Multicommutated flow system for spectrophotometric L(+)lactate determination in alcoholic fermented sugar cane juice using enzymatic reaction

Abstract

Neste trabalho foi desenvolvido um procedimento empregando multicomutação em fluxo e reação enzimática para determinação espectrofotométrica de L(+)lactato proveniente da fermentação alcoólica de vinho (mosto de cana de açúcar) O método espectrofotométrico foi baseado na reação com a enzima lactato oxidase, seguida pela reação com a enzima peroxidase e posterior condensação entre 4-clorofenol e 4 -aminoantipirina. O módulo de análise foi constituído por um conjunto de três válvulas solenóides, bomba peristáltica, espectrofotômetro e microcomputador equipado com interface de controle. Um programa escrito em QuickBASIC 4.5 foi desenvolvido para controlar as válvulas solenóides e realizar aquisição de dados. Amostras na faixa de concentração de 5,0 a 100,0 mg L -1 de L(+)lactato foram analisadas usando uma solução contendo 150 UI de lactato oxidase e 200 UI de peroxidase. Outras características analíticas tais como, desvio padrão relativo de 2% (n = 15) para uma amostra típica contendo 38,5 mg L -1 de L(+)lactato; freqüência de amostragem de 36 determinações por hora; e baixo consumo de reagentes (1,0 µg de 4-clorofenol e 0,1 µg 4-aminoantipirina) por determinação também foram obtidas.A multicommutation flow system for the spectrophotometric determination of L(+)lactate in alcohol fermented sugar cane juice is proposed. The spectrophotometric method was based on the L(+)lactate conversion to pyruvate plus hydrogen peroxide by the lactate oxides enzyme, followed by reaction with peroxidase enzyme and condensation between 4-chlorophenol and 4-aminoantipyrine. The flow system network comprised three three-way solenoid valves controlled by a microcomputer that running a software wrote in Quick BASIC 4.5 provided also facilities to perform data acquisition. After optimization of system parameters, a set of alcohol fermented juice was analyzed to prove the procedure feasibility. Samples with concentration raging from 5.0 to 100.0 mg L -1 L(+)lactate were analyzed using a 150 UI lactate oxidize and 200 UI peroxidase enzymatic solution. Other advantageous features such as a relative standard deviation of 2% (n = 15) for a typical sample containing 38.5 mg L -1 L(+)lactate; an analytical throughput of 36 determination per hour; and a low reagent consumption 1.0 µg 4-chlorophenol and 0.1 µg 4-aminoantipyrine per determination were also achieved.

Selective application of two rapid, low-cost electrochemical methods to quantify glycerol according to the sample nature

“…Many techniques have been used for glycerol quantitation purposes, such as spectrophotometric/spectrofluorometric ones [4][5][6], including enzyme-based ones [7][8][9], which incidentally, have been considered to be expensive [7], electrochemical ones [10][11][12], and the chromatographic ones, the latter having been officially recommended by the AOAC, as methods 968.09 and 972.10 [13]. However, even though the chromatographic methods permit to effectively determine glycerol in the presence of many interfering compounds, this technique is considered to be ecologically unfavorable because of the use of solvents which may be dangerous for environment, and intrinsically onerous because of the usually high cost of solvents and equipment [11].…”

“…On the one hand, the enzymatic spectrophotometric proposals to determine glycerol usually rely on the use of relatively expensive either single or multi-enzymatic systems, which are disposed after have being used for a single determination. In these cases, expensive enzymes and/or cofactors such as nicotinamide-adenine dinucleotide (NAD + ) or adenosine 5 -triphosphate (ATP) are often consumed [7][8][9]12]. As an example, Li's proposal to determine glycerol in complex matrixes is based on the use of three enzymes, namely glycerol kinase (GK), pyruvate kinase (PK), and lactate dehydrogenase (LDH), apart from using ATP and NAD + [8].…”

A chemiluminescence flow-based procedure for determination of carbaryl in natural waters exploiting multicommutation and enzymatic reaction

“…This requisite could be easily achieved by implementing the analytical procedure in a flow system based on multicommutation process, which afforded ability to handle small volumes of sample and reagent solutions. 18 In this paper we intend to develop an automatic flow procedure based on multicommutation [19][20][21] for the determination of carbaryl in water. The procedure is based on the enzymatic reaction and detecting by chemiluminescence using an inexpensive instrument based on photodiode.…”

Multicommuted flow system employing pinch solenoid valves and micro-pumps

“…In these cases, the pumping devices have installed near of the solution storing vessel and the flow system worked with inner pressure higher than the atmosphere. An interesting feature of the multicommutation flow process is its ability to handle two or more solutions using a single pumping channel [23,27]. In this case, the peristaltic pump had been installed between the detector and waste-storing vessel, sample and reagent solutions flowed through the flow network by suction, therefore the inner pressure was lower than the atmosphere.…”

“…Analytical procedures based on multicommutation process have been implement employing flow system manifold designed utilizing three-way solenoid valves as the core devices [23,27,28]. Pinch solenoid valves present a functioning structure more simple than that of the three-way solenoid valve, nevertheless it has been rarely employed to develop analytical procedures [29,30].…”