Reliable Food Analysis with Reproducible Sample Preparation

Food is often inhomogeneous and occurs in a variety of consistencies. Representative samples are required by food testing laboratories to produce reproducible and meaningful analysis results. Hence, food samples should be homogenized and pulverized to the desired analytical fineness, preferably with as little time and effort as possible. In addition, the entire sample preparation process must be performed reproducibly to obtain reliable analytical results, which means that the prepared part sample, from which just a few grams or milligrams are needed for analysis, has to represent both the lab sample and the original sample from which the lab sample was extracted. The original material is not represented by an inhomogeneous sample because some components may be either overrepresented or they might be missing completely. Therefore, a homogeneous sample forms the basis for representative and reliable analytical results.

Fat analysis of pizza is a good example to recognize the importance of sample homogeneity. For analysis, just a few milligrams of pizza are more than sufficient. However, random sampling may yield a piece of salami/ cheese/ mushroom which would falsify the total fat content in the following analysis (Figure 1).  A homogeneous and representative analysis sample, however, can be acquired by reducing the pizza to coarse particles < 5 mm and then pulverizing it to fine particles < 0.5 mm.

Homogenization of the analytical sample and particle size reduction considerably reduce the standard deviation of any subsequent analysis as will be demonstrated with the pizza example (Figure 2). In the pizza samples, the fat content differs with particle sizes around 5 mm, while in the homogenized samples it is much more consistent. The standard deviation (SD) is reduced from 0.21% to 0.03% (relative SD from 2.10% to 0.35%).

From left to right: a whole pizza; sample after grinding to coarse particles < 5 mm; fully homogenized sample with particle sizes < 0.5 mm

Figure 1. From left to right: a whole pizza; sample after grinding to coarse particles < 5 mm; fully homogenized sample with particle sizes < 0.5 mm

Left: fat content varies in coarse pizza samples but is stable in the pulverized samples; right: mean values of each batch of five samples, the relative standard deviation of the fat content is reduced from 2.10% to 0.35% by homogenization

Figure 2. Left: fat content varies in coarse pizza samples but is stable in the pulverized samples; right: mean values of each batch of five samples, the relative standard deviation of the fat content is reduced from 2.10% to 0.35% by homogenization

How to Select a Suitable Laboratory Mill and Accessories

When looking for an appropriate mill and grinding tools, it should be remembered that the sample properties to be determined, like heavy metal content or moisture, are not to be modified in any way during the process. In order to select the best suited mill for a particular application, some aspects should be taken into account beforehand, for instance the sample size. This makes a huge difference when small particles such as crop grains or large samples like a whole fish have to be homogenized. Usually, mills that accept bigger initial particle sizes are not ideal for producing very fine particles, i. e. particles that are sufficiently small for following analysis. There is another important aspect that relates to the sample properties: the comminution principle of the mill should correspond with the sample’s breaking behavior to create size reduction effect. Properties such as hardness, density, fat content, consistency, and residual moisture should be taken into account. Tendency of the sample to agglomerate or temperature stability are other properties which may affect the success of the grinding process.

Laboratory mills use different size reduction principles. For instance, friction, pressure, and impact effects are best suited to pulverize brittle and hard materials. However, these size reduction principles do not have a major effect on soft, fibrous, elastic samples. For example, a freeze-dried fish cannot be homogenized by using impact or pressure but shearing and cutting are more suitable. Another important point is the sample throughput and the mill’s capacity. Grinding some kilograms of wheat in a rotor mill would be much quicker than grinding the same amount in a mixer mill with a maximum jar volume of 50 ml. Moreover, the type of grinding tool material can have a major effect on the grinding efficiency. To prevent excessive wear, the grinding set material should be harder than the sample. With regards to food samples, this holds true for the majority of the grinding materials used. Mechanical size reduction invariably causes a certain degree of abrasion which may affect the following analysis. As a result, the sample may contain traces of materials like zirconium oxide or steel. In any case, the amount is often below the detection limit for most analyses and hence can be neglected. Irrespective of the grinding tool material, the type of suitable accessories can have a major impact on the grinding efficiency.

Drying or Embrittlement of the Sample

Knife mills alone can handle moist or wet sample materials without unfavorable side effects like sample loss or blockage of the machine. As a result, material is lost and the mill has to be cleaned which can take a considerable amount of time. Hence, it would be better to dry the material prior to further processing. When selecting the drying method and temperature, it is important to ensure that the properties of the sample to be determined are not modified in any way. That is very critical when it comes to volatile or temperature-sensitive components. Often, samples of these types can only be air-dried at room temperature. For gentle and fast drying of many products, the Fluid Bed Dryer TG 200M provides a suitable solution.

