

Flour Color

Flour color is important because it affects the crumb color of the finished product. The color of the flour used for variety breads, that have a dark color because of non-wheat components in the formula, is not important. Unbleached flours have a creamy color because of the presence of carotenoid pigments in the endosperm. The level of these pigments and therefore the color of the flour will vary from one flour to another. The level of pigments is under genetic control. The pigments can be readily bleached with benzoyl peroxide (mixed with the dry flour at the mill) or by enzyme active soy flour in the bread formula.

Flour color can be judged by visual comparison with a standard patent flour. In the Pekar (slick test), the sample flour is slicked alongside the standard sample and their colors compared visually. This procedure is also useful to determine if the sample is contaminated with bran.

In the procedure, 10-15 grams of the flour to be tested is placed on a glass, plastic, or metal plate. The surface of the flour is smoothed with a clean flour slick to a wedge approximately one-fourth inch thick at the top end of the flour sample down to a thin film at the bottom edge of the plate. The sides of the flour sample are trimmed so they form a straight edge. Next, similarly slick a second flour beside the first making certain that the two flours join and a straight edge forms between the two samples. If addition flours are to be compared, they can be placed on the plate next to the other flours and “slicked” so that there is one continuous wedge of all the flours, with a distinct line of demarcation between them. Any color differences between the samples can then be readily evaluated.

Color difference attributable to bran can be further accentuated by submerging the same samples at an angle into fresh clean water until air bubbles cease to rise (1-2 minutes). The plate is then carefully removed and placed in a warm place for the surface to dry. The relative intensity of the sample colors can then be noted after the surface has dried. The above experiment can also be carried out by dripping the glass plate containing freshly prepared flour wedges into a solution containing pyrocatechin. The bran contains the enzyme polyphenol oxidase that will convert the pyrocatechin into brown pigments. After the surface has dried, the samples are inspected for the presence of bran specks.

A number of instruments have been developed to measure the color of solids and foods. Although the may be useful with flour and baked products, they have not been readily accepted by the milling or baking industries.

Enzyme Activity

Although flour contains a large number of enzymes, only a few are measured and/or controlled. Clearly, the most important enzymes in bread flour are the amylases. Beta amylase is found in sufficient quantities in all flours. It has no action on native starch granules but does attack gelatinized and damaged starch. It acts from the non-reducing end of the gelatinized starch chain to produce maltose. It cannot go past a branch point so its action is stopped with a large part of the molecule still intact. This is called the beta limit dextrin. It will convert about 30% of the amylase and 45% of the amylopectin to maltose.

The other amylase of importance in wheat flour is α-amylase. Flour milled from sound wheat contains little or no α-amylase. Bread produced from flours with low levels of α-amylase will be low in volume and have a rough textured crumb. Thus, it is common to add malted barely or malted wheat flour to increase the α-amylase activity. Some millers will add fungal amylase preparations to increase the α-amylase activity. This requires a modified method of analysis.

Although sound grain contains low levels of α-amylase, the level of activity increases rapidly if the grain is sprouted. After the grain is mature, raising the moisture content (i.e. rain) may cause the grain to lose its dormancy and it may start to sprout while still in the field before harvest. This greatly increases the level of α-amylase and other enzymes.

α-Amylase Activity

α-Amylase breaks the α-1 – 4 bonds in starch in a more or less random attack. It is not truly random as it does not break those bonds near an α-1 – 6 branch point. Because of its attack pattern, each break dramatically reduces the size of the resulting dextrin. As a result the viscosity of the starch-water paste decreases rapidly. This is why α-amylase is sometimes referred to as the liquefying enzyme. Because of the rapid decrease in viscosity with each bond broken, measurement of viscosity is a sensitive measure of enzyme activity. The following three methods to measure α-amylase activity are all viscosity measuring procedures.

Falling Number. The falling number apparatus consists of a boiling water bath, matched test tubes (to conduct heat at the same rate), a stirrer, a stirring apparatus, and a timing mechanism. Flour plus a known amount of excess water is placed in a test tube and shaken to disperse the flour. The tube is placed in the apparatus that stirs the sample as if it is heated. At the end of stirring, the stirrer is dropped from the top position. The number of seconds required for the stirrer to fall through the flour-water paste is the falling number.

