FIA-FAAS method for tannin determination based on a precipitation reaction with hemoglobin

Abstract

Um sistema em fluxo acoplado a espectrômetro de absorção atômica com chama (FIA-FAAS) foi desenvolvido para determinação de taninos em amostras de feijão guandu, explorando a reação de precipitação entre os taninos e proteínas. Extratos obtidos pela sonicação das amostras com solução de metanol 50% (v/v), foram introduzidos no sistema e induzidos à reação com hemoglobina, sendo o precipitado produzido pela reação retido em um filtro localizado no percurso analítico. Solução 1% (m/v) dodecil sulfato de sódio, fluindo em sentido inverso ao fluxo de filtração, foi empregado para solubilizar e conduzir o complexo tanino-hemoglobina ao detector, espectrômetro de absorção atômica com chama, para quantificação dos íons ferro presentes na estrutura da hemoglobina. Uma solução de ácido tânico foi utilizada para construção da curva analítica. O método proposto apresentou freqüência analítica de 30 amostras por hora, desvio padrão relativo de 9,7 % (n=10) e limite de quantificação 0,27 mg L -1 de ácido tânico.A flow system, coupled with flame atomic absorption spectrometry (FIA-FAAS), was developed for tannin determination in pigeon pea samples, exploring the precipitation reaction between tannins and proteins. Sample extracts obtained by sonication with a 50% (v/v) methanol solution were introduced into the system and induced to react with a hemoglobin solution. The precipitate produced was retained on a filter located in the analytical flow. A reversed flow of 1% (w/v) sodium dodecyl sulfate solution was used for solubilization of the precipitate from the filter and to conduct the tanninhemoglobin complex to the FAAS, to quantify the iron ions present in the hemoglobin structure. A tannic acid solution was used to prepare the analytical curve. The proposed method allowed determination of 30 samples per hour, a standard deviation of 9.7% (n=10), and a quantification limit of 0.27 mg L -1 for tannic acid.Keywords: tannin-protein complexes, hemoglobin, FIA, flame atomic absorption spectrometry IntroductionThe term tannin originated from "tanning", an expression which refers to tan animal skin. 1 This is possible because tannins link to collagen, which is the protein present in animal skin. 2 Thus, the ability to link and precipitate proteins is a property that defines tannins. For this reason, the reaction between tannins and protein has been used in many analytical methods for tannin determination, 3-6 especially when results that measure the correlation with the nutritional value of food are desired. 7 Moreover, this reaction is very specific, which minimizes interference effects in an analytical method.Tannin-protein complexes are dissolved by detergents, that break hydrophobic interaction between them, or by high pH values that ionize hydroxyphenolic groups so that it becomes unable to maintain the hydrogen bonding. 4 Usually tannin-protein precipitates are dissolved using sodium dodecyl sulfate. 7 Bate-Smith 6 proposed the use of hemoglobin for tannin determination as a chromoprotein in a spectrophotometric methodology...

Separation and Determination of Iron-Containing Proteins via Liquid Chromatography−Particle Beam/Hollow Cathode−Optical Emission Spectroscopy

“…Furthermore, the PAR assay is quantitative only when there is complete release of the metal from the denatured proteins into solution; therefore, PAR is not useful for proteins that bind the metals very tightly or in the presence of competing chelators. An assortment of atomic absorption spectrophotometry techniques, including graphite furnace atomic absorption spectrometry (GFAAS) and flame atomic absorption spectrometry (FAAS), have been employed for the detection of Fe in proteins. Inductively coupled plasma optical emission spectroscopy (ICP−OES) − and inductively coupled plasma mass spectrometry (ICP−MS) − have demonstrated accurate detection of iron-containing metalloproteins.…”

Atomic spectrometry update. Environmental analysis

“…In another study, a flow system coupled with FAAS was developed for tannin determination in pigeon pea samples. 89 The method, which utilised the precipitation reaction between tannins and proteins, in this case haemoglobin, allowed 30 samples per hour to be analysed with a quantification limit of 0.27 mg l 21 for tannic acid, from monitoring the haemoglobin Fe.…”

Determination of “free” iron and iron bound in metalloproteins via liquid chromatography separation and inductively coupled plasma-optical emission spectroscopy (LC-ICP-OES) and particle beam/hollow cathode-optical emission spectroscopy (LC-PB/HC-OES) techniques

“…[12][13][14] Beyond this approach, some form of chemical differentiation must be employed to segregate the forms of the metal, such as extraction or chromatographic methods. Both flame 15 and graphite furnace 16 atomic absorption spectrophotometry have been exploited for the analysis of iron and iron metalloproteins. Inductively coupled plasma optical emission spectroscopy (ICP-OES) has been the technique of choice for the determination of trace amounts of iron, 9,17,18 due to its short analysis time, sensitivity, experimental simplicity, multielemental capabilities, large dynamic range and its ability to completely dissociate complex (molecular) analyte species down to the individual elemental forms.…”