There are certain types of soft, sticky, fatty or tough food samples which should be cooled before subjecting them to preliminary or fine size reduction. For example, raisins or chocolate can be easily pulverized by cryogenic grinding while at room temperature, only a rather inhomogeneous paste is produced. For easy pulverization, the sample can be embrittled in liquid nitrogen (LN2) prior to grinding. At a temperature of -196 °C, even soft jelly bears become so hard and brittle that they can be easily pulverized without any issues. Mixing the sample with dry ice (solid CO2) is yet another possibility. Cryogenic grinding is also the preferred method when volatile substances are present in the sample. Materials which must not become moist should never be directly treated with cooling agents. This is because the humidity of the air condenses on the cold sample.

Overview of Mills Commonly Used for Food Sample Preparation

Naturally, more than one mill type may be used for grinding a specific sample. As described before, the choice of the most appropriate mill for a particular sample depends on the material properties, the sample volume, the throughput, the required final fineness, and the subsequent analysis. Optimum choice for a specific application can be made by knowing the basics and working principles of different types of mills.

Rotor Mills

Ultra Centrifugal Mill ZM 200

Typical applications: seeds, corn, maize, wheat, dried algae, salt, sugar, dried fish, peas, nuts, almonds, coconut, coffee, tea, roots, gelatin, dried leaves, rice, spices, herbs, soya meal, etc.

The size of soft, brittle, medium-hard, and fibrous materials is rapidly reduced using the Ultra Centrifugal Mill ZM 200. Size reduction is effected by impact and shearing forces between horizontal rotor and ring sieve. 10 mm is the maximum feed size. A final fineness of 40 µm (d90) and below may be obtained, particularly with maximum speed, but depends on the material. This is the highest fineness that can be achieved amongst the rotor mills. The grind size is determined by the aperture sizes of the exchangeable ring sieves, ranging between 0.08 mm and 10 mm.

Ultra Centrifugal Mill ZM 200 with cyclone

Ultra Centrifugal Mill ZM 200 with cyclone

The revolution speed of the Ultra Centrifugal Mill ZM 200 ranges from 6,000 to 18,000 min-1. The cassette principle ensures 100% sample recovery and easy cleaning. A Vibratory Feeder DR 100 is recommended for automatic and uniform feeding of large quantities of free-flowing materials. If temperature-sensitive or large amounts of materials are processed, it is recommended to use a cyclone, for example, with a 5-liter or 3-liter collector. During the grinding process, frictional heat is produced that is partially discharged through the cooling effect of the cyclone.

Rotor Beater Mill SR 300

Typical applications: seeds, corn, maize, wheat, dried algae, salt, sugar, dried fish, peas, nuts, almonds, coconut, coffee, tea, roots, gelatin, dried leaves, rice, spices, herbs, soya meal, etc.

The Rotor Beater Mill SR 300 is suitable for preliminary and fine size reduction of soft, brittle, and medium-hard materials with a maximum feed size of 25 mm. The final fineness of the materials is established by the aperture size of the exchangeable ring sieves (0.08 – 10 mm). It is possible to achieve a fineness as low as 50 µm and below, based on the properties of the sample material.

Rotor Beater Mill SR 300

Rotor Beater Mill SR 300

Size reduction in the Rotor Beater Mill SR 300 is effected through impact and shearing forces. The revolution speed of the mill can be adjusted between 3,000 and 10,000 min-1. In the case of larger quantities of sample, the Vibratory Feeder DR 100 may be employed for automated feeding. Contrary to other rotor mills, the Rotor Beater Mill SR 300 is suitable for grinding large amounts of sample up to 30 liters in a single step.

Cyclone Mill Twister

Typical applications: seeds, corn, maize, wheat, peas, tea, dried leaves, rice, spices, herbs, etc.

The Cyclone Mill Twister is designed for the processing of feedstuff and non-fatty food samples for subsequent Near Infrared Spectroscopy (NIR analysis). It can process soft and fibrous products quickly and gently to the desired analytical fineness of about 0.5 m. The Twister is equipped with a rotor and grinding ring with sieve insert. The geometry of the grinding chamber and rotor and the high revolution speed of up to 14,000 min-1 create an air stream which carries the sample into the sample bottle through the integrated cyclone. Thus, cross contaminations may be prevented. Additional cooling of grinding tools and the sample is provided by the cyclone. This prevents thermal degradation and loss of moisture and makes it possible to preserve the sample properties. After separating the ground material in the cyclone, it is collected in a sample bottle for complete recovery. The speed of the rotor can be adjusted in three steps, permitting perfect adaptation to sample requirements.

Cyclone Mill Twister

Cyclone Mill Twister

Knife Mills GRINDOMIX GM 200 and GM 300

Typical applications: fresh meat, herbs, milk powder, fresh bacon, convenience food, cereal bars, soy beans, cakes, fresh fish, salad, raisins, tomatoes, fresh vegetables, sweets, jelly bears, bread, cheese, liver, fruits, chocolate, salami, soups, potatoes, cookies, ground meat, berries, nuts, seeds, boiled eggs, etc.