Sound flour will have a falling number of 400 seconds or greater. Increased enzyme activity will decrease the falling number. Flour milled from badly sprouted wheat may have falling numbers of 50 to 100 sec. Bakery flours are generally adjusted to 250-300 seconds. The procedure is rapid and reasonably reproducible. It can be used for either whole-wheat meal or flour.

Amylograph. In this procedure, flour and a buffer solution are stirred in a rotating bowl that is heated by an air bath. The sample is heated from room temperature to 95˚C (203˚F) at a rate of 1.5˚C/minute. If one is only interested in the α-amylase activity, the test can be ended when the slurry reaches 95˚C (203˚F). If the flour contains no α-amylase activity the viscosity (consistency) of the sample will continue to increase as the temperature rises to 95˚C. Optimumly treated bread flours are in the range of 400-600 BU. If there is increased enzyme activity, the curve will peak at a lower viscosity (consistency) and at a lower temperature. The peak height is taken as the measure of enzyme activity. The amylograph procedure is relatively slow and requires a relatively are sample. The procedure is reproducible and still widely used to control the level of malt addition.

Rapid ViscoAnalyzer (RVA). The RVA was developed as a faster and more rugged version of the amylograph. Stimulating the amylograph, the temperature control can be programmed to heat at various rates. This viscosity is determined by the load on the stirring motor. As is the case with the amylograph, the height of the viscosity vs. temperature curve is related to the α-amylase activity of the sample. Because of the flexibility in controlling heating/cooling profile, the RVA has found many uses in cereal laboratories in addition to determining α-amylase activity. The RVA can also stimulate the falling number method when samples are heated at 95˚C (203˚F) for three minutes. Stirring number is reported as the viscosity at the test’s end.

Proteolytic Activity

Proteolytic enzymes hydrolyze proteins. Proteolytic activity can be divided into two basic types. Some enzymes hydrolyze an amino acid from the end of a protein molecule while other proteolytic enzymes attack the protein chain internally. The attack is not random but instead occurs between specific amino acids. The two types of enzyme are classified as exo- (which releases amino acids from the exterior) and endo- (which breaks the protein chain internally).

Soluble Nitrogen. In general the determination of proteolytic activity is difficult. The most popular method is to measure soluble nitrogen produced from a suitable substrate. The buffered enzyme is incubated with hemoglobin (substrate) for a suitable time. The protein is precipitated and the remaining soluble nitrogen determined. The results are reported as hemoglobin units (H.U.). This is a very popular method to measure proteolytic activity but it can be misleading. The test is biased to measure exo-enzyme activity. There can be considerable endo-activity with little or no soluble nitrogen produced. Additionally, flour proteins may be degraded differently than hemoglobin.

Rheological Measurement. The chemical determination of endo-proteolytic activity is complicated and difficult. Because the endo-proteolytic enzyme significantly reduces the size of the protein molecule by its activity, it changes the rheological properties (viscosity or consistency) of the system. Thus, a dough becomes more viscous and less elastic as the result of endo-proteolytic activity. The enzyme activity can then be estimated by following the change in rheological properties as a function of time. One of the advantages of using a rheological test is that it is not affected by exo-proteolytic activity. Reducing the size of the protein by one amino acid is insignificant from a rheological viewpoint. The other advantage is that the substrate used (native gluten) and the conditions of the test (dough) both apply directly to our area of concern.

A number of rheological tests have been used to follow endo-proteolytic activity. The most appropriate appear to be the extensograph, alveograph, and lubricated compression. All of these tests will be discussed later in the chapter.

Wet gluten

Wet gluten provides a quantitative measure of the gluten forming proteins in flour that are primarily responsible for its dough mixing and baking properties.


Wheat Quality & Carbohydrate Research
Department of Plant Sciences
NDSU Dept. 7670
PO Box 6050
Fargo , ND 58108-6050
166 Loftsgard Hall
(701) 231-7737
Fax: (701) 231-8474
Site Manager: Senay Simsek
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