Search: Vitamin B6

Vitamin B₆ is separated by ion-pair RP-HPLC, detected using UV absorption and quantified through comparison with external standards. If necessary, the identity of the substances can be confirmed via spectra comparison.

HPLC methods have been successfully used for the analysis of vitamin content in non-alcoholic beverages. However, for the most part, these methods cannot be employed with a wort or beer matrix due to the vitamin concentrations present.

The reagent kit for the determination of vitamins B₁ and B₂ in blood from Chromsystems GmbH in Munich, Germany was used for the analysis. This company specializes in the manufacture of reagent kits for clinical applications. The kits are easy to use and do not require much additional equipment; a simple isocratic HPLC system is sufficient. In cooperation with the company, this kit was modified to accommodate a different sample matrix (mash, wort and beer).

The VitaFast® microbiological assay for vitamin B₆ (pyridoxine) is carried out on microtiter plates and is suitable for the determination of total amount of vitamin B₆ in foods and beverages.

Vitamin B₆ is added as pyridoxine hydrochloride or as pyridoxine-5-phosphate. Since the phosphorylated vitamin B₆ form is not recognized by the microbes in the kit, the beverage is treated with acid phosphatase to determine the amount of supplemental vitamin B₆; if necessary, the beverage is diluted with sterile water from the test kit.

The sample must be digested through enzymatic hydrolysis in order to measure the total content of vitamin B₆ (naturally occurring and added). 

The diluted sample and the vitamin B₆ assay medium are added to the wells of the microtiter plate, which are coated with Saccharomyces cerevisiae. Saccharomyces cerevisiae requires vitamin B₆ to grow. After an addition of vitamin B₆, the microorganism exhibits growth until the vitamin is completely utilized. The plates are incubated at 30 °C for 44–48 h.

The growth of Saccharomyces cerevisiae is dependent upon the concentration of vitamin B₆ and is determined through measuring the turbidity. These results are compared with those obtained from a series of standard concentrations. The measurement is performed with a microtiter plate photometer at 610–630 nm (alternatively at 540–550 nm).

Xanthohumol and isoxanthohumol are dissolved with acetonitrile from the sample and following separation, are determined using a Nucleodur C18 column and UV detection.

Inductively coupled plasma optical emission spectroscopy (ICP-OES) is a fast and reliable method for the laboratory analysis of metals. Inductively coupled plasma (ICP), a high frequency field of ionized gas, serves as a medium for atomizing and exciting the substances found in samples. Liquid, dissolved or aerosol samples are injected into the ionized gas stream. In emission spectroscopy, ICP can be used in conjunction with a number of optical and electronic systems either simultaneously or sequentially in multi-element spectrometers. In the plasma, the atoms and ions are excited to a higher energy state bringing about the emission of electromagnetic radiation (light), primarily in the ultraviolet and visible region of the spectrum. Metals ordinarily occur as ions in the temperature range typical for ICP of 6000 to 10000 K; however, non-metals and metalloids are only partially ionized.

ICP-OES operates within a very wide range. This usually encompasses six orders of magnitude in concentrations smaller than μg/l up to g/l, depending upon the element and the concentrations used for the set of analysis data. With ICP-OES, beer and wort can also be analyzed without prior processing of the samples, in contrast to AAS. Methods for determining the following in beer and wort will be described below: Al, B, Ba, Ca, Co, Cu, Fe, K, Mg, Mn, Mo, Na, P, Si, Sr, Sn and Zn.

Volatile aroma compounds are driven out of the sample through steam distillation. The ethanol distillate is adjusted to be alkaline and saturated with NaCl. The extraction of the aroma compounds is performed by shaking out with dichloromethane and the phases separated by centrifuging. The organic phase is further concentrated in a stream of nitrogen gas. An ammonia solution is added to remove the acids, because the acids would co-elute, thus preventing quantification of the target substances.