Feasibility of Pressurization To Speed Up Enzymatic Hydrolysis of Biological Materials for Multielement Determinations

Abstract

The feasibility of pressurized solvents (liquids at a high pressure and/or high temperature without the subcritical point being reached) has been newly investigated to accelerate enzymatic hydrolysis processes of mussel tissue for multielement determinations. The target elements (Al, As, Cd, Co, Cu, Fe, Hg, Li, Mn, Pb, Se, Sr, V, and Zn) were released from dried mussel tissue by action of two proteases (pepsin and pancreatin), and they have been evaluated by inductively coupled plasma optical emission spectrometry (ICP-OES). Variables inherent to the enzymatic activity (pH, ionic strength, temperature, and enzyme mass) and factors affecting pressurization (static time, pressure, and number of cycles) were simultaneously studied by applying a Plackett-Burman design (PBD) as the screening method. Results showed that pH, ionic strength, and temperature were the most statistically significant factors (confidence interval of 95%) under pressurized conditions for pepsin, while pH and ionic strength affected pancreatin activity. This means that metal extraction is mostly attributed to enzymatic activity. The static time (enzymatic hydrolysis time) was found statistically nonsignificant for most of the elements, meaning that the hydrolysis procedure can be finished within a 2-15 min range. For pepsin, optimized conditions (pH 1.0, temperature 40 degrees C, pressure 1500 psi, static time 2 min, and number of cycles 3) gave quantitative extractions for As, Cd, Co, Cu, Hg, Li, Mn, Pb, Se, Sr, V, and Zn. The pepsin mass was 0.05 g, and the solution was Milli-Q water at pH 1.0 (adjusted with hydrochloric acid). For pancreatin, quantitative recoveries were only reached for As, Cd, Cu, Li, Pb, and Sr at room temperature, at a pressure of 1500 psi, for a static time of 2 min and a number of cycles of 3. The extraction solution was a 0.3 M potassium dihydrogen phosphate/potassium hydrogen phosphate buffer at a pH of 7.5 working at room temperature. Around 0.5 g of diatomaceous earth was used as dispersing agent for hydrolyses with either enzyme. Analytical performances, such as limits of detection and quantification and repeatability of the overall procedure, have been established. Finally, accuracy of the methods was assessed by analyzing seafood certified reference materials (GBW-08571, DORM-2, DOLT-3, TORT-2), fatty tissues certified reference materials (BCR 185, NIST 1577b), and fibrous certified reference materials (BCR 62, GBW-08501).

Extraction and Characterization of Bioactive Compounds with Health Benefits from Marine Resources: Macro and Micro Algae, Cyanobacteria, and Invertebrates

“…The developed process was preferred over more conventional extraction/hydrolysis processes based on methanol extraction and hydrochloric acid hydrolysis at 80°C, regarding primary energy consumption and global warming potential (Lindahl et al 2010 ) . Following a similar approach, integrated treatments considering enzymatic hydrolysis plus extraction carried out on different algae, such as Hizikia fusiformis (164), Ecklonia cava, and other brown algae (Athukorala et al 2006 ;Moreda-Piñeiro et al 2007 ;Turner et al 2006 ;Heo et al 2005 ) could be optimized in a more effi cient and Green way using the abovementioned concepts and ideas. Moreover, studies have demonstrated that processes such as enzymatic hydrolysis are accelerated under pressure conditions, thus giving even stronger support to the possibility of improving the processes through the use of integrated pressurized fl uid technologies (Moreda-Piñeiro et al 2007 ) .…”

Simultaneous pressurized enzymatic hydrolysis extraction and clean up for arsenic speciation in seafood samples before high performance liquid chromatography–inductively coupled plasma-mass spectrometry determination

“…The reactors were then subjected to a conventional microwave program [40]. After acid digestion was completed, the acid digests were made up to 25 mL with ultra-pure water.…”

“…A similar effect can be obtained when assisting enzymatic hydrolysis by pressurization. Thus, Moreda-Piñeiro et al [38] have proved the feasibility of pressurization (pressurized liquid extraction, PLE) to speed up enzymatic hydrolysis of marine biota (mussel), fatty biological tissues and plants for total metal extraction. The pre-treatment time was shortened to 6 min (3 PLE cycles of 2 min each one at 50 • C and 1500 psi) using pepsin or pancreatine as proteases.…”

Flame Atomic Absorption Spectrometric Determination of Trace Amounts of Silver in Radiological Waste Samples After Solid Phase Extraction Using Multi-Walled Carbon Nano-Tubes Modified by Sodium Dodecyl Sulfate

“…Thus, developing a sensitive, selective, highly precise, and economical analytical method for the monitoring of silver in environment samples with complex matrices is required. There are many sensitive but expensive analytical methods such as electrochemical analysis, 3 inductively coupled plasma mass spectrometry 4 and electrothermal atomic absorption spectrometry. 5 In addition, these instrumental techniques involve complicated processing step.…”