Knife Mills are used for the size reduction and homogenization of samples with a high oil, fat, or water content. Sample amounts up to 4,500 ml are homogenized by the GM 300 making it the only mill which can homogenize a loaf of bread or an entire pizza in a single batch. The Knife Mills have a flexible speed range, allowing for optimum adaption to the specific properties of the sample.

Operating the mills in reverse mode causes the blunt edge of the blades to hit the sample with impact and crush it (rather than cutting in forward mode). A variety of accessories is available such as different lids and knives, and containers of polycarbonate, polypropylene, glass, and stainless steel. All containers can be autoclaved apart from the polypropylene grinding container. Neutral-to-analysis knives are available for heavy-metal-free grinding processes. When a gravity lid is used, the container volume is reduced and automatically adapted to the sample amount.

Knife Mills GRINDOMIX GM 300 and GM 200

Knife Mills GRINDOMIX GM 300 and GM 200

Mixer Mill MM 400 and CryoMill

Typical applications: chocolate cream, spices, herbs, tea, olive pulp, lactose powder, egg shells, jelly bears, liver, vanilla pods, berries, cookies, tobacco, chewing gum, wheat, waffles, frozen fish, seeds, etc.

The Mixer Mill MM 400 is designed for grinding small quantities of samples up to 20 ml. Radial oscillations in a horizontal position are performed by the grinding jars with a maximum frequency of 30 Hz. Size reduction is effected by impact forces, producing a final fineness as low as 5 µm (d90), depending on the properties of the sample. With a size range of 1.5 to 50 ml, the screw-top grinding jars are suitable for wet grinding. Balls and jars are available in many different sizes and materials, for example agate or ceramics such as zirconium oxide. It is possible to use different adapters which can hold up to 8 x 50 ml conical centrifuge tubes or 10 x 5 ml or 20 x 2 ml reaction vials. The CryoMill has been specifically developed for cryogenic grinding and this will be discussed in the sub-chapter of cryogenic grinding.

Mixer Mill MM 400

Mixer Mill MM 400

Cutting Mills SM 100, SM 200, SM 300

Typical applications: roots, tea, corn, freeze dried fish, bones, mushrooms, spices, orange peel, sugar beet pellets, shea nuts, sugar cane, herbs, potatoes, lumps of cocoa butter, etc.

Cutting Mills are suitable for preliminary size reduction of fibrous, soft, or medium-hard materials such as bones, nut shells, or roots. The revolution speed of the cutting mill is either variable up to 3,000 min-1 or fixed, depending on the model. The achievable grind size is determined by the breaking properties of the sample material and the aperture size of the exchangeable bottom sieve ranging between 0.25 mm and 20 mm. There are three types of rotors available to select the best way to crush a particular sample. With the help of a cyclone, the sample is discharged more quickly from the grinding chamber, and the generated air stream results in a cooling effect.

The cutting mill series

The cutting mill series

Application Examples: Homogenization of Food

Fat Content in Sausages (GM 300)

Large fatty particles are often present in sausages, which should be completely homogenized to ensure reliable analysis results. If the few grams needed for fat content analysis are randomly extracted from the sample, there would be increased standard deviations of the analysis results. Sausages of 400 g were cut manually into pieces of about 20 mm and then ground in the GM 300 mill in two steps. Using a knife with serrated blades, the first grinding cycle was performed at a revolution speed of 4,000 min-1. It took only 25 seconds to cut the sample into pieces smaller than 5 mm. The serrated blades make it easier to tear the fibrous meat. For fat analysis, a part sample was taken directly and the remaining sample was pulverized under cryogenic conditions. For this purpose, the sample was combined with dry ice snow in 1:2 ratio after the initial grinding step, and the mixture was subsequently filled into the grinding container of stainless steel. Using a combination of the full metal knife and a lid specially designed for cryogenic grinding, the sample was pulverized at 4,000 min-1 for 3 x 20 seconds, as shown in Figure 3.

Homogenization of sausages; from left to right: original sample; pre-cut with large fatty parts; ground to < 5 mm; pulverized sample < 300 µm

Figure 3. Homogenization of sausages; from left to right: original sample; pre-cut with large fatty parts; ground to < 5 mm; pulverized sample < 300 µm

The homogenized and coarse samples were analyzed for their fat content for five consecutive times using microwave-induced drying with a Smart 6 (CEM GmbH, Germany) together with NMR spectroscopy in an Oracle fat analyzer (CEM GmbH, Germany). For every measurement, 4 g of sample were dried in 2.5 minutes and then analyzed in less than 1 minute. Compared to the finely ground samples, the fat content of the independent samples of the coarse sausage was found to differ more. The fat content of the coarser fraction was measured in a range from 14.85% to 17.12% with a SD of 0.88%. As shown in Figure 4, the SD was reduced more than 10-fold to 0.07% in the homogenized sample with a fat content ranging between 15.84% and 16.02% (relative SD reduced from 5.63% to 0.45%).

