An efficient sample preparation procedure for rapid, reliable and reproducible analytical results is becoming increasingly important today. Ever more stringent requirements are being set both in research and production, e.g. for product monitoring and quality control. Particularly in the food, pharmaceutical and biotechnological industries as well as in medicine a meticulous sample preparation procedure is a prerequisite for accurate analysis in order to be able to observe the given limits and tolerances. The food industry depends on rapid and safe sample preparation techniques for production and quality monitoring in order to be able to intervene in running processes while losing as little time as possible. In such cases official and private food chemistry institutes and laboratories have an important key function in checking that food is generally recognized as being safe and pure. Some sample preparation techniques with RETSCH mills are described below using a current topic as an example: the detection of acrylamide in foodstuffs.
The monomer acrylamide (acrylic acid amide or, according to IUPAC, 2- propenamide) is used in industry as the basis for the manufacture of the polymer polyacrylamide, which is used in plastic packaging (e.g. also for food packaging), sealants, cosmetics and as a binder in paper and cardboard. It is also used for membranes in reverse osmosis (desalination of water) and as a flocculating agent in wastewater treatment. As monomer fractions can escape from the polyacrylamide by migration, an attempt has been made to reduce the accumulation of this hazardous substance to a minimum by the introduction of a legal limiting value. From animal studies it is known that acrylamide is a neurotoxic, carcinogenic and mutagenic substance, for whose working mechanism no significant threshold value can be assumed. As no long-term studies have yet been carried out which allow statements to be made about late-occurring damage in human beings, the risk of cancer has initially been estimated to be high even at small concentrations of a few micrograms per day. During a control test on Swedish workers exposed to acrylamide it was discovered that even apparently non-contaminated reference persons had considerable concentrations of acrylamide in their blood. Further intensive research showed that the cause of the increased acrylamide values in the reference persons could be traced to the consumption of certain foodstuffs. High concentrations were found in numerous foodstuffs such as crispbread, French fries, biscuits, chips, cookies, but also in coffee, beer, cereals, bread, etc. The trigger for the accumulated acrylamide concentrations was found to be the roasting, frying, deep-frying and baking processes used in both the industry and in private households; this provided the explanation of how acrylamide could enter the food chain. In the meantime the etiology of the formation of acrylamide in the production or finishing processes of foodstuffs has shown that not only excessive baking temperatures but also the water and fat content of the products treated during the manufacturing processes have a decisive influence. The reaction most likely to cause the formation of acrylamide is the so-called Maillard reaction, in which amino acids (aspartamic acid) react with sugars (glucose). These are present in many foodstuffs in differing concentrations and therefore provide a basis for the formation of the hazardous substance. Because of the danger resulting from an increased risk of cancer it has become essential to carry out a constant check on the acrylamide contamination in foodstuffs in order to ensure that its concentration is minimized. For this reason a rapid, reliable and accurate analytical procedure is required; a prerequisite for this is an efficient, reproducible and careful sample preparation procedure in which no analyte is lost.
Frequently used and reliable methods for detecting acrylamide are combined gas-chromatography – mass spectrometry (GC-MS) or liquid chromatography/mass spectroscopy (LC-MS/MS). In GC-MS analysis a reliable method has proved to be negative chemical ionization (CI-) with methane or ammonia, while in LC-MS/MS analysis the electro-spray technique (ESI+) is used. The quantitative determination of acrylamide is made from the relationship between the sample and measured standard solutions of D3-acrylamide, acrylamide (AA) and methacrylamide (MAA). However, before an analytical sample can be taken from the large laboratory sample that meets the requirements of the above-mentioned analytical method, the laboratory sample must be carefully prepared. Different preparation methods are used for high-fat, low-fat and fat-freeproducts. The first extremely important steps are the comminution down to particle sizes of less than 1 mm and the complete homogenization of the sample . Experience has shown that the distribution of acrylamide is very inhomogeneous in individual foodstuffs, caused for example by temperature gradients during baking or roasting. This means that the acrylamide concentration at the surface can be many times higher than that inside the food sample and could mean that a sample from a particular location could have a considerably different hazardoussubstance concentration from a sample taken at a different location. This would produce incorrect analytical results. It is also advisable to carry out a sample division after a sufficiently large laboratory sample has been homogenized in order to obtain a representative analytical sample. Comminution should take place at a temperature kept below 60 °C in order to avoid large analyte loss.
Gentle and rapid preparation of fatty samples such as chips, French fries, etc. without any noticeable temperature increase can be carried out with, for example, a knife mill (Retsch Grindomix GM 200). This uses two offset knives rotating at up to 10000 min-1 to produce a completely homogeneous pasty sample mixture in a few seconds by the effect of cutting and impact.
Dry and brittle low-fat and fat-free products such as crispbread and bread can also be comminuted and homogenized in a very short time in an ultracentrifugal mill (Retsch ZM 100). The sample material impacts at the center of a rotor rotating at 18000 min-1 and is then comminuted by impact and sheer forces between the rotor and its surrounding distance sieve ring. After passing through the sieve ring it is collected in a cassette. Depending on the selected sieve openings, this produces a particle fineness of 1 mm or less.
By the addition of water to the light-sensitive homogenizate, which therefore must be rapidly processed, a wet suspension (slurry) is produced which is then treated with 1-propanol. This extracts the acrylamide contained in the sample. The extract is concentrated, purified by the addition of acetonitrile and defatted with hexane. The concentrated mixture is then analyzed by one of the above-mentioned methods and the acrylamide concentration of the sample determined by comparison with the standard solutions.<(p>