Analytical techniques for serratiopeptidase: A review

Gupte, Vandana; Luthra, Umesh

doi:10.1016/j.jpha.2017.03.005

Cited by 23 publications

(19 citation statements)

References 28 publications

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“…The crude extract, ow-through, puri ed enzyme, induced cell extract; wild type strain cell extract was loaded in the SDS PAGE to check the molecular weight of the protein and to show the difference in protein expression and concentration [24]. The separating gel was 12% and the stacking gel was 5 % was used to perform the SDS PAGE.…”

Section: Sds Pagementioning

confidence: 99%

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Cloning, expression, purification and amyloid degradation study of recombinant serrapeptase from Serratia marcescens

Venkataprasad

Gopinath

Ethiraj

2021

Preprint

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Background Serrapeptase (SP) is a proteolytic enzyme which plays a prominent role in the degradation of dead tissue. It can also use as a therapeutic enzyme for the treatment of Insulin amyloids. It has reported that the SP protein has the fibrinolytic activity, but the activities towards insulin amyloid degradation not reported. Our study has focused the cloning and expressing SP gene and studying its cytotoxic reactivity studies and amyloids degradation. Results SP gene was cloned in pET-28 a (+), and the enzyme was expressed using E.coli BL 21 (DE3) strain. The clone was confirmed using colony PCR, sequencing and restriction digestion. Overexpression of enzyme and purification performed using IPTG and Ni-NTA column. The molecular weight of serrapeptase protein was determined using SDS-PAGE, which found to be 52 kDa. The purified enzyme showed a zone of clearance on casein plate, slight cytotoxic reactivity to PC-12 cells after 24 h contact and 25% insulin amyloid degradation confirmed after 24 h of incubation. Conclusion This study shows that cloning and the expression of SP will reduce the time and cost of production of SP from wild type strain which can be used for the treatment of amyloid-related diseases.

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Section: Sds Pagementioning

confidence: 99%

“…The puri ed sonicated sample and supernatant was loaded in the casein plate. The plate was incubated at 37°C for 16 hours and checked for the zone clearance at patched sites [24].…”

Section: Caseinolytic Assaymentioning

confidence: 99%

Cloning, expression, purification and amyloid degradation study of recombinant serrapeptase from Serratia marcescens

Venkataprasad

Gopinath

Ethiraj

2021

Preprint

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“…Numerous studies have been carried out for quantitative and qualitative estimation of SER [12]. Thin-layer chromatography [13], SDS-PAGE gel electrophoresis [14], and X-ray powder diffractometry [15] are some of the techniques reported for successful qualitative estimation of SER.…”

Section: Manasi Et Almentioning

confidence: 99%

“…Thin-layer chromatography [13], SDS-PAGE gel electrophoresis [14], and X-ray powder diffractometry [15] are some of the techniques reported for successful qualitative estimation of SER. Quantitative instrumental methods use chromatography, field flow fractionation, electrophoresis, etc., for separation, and physical properties such as absorption, fluorescence, conductivity, and light scattering are used to measure the analyte accurately and precisely [12]. For quantitative analysis, caseinolytic assay method [16][17][18][19][20][21][22][23][24][25][26][27], spectroscopy [28][29][30][31][32][33][34][35], fluorescence spectrometry [36], enzyme-linked immunosorbent assay (ELISA) [37][38][39], radioimmunoassay [40], ultraviolet (UV) microplate method [41], high-performance LC (HPLC) [42], and LC-MS method [43] are reported.…”

Section: Manasi Et Almentioning

confidence: 99%

An Experimental Design Approach for Optimization of Modified Colorimetric First-Order Derivative Method for Estimation of Serralysin in Bulk and Pharmaceutical Formulation

2018

Asian J Pharm Clin Res

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Objective: The aim of the present work is to use experimental design to screen and optimize experimental variables for developing a colorimetric first-order derivative method for determining content of serralysin (SER) using biuret and Folin–Ciocalteu phenol reagent for stable color development. The method is based on the reaction of peptide bond in the protein with Biuret reagent in alkaline medium and further reaction of remaining tryptophan and tyrosine residues with Folin–Ciocalteu Phenol reagent to form a stable blue-colored complex (first-order derivative λmax 620 nm).Materials and Methods: A two-level full factorial design was utilized to screen the effect of Volume of NaOH (A), volume of biuret reagent (B), volume of Folin–Ciocalteu phenol reagent (C), and concentration of NaOH (D) on the formation of blue-colored SER - reagent complex (response - absorbance). A box Behnken experimental design with response surface methodology was then utilized to evaluate the main interaction and quadratic effects of these factors on the selected response.Results: With the help of a response surface plot and contour plot, the optimum values of the selected factors were determined and used for further experiments. These values were volume of NaOH (A) of 1.0 mL, volume of biuret reagent (B) of 0.25 mL, and volume of Folin–Ciocalteu phenol reagent (C) of 10 μL. The proposed method was validated according to the ICH Q2(R1) method validation guidelines. The developed colorimetric first-order derivative method was found to be simple, accurate, rapid, sensitive, precise, and economic. Further optimization of the method with experimental design approach makes it convenient for use in laboratory.Conclusion: The results of present study have clearly shown that an experimental design approach may be effectively applied to the optimization of a modified visible spectrophotometric method for estimation of SER in bulk and in pharmaceutical formulation with the least number of experimental runs possible. The method can be employed successfully for the estimation of SER in both bulk and tablet dosage form.

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“…The enzyme has an EC number 3.4.24.40 and belongs to the group Serralysin. It is originally obtained from Serratia marcescens isolated from the intestine of the silkworm Bombyx mori L. Extensive review on analytical techniques used in the qualitative and quantitative analysis of serratiopeptidase has been published by [ 3 ], where the authors pointed out a need for selective and specific techniques for the quantification of serratiopeptidase. A detailed review of existing evidences for serratiopeptidase has been reported by [ 4 ].…”

Section: Introductionmentioning

confidence: 99%