Left: fat content varies in coarse sausage samples, but is stable in fine ground samples; right: mean values of five samples each, fine grinding greatly reduced the relative standard deviation of the fat content.

Figure 4. Left: fat content varies in coarse sausage samples, but is stable in fine ground samples; right: mean values of five samples each, fine grinding greatly reduced the relative standard deviation of the fat content.

NIR Analysis of Wheat Samples (Twister)

Near Infrared Spectroscopy (NIR) is an analytical method that is often used for determining fat, moisture, protein content, and ash in a single run. This method is therefore used whenever great flexibility and high sample throughput are needed. The necessity of sample preparation is a much-discussed problem. What are the benefits of sample preparation before NIR analysis? Since the penetration depth of NIR radiation is 1 mm maximum, it is not possible to detect everything that lies underneath That is not a problem, However, if the sample is completely homogeneous; but if a sample has different layers, like seeds or grains, then only the layers as low as 1 mm are analyzed and are thus overrepresented in the measurement results. In order to show this effect, NIR was used to analyze the different properties of unground and ground wheat samples. The samples were analyzed 10 consecutive times, and for each measurement the spectrometer was refilled. In the Twister, the samples were pulverized at a revolution speed of 14,000 min-1. The results for wheat reveal a large difference between unground and ground sample, particularly with regards to the fiber and ash content (Table 1). As mentioned before, only the unground wheat surface is analyzed which led to an overrepresentation of the kernel shell. Reliable and meaningful analysis results can be obtained only through previous sample homogenization.

Table 1. Different ash and fiber contents in ground and unground samples

  Ash Moisture Fiber Fat Protein
Ground wheat
Average [%] 2.80 9.68 1.10 1.17 9.02
Standard deviation [%] 0.03 0.09 0.05 0.03 0.07
Unground wheat
Average [%] 0.10 9.80 6.90 1.38 8.46
Standard deviation [%] 0.10 0.25 0.62 0.16 0.45

 

 

Detection of Mycotoxins in Nuts (SM 300 and ZM 200)

There are certain types of food that show an increased risk of mycotoxin release caused by fungal infestation, particularly when food is stored in an unsuitable way and for a long time. Usually, fungal infestation occurs in nests and hence a random sample collected from the bulk should be large enough to enable the detection of contaminants. The first step involves the preliminary size reduction of a representative amount of 1 to 2 kg per ton of nuts in the cutting mill SM 300 to particles less than 3 mm by using the 4-mm bottom sieve. The 6-disc rotor should be used because the nut shells are very hard for the cutting effect of the other rotors. The ZM 200 is used to perform the subsequent fine size reduction. It is recommended to use distance sieves for the processing of hazelnuts. Since mycotoxins are lipophilic, the grinding process should be as gentle as possible so that fat is not released from the sample. For subsequent extraction of the mycotoxins and for HPLC analysis, a fineness of 300 µm (Figure 5) is more than adequate.

Homogenization of nuts; from left to right: original sample; sample ground to < 3 mm; pulverized sample < 300 µm

Figure 5. Homogenization of nuts; from left to right: original sample; sample ground to < 3 mm; pulverized sample < 300 µm

Detection of Polychlorinated Biphenyls in Fish (SM 300)

Fish homogenization presents a major challenge. Bones, skin, and scales are relatively resistant to size reduction which means the sample still contains larger pieces even after grinding in most mills (e. g. fresh fish in Knife Mills).

Fish has a high fat content which makes the process even more complicated, because fatty particles produce large lumps by sticking together, blocking the mill and keeping the sample inhomogeneous. This problem can be solved by freeze drying the fish and further milling in the SM 300. In the SM 300, 125 g (4 fishes, pre-cut once) of turbot or carp were pulverized at a revolution speed of 3,000 min-1, using a V-rotor which also cuts the scarp and fish bones. The sample is cooled with the use of a cyclone. After 2 minutes of grinding with a bottom sieve of 1 mm without considerable heat build-up, the fish has a particle size of approx. 1 mm (Figure 6).

Homogenization of fish; left: original sample; right: sample ground to < 1 mm

Figure 6. Homogenization of fish; left: original sample; right: sample ground to < 1 mm

Pyrrolizidine Alkaloids in Tea (ZM 200)

The group of pyrrolizidine alkaloids contains 500 chemical compounds which are largely found in leguminous plants, composite flowers, and borage family. The following parameters were used to process dried chamomile flowers: using a 0.2 mm ring sieve, 25 g of sample with a maximum particle size of 5 mm were pulverized at a revolution speed of 18,000 min-1 in the ZM 200. After a period of 2 minutes, the complete sample was ground to a final fineness of less than 100 µm, as shown in Figure 7. Continuous discharge of the material and cooling of the sample are ensured by using a cyclone. Thus, the properties of the heat-sensitive pyrrolizidine alkaloids are preserved.

