Purification of α-galactosidase from pepino (Solanum muricatum) by three-phase partitioning

Şen, Aysu; Eryılmaz, Melike; Bayraktar, Hasan; Önal, Seçil

doi:10.1016/j.seppur.2011.09.026

Cited by 42 publications

(15 citation statements)

References 41 publications

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“…After mixing crude enzyme extract and selected salt from previous step, the pH of the system was adjusted to each pH values (6,7,8,9, and 10) by addition of 2M HCl or NaOH. Next, the selected organic solvent was added to the mixture with the ratios of 1: 0.5 (v/v).…”

Section: Effect Of Phs On Proteases Partitioningmentioning

confidence: 99%

“…From the previous works showed that ammonium sulfate were superior to other salts [4], [9], [11], [14] and [15]. This might be due to the Hofmeister series, which is a classification of ions based on their salting-out ability [31] and [32].…”

Section: Effect Of Salts On Proteases Partitioningmentioning

confidence: 99%

“…It has been reported that, it might not be necessary to use t-butanol always, other C3, C4 and cosolvents may also serve because they too act as a differentiating cosolvents [1]. t-Butanol binds to the precipitated proteins, thereby increasing their buoyancy and causing the precipitates to float above the denser aqueous salt layer [9]. Pigments, lipids or hydrophobic materials are concentrated in the upper phase while the polar components like remaining proteins, saccharides and cell debris are enriched in the lower phase [1] and [2].…”

Section: Effect Of Solvents On Proteases Partitioningmentioning

confidence: 99%

“…Enzymes show different behavior in TPP systems depending on the pH of the system. The efficiency of the salting out of proteins will also depend on the net charge of the proteins which is highly pH dependent [9]. Proteins tend to precipitate most readily at their isoelectric point (pI) [10].…”

Section: Effect Of Phs On Proteases Partitioningmentioning

confidence: 99%

“…TPP is a concentrating or dewatering step and some enzymes have enhanced catalytic activities in these conditions within short periods of time (about 1 h) [4]. TPP has been widely used to separate and purify various enzymes such as Aspergillus oryzae [4], protease/amylase inhibitor from wheat germ [5] and ragi (Eleusine coracana) [6], phospholipase D from Dacus carota [7], α-galactosidase form tomato [8], and pepino (Solanum muricatum) [9], invertase from baker's yeast [10], and tomato [11], proteases from papaya peels [12], Calotropis procera latex [13] and giant catfish viscera [14], inulinase from Aspergillus niger [15]. Rawdkuen et al [14] showed the best condition for separating of alkaline proteases from farmed giant catfish viscera that was consisted of the crude enzyme extract to t-butanol ratio of 1.0: 0.5 in the presence of 50% (NH 4 ) 2 SO 4 .…”

Section: Introductionmentioning

confidence: 99%

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Three-phase partitioning and proteins hydrolysis patterns of alkaline proteases derived from fish viscera

Ketnawa

Benjakul

Martínez-Álvarez

et al. 2014

Separation and Purification Technology

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In this study, the recovery alkaline proteases from farmed giant catfish viscera were isolated by using three-phase partitioning (TPP). Factors affecting partitioning efficiency such as salts, solvent types, pH, and incubation temperatures were studied. Furthermore, the application of extracted alkaline proteases on proteins hydrolysis was also determined. The system consisted of crude enzyme extract: t-butanol 1: 0.5 (w/v), 50% sodium citrate, pH 8.0 with incubation temperature of 25ºC provided the highest enzyme recovery (220%). The smear protein bands with molecular weight of 20, 24, and 215 kDa of TPP fractions appeared on the protein stained gel. Two major clear zones (24 and 130 kDa) in the interphase were observed on casein-substrate gel electrophoresis. Extracted alkaline proteases showed relatively high effective in protein hydrolysis. As a result, TPP provided high enzyme recovery and could be applied to other enzymes. The obtained alkaline proteases can be further applied in preparation of protein hydrolysates.