Homogenization of chamomile; left: original sample; right: sample ground to < 100 µm

Figure 7. Homogenization of chamomile; left: original sample; right: sample ground to < 100 µm

Ginsenoide in Ginseng (MM 400)

In traditional Chinese medicine, ginseng has been known for many years to have beneficial health effects such as supporting the cardiovascular system and boosting immune reaction. A group of chemical substances, like ginseng saponins, appears to contribute to these beneficial effects. In the MM 400, small amounts of ginseng roots can be easily pulverized but they have to be smaller than 8 mm. First, larger sample pieces should be cut, for instance by using a Cutting Mill with a parallel section rotor. Then, 17 ml of pre-cut ginseng particles were pulverized in the Mixer Mill MM 400 in a 50 ml stainless steel grinding jar. Also used were 15 grinding balls with 10 mm diameter. After a period of 4 minutes at a frequency of 30 Hz, a final fineness of less than 100 µm was obtained (Figure 8).

Homogenization of ginseng; left to right: original sample, sample ground to < 8 mm, sample ground to < 100 µm

Figure 8. Homogenization of ginseng; left to right: original sample, sample ground to < 8 mm, sample ground to < 100 µm

Pulverizing Large Quantities of Salt (SR 300)

In addition to sodium chloride, sea salt and rock salt may also contain other silicates and minerals based on the mining area and method. Salt usually has very low element concentrations, making it necessary to process amounts in the kilogram range. Theoretically, a Cutting Mill can handle large amounts of samples but the wear would be far greater than in the SR 300, since the Cutting Mill’s cutting bars cannot process abrasive materials like salt. Charges of several kilograms can be easily pulverized with the SR 300. Frictional heat is reduced using the distance rotor. With a 5-liter collecting vessel, 5 kg of sample with a feed size up to 10 mm are pulverized in a single run at a revolution speed of 10,000 min-1. The entire sample is pulverized to less than 200 µm in just 6 minutes, as illustrated in Figure 9.

Homogenization of rock salt; left to right: original sample, sample ground to < 20 mm, sample ground to < 200 µm

Figure 9. Homogenization of rock salt; left to right: original sample, sample ground to < 20 mm, sample ground to < 200 µm

Vitamin C Analysis in Hard Candy (GM 200)

Confectionery comes in a variety of textures: it can be moist, hard, greasy, or sticky and is often inhomogeneous. For HPLC analysis, which is used to detect vitamin C content, for example in hard candy, a particle size distribution from 0.5 to 0.75 mm is ideal. For a standard homogenization process in the GM 200, 100 g of hard candy is taken which is first roughly ground for several seconds in reverse mode using the blunt side of the knife. The subsequent step involves operation in forward mode for another 15-second interval at a revolution speed of 4,000 min-1. Additional pulverization to a size less than 0.5 mm is obtained by grinding at a revolution speed of 6,000 min-1 for 6 to 12 seconds, as depicted in Figure 10. Since the sample has a high starch and sugar syrup content, this step-by-step process prevents it from adhering to the knife as is usually the case in household mixers.

Homogenization of foamy sugar; left: original sample, right: sample ground to < 500 µm

Figure 10. Homogenization of foamy sugar; left: original sample, right: sample ground to < 500 µm

Detection of GMO in Soy Beans (GM 200)

Genetically modified organisms (GMO) in food are detected using polymerase chain reaction (PCR). The sample should be homogenized before PCR. Sampling and acquiring a representative part-sample are a precondition to ensure sensitive and meaningful GMO testing. first, a laboratory sample of roughly 2.5 kg is extracted from a 20 t bulk of e.g. soy beans. For the detection of GMO, a smaller analysis sample, about 1000 g in case of soy beans or corn, is extracted from the lab sample and thoroughly homogenized in the GM 200. Only 2 mg of sample material is needed for PCR analysis. Thanks to the homogenization step, this 2 mg is representative of the entire sample. Soy bean is a grainy food that is processed in a steel container at a revolution speed of 10,000 min-1. With batches of 4 x 250 g, a grind size of less than 0.5 mm is achieved within 30 seconds, as illustrated in Figure 11.

Homogenization of soy beans; left: original sample; right: sample ground to < 500 µm

Figure 11. Homogenization of soy beans; left: original sample; right: sample ground to < 500 µm

Further Applications: Food Homogenized at Room Temperatures

Additional application examples for homogenization of food samples at room temperature are provided in the following sections.

Table 2. Application examples of food homogenized at room temperature

Sample Mill Parameters & Accessories Size reduction Remark
200 g lemons Knife Mill GRINDOMIX 8000 min-1, 10 s; gravitiy lid with overflow channels 80 mm (paste) High water content and large particle size: milling only possible in a Knife Mill
280 g lasagna Knife Mill GRINDOMIX 10 s at 4000 min-1,
20 s at 8000 min-1
80 mm (paste) Starting with short intervals helps to avoid material sticking on grinding container
500 g bread Knife Mill GRINDOMIX 1 min at 4000 min-1, knife with titanium niob coated blades 160 mm to 1.5 mm Heavy metal determination: knife with titaniumniob coated blades was used
100 g dry pear Knife Mill GRINDOMIX 15 s at 4000 min-1,
15 s at 7000 min-1
50 mm to 1 mm Homogenization of sticky material
800 g soup Knife Mill GRINDOMIX 30 s at 4000 min-1 with interval 50 mm to paste Double sealed lid for liquid samples, interval mode improves sample mixing
5 eggs Knife Mill GRINDOMIX 10 s at 10,000 min-1 70 mm to paste Very fast homogenization
100 g field beans Ultra Centrifugal Mill ZM 200 12 tooth rotor, distance sieve 1 mm, 60 s, 18,000 min-1 15 mm to 0.5 mm To avoid warming, the sample is filled into the mill slowly but continuously. The distance sieve reduces heat.
150 g gelatine Ultra Centrifugal Mill ZM 200 12 tooth rotor, distance sieve 1 mm and 0.35 mm, 45 and 120 s, 18,000 min-1, cyclone 70 mm to 0.5 mm Distance sieve to avoid warming, slow feeding required, cyclone helps to cool sample and improve sample discharge
50 g green coffee Ultra Centrifugal Mill ZM 200 12 tooth rotor, distance sieve 0.75 mm, 3 min, 18,000 min-1, cyclone 15 mm to 0.75 mm Distance sieve and cyclone reduce heat and fat release. Fat release may block sieves with small aperture sizes.
150 g corncob 1.Cutting Mill SM 300
2. Ultra Centrifugal Mill ZM 200
1. Parallel section rotor, bottom sieve 4 mm, 1500 min-1; 20 s;
2. 12 tooth rotor,
ring sieve 0.5 mm,
20 s, 18000 min-1
150 mm to 400 µm Grinding in two steps as initial sample is too big for direct feeding into the ZM 200; required final fineness achieved efficiently in the ZM 200
50 g viola roots 1.Cutting Mill SM 300
2. Ultra Centrifugal Mill ZM 200
1. 6-disc rotor, bottom sieve 4 mm, 1500 min-1, 20 s;
2. 12 tooth rotor,
ring sieve 0.5 mm,
15 s, 18000 min-1
100 mm to 200 µm Sample is too hard for manual pre-crushing, fine grinding in the ZM 200 as second step yields very fine material
5 kg tea Cutting Mill SM 300 V-rotor, 0.25 mm bottom sieve, 3000 min-1, 25 min 6 cm to 200 µm Less warming of the sample compared to the ZM 200, but same fineness & time
10 x gelatin blocks Cutting Mill SM 300 V-rotor, 6 mm bottom sieve, 3000 min-1, 10 s, cyclone 80 mm to 6 mm The cyclone is used to increase sample discharge (very light material)
10 kg oat Cutting Mill SM 300 Parallel section rotor, 6 mm bottom sieve, 700 min-1, 60 s, cyclone 6 mm to
3 mm
Reduction of speed increases required particles size, fine fraction is reduced
50 g mushrooms Cutting Mill SM 300 Parallel section rotor, 6 mm bottom sieve, 1500 min-1, 10 s 30 mm to 4 mm Sample was ground piece by piece, high coarse particle content required
2 l manioc Rotor Beater Mill SR 300 0.25 mm 360° sieve, cyclone, feeder, 10,000 min-1, 11 min 2 mm to 200 µm Vibratory feeder for uniform feeding of large quantities
20 kg roasted milk with sugar Rotor Beater Mill SR 300 2 mm 360° sieve, cyclone and feeder, 30 l receptacle, 10,000 min-1, 38 min 3 mm to 1 mm Distance sieve reduces sticking of sample; 30 l receptacle required for large sample quantity
2 kg herbs Rotor Beater Mill SR 300 0.08 mm 360° sieve, cyclone, feeder, 30 l receptacle, 10,000 min-1, 80 min 15 mm to 120 µm Vibratory feeder for uniform feeding of large quantities; 80 min processing time for 2 kg
30 g corn Cyclone Mill Twister 0.5 mm sieve, 14,000 min-1, 15 s 10 mm to 0.3 mm Quick and contamination-free grinding of nonfatty samples, high sample throughput
100 g barley Cyclone Mill Twister 1 mm sieve, 14,000 min-1, 10 s 10 mm to 1 mm Quick and contamination-free grinding of nonfatty samples, high sample throughput
50 g dry noodles Cyclone Mill Twister 2 mm sieve, 14,000 min-1, 20 s 15 mm to 0.75 mm Quick and contamination-free grinding of nonfatty samples, high sample throughput

 

Special Application: Cryogenic Grinding of Food Samples

The majority of sample materials may be ground to the preferred analytical fineness at room temperature. But there are certain limits, for instance when a slight increase in temperature negatively affects the sample; or when the material is highly elastic and will only be deformed. In addition, sticky or fatty food samples may block the mill.