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Section: Effect Of Phs On Proteases Partitioningmentioning

confidence: 99%

Section: Effect Of Salts On Proteases Partitioningmentioning

confidence: 99%

Section: Effect Of Solvents On Proteases Partitioningmentioning

confidence: 99%

Section: Effect Of Phs On Proteases Partitioningmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Three-phase partitioning and proteins hydrolysis patterns of alkaline proteases derived from fish viscera

Ketnawa

Benjakul

Martínez-Álvarez

et al. 2014

Separation and Purification Technology

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Simultaneous extraction of chlorophylls, proteins, and carbohydrates from isolated Chlorella thermophila using a triphasic separation technique: A biorefinery approach

Sarkar

Bhowmick

Gayen

2023

Biofuels Bioprod Bioref

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The separation of multiple products from microalgae using a biorefinery approach is essential for the economic sustainability of microalgae-based products. In the current study, a technique for the simultaneous extraction of such products was investigated for the complete valorization of microalgae biomass. Three principal metabolites -chlorophylls, carbohydrates and proteins -were simultaneously fractionated from the biomass of isolated Chlorella thermophila using a modified version of the triphasic separation (TPS) process. Optimized process parameters were determined as: (i) ammonium sulfate as the salt; (ii) 40% (w/v) as the salt concentration; (iii) 1:1 as the salt/alcohol ratio; (iv) 0.25% (w/w) as biomass loading; (v) 6 min of homogenization time, and (vi) 10 000 rpm as homogenization speed. These optimized process parameters resulted in the simultaneous recovery of 71%, 40%, and 22% yields for proteins, carbohydrates, and chlorophylls, respectively. Ultrastructure analysis revealed that intracellular metabolites could be extracted without complete cell disruption. Finally, a comparative analysis between triphasic separation with and without homogenization demonstrated the economic feasibility of the optimized extraction process.

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Ultrasound-assisted three-phase partitioning of polyphenol oxidase from potato peel (Solanum tuberosum)

Niphadkar

Rathod

2015

Biotechnol Progress

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Conventional three phase partitioning (TPP) and ultrasound assisted three phase partitioning (UATPP) were optimized for achieving the maximum extraction and purification of polyphenol oxidase (PPO) from waste potato peels. Different process parameters such as ammonium sulfate (NH4)2SO4 concentration, crude extract to t-butanol ratio, time, temperature and pH were studied for conventional TPP. Except agitation speed, the similar parameters were also optimized for UATPP. Further additional parameters were also studied for UATPP viz. irradiation time at different frequencies, duty cycle and, rated power in order to obtain the maximum purification factor and recovery of PPO. The optimized conditions for conventional TPP were (NH4)2SO4 0-40% (w/v), extract to t-butanol ratio 1:1 (v/v), time 40 min and pH 7 at 30°C. These conditions provided 6.3 purification factor and 70% recovery of PPO from bottom phase. On the other hand, UATPP gives maximum purification fold of 19.7 with 98.3% recovery under optimized parameters which includes (NH4)2SO4 0-40% (w/v), crude extract to t-butanol ratio 1: 1 (v/v) pH 7, irradiation time 5 min with 25 kHz, duty cycle 40% and rated power 150W at 30°C. UATPP delivers higher purification factor and % recovery of PPO along with reduced operation time from 40 min to 5 min when compared with TPP. SDS PAGE showed partial purification of PPO enzyme with UATPP with molecular weight in the range of 26-36 kDa. Results reveal that UATPP would be an attractive option for the isolation and purification of PPO without need of multiple steps.

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Purification of α-galactosidase from pepino (Solanum muricatum) by three-phase partitioning

Cited by 42 publications

References 41 publications

Three-phase partitioning and proteins hydrolysis patterns of alkaline proteases derived from fish viscera

Three-phase partitioning and proteins hydrolysis patterns of alkaline proteases derived from fish viscera

Simultaneous extraction of chlorophylls, proteins, and carbohydrates from isolated Chlorella thermophila using a triphasic separation technique: A biorefinery approach

Ultrasound-assisted three-phase partitioning of polyphenol oxidase from potato peel (Solanum tuberosum)

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