When food samples are fatty, sticky, or semi-liquid (e.g. marzipan, cheese, wine gum or raisins) and simply clump together when ground at room temperature, then cryogenic grinding is the best way to pulverize such samples.

In a cryogenic grinding process, the samples do not clump together and are hence homogenized effectively. Under cryogenic conditions, the loss of volatile substances like alcohol, is limited. Residues of, for example, softeners which migrate from plastic wrappings into food, are preserved. These ingredients would escape on warming the sample during grinding. Cold milling also preserves the original structures of proteins or vitamins. Grinding aids such as dry ice (solid CO2; -78 °C) or liquid nitrogen LN2 (-196 °C) are used to perform cryogenic grinding. These grinding aids embrittle the sample and make it break more easily. The unique requirements for cryogenic grinding in different types of mills are discussed in this section. Basically, all rules and recommendations explained for room-temperature grinding should be followed for cryogenic grinding as well.

Cryogenic Grinding in the Mixer Mill MM 400 or in the CryoMill

Before embrittling, the MM 400 jar should be first filled with the grinding ball(s) and with the sample and closed tightly. No LN2 should be enclosed in the grinding jars because evaporation of this gas would significantly increase the pressure inside the grinding jar. The closed grinding jars, and hence the sample, are embrittled in a LN2 bath for 2 to 3 minutes. Appropriate grinding jars for cryogenic grinding are made of PTFE or steel, but jars made of different materials are not recommended for use. This is very important, because two different materials may react differently to high temperatures of -196 °C which may damage the jar. For cryogenic grinding, single-use vials of 1.5, 2 and 5 ml are also available. Owing to the high-energy input and the ensuing frictional heat, the grinding process should not take more than 2 minutes to prevent the sample from warming up and to retain its breaking properties. If longer grinding times are needed, these should be interrupted by intermediate cooling of the closed grinding jars. The CryoMill provides the benefit of continuous cooling of the grinding jar with LN2. Therefore, a consistent temperature of -196 °C is ensured even for prolong grinding times without the need for intermediate cooling breaks. Furthermore, the user does not come into contact with LN2 at any point. There is an automatic pre-cooling function that ensures that the grinding process does not begin before a temperature of -196 °C is reached and maintained. A zirconium oxide grinding jar can be used for heavy-metal-free grinding.

CryoMill with 50 l dewar

CryoMill with 50 l dewar

Cryogenic Grinding in the ZM 200 or in a Cutting Mill

Before embrittlement, the MM 400 jar should be filled with the grinding ball(s) and with the sample and closed tightly. No LN2 must be enclosed in the grinding jars because evaporation of this gas would significantly increase the pressure inside the jar. The closed grinding jars, and hence the sample, are embrittled in a LN2 bath for 2 to 3 minutes. Appropriate jar materials for cryogenic grinding include PTFE or steel, but jars consisting of different materials are not recommended for use. This is very important, because different materials may react differently to high temperatures of -196 °C which may cause damages to the jar. For cryogenic grinding, single-use vials of 1.5, 2 and 5 ml are also available. Owing to the high-energy input and the ensuing frictional heat, the grinding process should not take more than 2 minutes to prevent the sample from warming up and to maintain its breaking properties. If longer grinding times are needed, these should be interrupted by intermediate cooling of the closed grinding jars. The CryoMill provides the benefit of continuous cooling of the grinding jar with LN2. Therefore, a consistent temperature of -196 °C is ensured even for prolonged grinding times without the need for intermediate cooling breaks. Furthermore, the user does not come into contact with LN2 at any point. There is an automatic pre-cooling function that ensures that the grinding process does not begin before a temperature of -196 °C is reached and maintained. A zirconium oxide grinding jar should be used for heavy-metal-free grinding.

Cryogenic Grinding in Knife Mills

A knife mill can be used to perfectly homogenize hard and sticky food samples such as marzipan, cheese, wine gum or raisins. When processed at room temperature, chocolates can become into paste-like substances, but even these products can be effectively pulverized cryogenically. It is not recommended to use LN2 because the knife mills are not designed for temperatures as low as -196 °C. The sample is mixed with dry ice in 1:2 ratio, and after several minutes it is completely cooled and the grinding process begins. The dry ice continuously cools the sample.

However, no plastic accessories should be used when performing cryogenic grinding in the knife mills as these can be easily damaged during the process. Ideal accessories include a full metal knife, a grinding container of stainless steel, and a lid with aperture to enable the evaporation of the carbon dioxide gas.

Table 3. Application examples of cryogenic grinding of food

Sample Mill Parameters & Accessories Size reduction Remark
10 jelly bears Mixer Mill MM 400 1 min, 30 Hz 20 mm to 300 µm Grinding in 50 ml grinding jar + grinding ball 25 mm (both stainless steel)
20 g chewing gum Mixer Mill MM 400 30 s, 30 Hz 15 mm to 500 µm Grinding in 50 ml grinding jar + grinding ball 25 mm (both stainless steel)
10 g liver Mixer Mill MM 400 2 min, 30 Hz 6 mm to 400 µm Grinding in 50 ml grinding jar + grinding ball 25 mm (both stainless steel)
3 g vanilla pod Mixer Mill MM 400 20 s, 30 Hz 10 mm to 500 µm Grinding in 50 ml grinding jar + grinding ball 25 mm (both stainless steel)
2 g cherries CryoMill 10 s, 30 Hz 15 mm to 600 µm Pre-cooling of approx. 5 min is typical. Grinding in 50 ml grinding jar + grinding ball 25 mm (both stainless steel)
6 g licorice CryoMill 2 min, 30 Hz 10 mm to 300 µm Pre-cooling of approx. 5 min is typical. Grinding in 50 ml grinding jar + grinding ball 25 mm (both stainless steel)
9 g coffee CryoMill 15 min, 30 Hz 10 mm to 150 µm Pre-cooling of approx. 5 min is typical. Grinding in 50 ml grinding jar + grinding ball 25 mm (both stainless steel)
5 g cheese CryoMill 2 min, 30 Hz 8 mm to 300 µm Pre-cooling of approx. 5 min is typical. Grinding in 50 ml grinding jar + grinding ball 25 mm (both stainless steel)
1 praline CryoMill 2 min, 30 Hz 10 mm to 400 µm Pre-cooling of approx. 5 min is typical. Grinding in 50 ml grinding jar + grinding ball 25 mm (both stainless steel)
500 g wine gum Knife Mill GRINDOMIX 40 s at 2000 min-1 reverse; 20 s at 4000 min-1 forward 20 mm to 0.8 mm Grinding container stainless steel, full metal knife, cryo lid with aperture; dry ice.
300 g block of marzipan Knife Mill GRINDOMIX 20 s at 2000 min-1 reverse; 20 s at 4000 min-1 forward 40 mm to 800 µm Grinding container stainless steel, full metal knife, cryo lid with aperture; dry ice.
400 g pure bacon Knife Mill GRINDOMIX 45 s at 2000 min-1 reverse; 30 s at 4000 min-1 forward 30 mm to 1 mm Grinding container stainless steel, full metal knife, cryo lid with aperture; dry ice.
800 g raisins Knife Mill GRINDOMIX 45 s at 2000 min-1 reverse 15 mm to 0.5 mm Grinding container stainless steel, full metal knife, cryo lid with aperture; dry ice.
100 g cereals Ultra Centrifugal Mill ZM 200 12 tooth rotor, ring sieve 0.5 mm, 3 min, 18,000 min-1 8 mm to 250 µm Use of cyclone and LN2
100 g dried apples Ultra Centrifugal Mill ZM 200 12 tooth rotor, ring sieve 0.5 mm, 1 min, 18,000 min-1 5 mm to 250 µm Use of cyclone and dry ice.
15 g toffee candy Ultra Centrifugal Mill ZM 200 12 tooth rotor, ring sieve 2 mm and 0.5 mm, 1 min, 18,000 min-1 10 mm to 500 µm Use of cyclone and dry ice.
1 kg trout Cutting Mill SM 300 6-disc rotor, 20 mm bottom sieve, 700 min-1, 60 s 200 mm to 20 mm Use of cyclone and LN2; reduced speed leads to less heat built-up
500 g lump of cocoa butter Cutting Mill SM 300 6-disc rotor, 6 mm bottom sieve, 700 min-1, 90 s 100 mm to 6 mm Use of cyclone and LN2; reduced speed leads to less heat built-up
20 kg sweet potatoe Cutting Mill SM 300 6-disc rotor, 20 mm bottom sieve, 1500 min-1, 15 min 100 mm to 20 mm Use of cyclone and dry ice

 

Conclusion

In this article it was demonstrated by many application examples that sample preparation before any food analysis is an important step of the quality control process, because reliable and reproducible analysis results can only be obtained from fully homogenized samples. With a wide range of laboratory mills and accessories available, all aspects of the sample preparation process should be considered before choosing an appropriate device.

Only then can this significant step before sample analysis be performed in the most reliable and efficient way. Armed with the knowledge of the sample characteristics and the available types of mills and accessories, users can process their samples with best possible results, ideally with as little time and effort as possible.

This information has been sourced, reviewed and adapted from materials provided by RETSCH GmbH.

For more information on this source, please visit RETSCH GmbH.

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