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Low-cost paper-based analytical devices are the latest generation of portable lab-on-chip designs that offers an innovative platform for the on/off-site analysis (biosensing) of target analytes, especially in rural and remote areas. Recently, microfluidic paper-based analytical devices (μPADs) have attained significant recognition owing to their exciting fundamental features such as: ease of fabrication, rapid operation, and precise interpretations. The incorporation of enzymes with paper-based analytical devices significantly improves analytical performance while exhibiting excellent chemical and storage stability. In addition to that, these devices are highly compact, portable, easy-to-use, and do not require any additional sophisticated equipment for the detection and quantification of target analytes. This review provides a holistic insight into design, fabrication, and enzyme immobilization strategies for the development of enzyme-μPADs, which enables them to be widely implemented for in-field analysis. It also highlights the recent application of enzyme-μPADs in the area of: biomedical, food safety, and environmental monitoring while exploring the mechanisms of detection involved. Further, in order to improve the accuracy of analysis, researchers have designed a smartphone-based scanning tool for multi-variant point-of-care devices, which is summarized in the latter part of the review. Finally, the future perspectives and outlook of major challenges associated with enzyme-μPADs are discussed with their possible solutions. The development of enzyme integrated μPADs will open a new avenue as an exceptional analytical tool to explore various applications. Enzyme embedded paper-based analytical devices are a revolution in the field of biosensing. The design, fabrication, and enzyme immobilization on μPADs have been comprehensively discussed. The application of enzyme-μPADs food safety, environmental monitoring, and clinical diagnostic have been reviewed. Smartphones can be used as an on-site, user-friendly, and compact next-gen scanning tool for biosensing. [ABSTRACT FROM AUTHOR]

The porous material has been considered as a potential candidate for immobilizing enzymes. Recently, metal organic framework (MOF) has been emerged as a hybrid organic inorganic material with unique intrinsic properties such as well-defined pore structure, excellent chemico-thermal stability, and extremely high surface areas which make them as a suitable scaffold for enzyme immobilization. The outstanding improvement in catalytic performance, high enzyme loading capacity, remarkable interaction between enzyme and MOF are the key features of the novel enzyme–MOF biocomposites. Amongst different immobilization approaches of enzyme–MOF composite development, de novo strategy received immense attention due to rapid, facile, mild immobilization procedure which exhibits potentially superior catalytic activity and extraordinary operational stability. This review presents a holistic insight of two different de novo strategies i.e. co-precipitation and biomineralization with state-of-art examples. Further, the recent developments in enzyme–MOF composites along with their potential features and characteristics are exploited in terms of catalytic activity, thermal/chemical stability, Michaelis–Menten kinetics, recyclability and storage stability. The advanced de novo strategies such as multi-enzyme catalytic system and magnetic enzyme–MOF are explored in the latter part of highlights. This highlight presents a holistic insight of two different approaches of de novo strategies i.e. co-precipitation and biomineralization. Unlabelled Image • The facile de novo strategies have been explored for the preparation of enzyme–MOF composites. • The potential features of enzyme–MOF are exploited in terms of activity, stability, kinetics and reusability. • The advance strategies i.e. multi-enzyme MOF and magnetic enzyme–MOF have been illustrated. • New emerging trends and possible hurdles regarding enzyme–MOF composites have been discussed. [ABSTRACT FROM AUTHOR]

Mostly, enzyme activity is reduced after immobilization of enzyme within MOF due to unfavourable conformational changes occurred during the immobilization procedure. In this context, lipase was activated by surfactants (in order to get a highly active enzyme) followed by encapsulation within zeolitic imidazolate framework (ZIF)-8 via one-pot facile self-assembly method. The prepared lipase-SDS ZIF-8 exhibited 250% enhanced activity compared to native form. The prepared biocomposite was characterized and analysed by X-ray diffraction (XRD), Fourier transform infrared (FT-IR) and scanning electron microscopy (SEM). Thermo-stability was determined for prepared lipase-SDS ZIF-8 biocomposite in the range of 50–70 °C, which showed more than two-folds enhanced stability in terms of half-life. Further, immobilized lipase retained 76% of residual activity even after six repetitive cycles and, it showed 91% residual activity after twenty days of long term storage. Finally, lipase-SDS ZIF-8 was tested for chemical stability in polar denaturing solvents which showed excellent stability as compared to free lipase. An enhanced surfactant activated lipase-SDS MOF prepared by self-assembly biomineralization method which exhibited excellent catalytic activity and recyclability. Unlabelled Image • Surfactant modified lipase was entrapped within MOF with higher retention in activity. • The prepared Lipase-SDS MOF was analysed and characterized by using FT-IR, XRD and SEM. • Improved thermal stability of Lipase-SDS MOF was expressed as half-life and deactivation energy. • Lipase-SDS MOF exhibited enhanced reusability and storage stability for industrial application. • Improved tolerance towards polar denaturing solvents was exhibited by lipase-SDS MOF. [ABSTRACT FROM AUTHOR]

Enzyme embedded metal–organic frameworks (MOFs) are blooming in the field of enzyme immobilization on account of their superior catalytic efficiency, thermal and chemical stability as compared to native enzyme. However, the separation and recovery of enzyme embedded MOF composite is quite challenging due to its nanometer size, high dispersity and low density which limits the reusability. In this work, α-amylase embedded zeolite imidazole framework (ZIF)-67 was fabricated onto melamine sponge using surfactant assisted dip coating technique. This was achieved in single pot making the method facile and rapid. α-Amylase MOF-sponge was analysed and characterized by FT-IR, XRD, SEM and TGA. We demonstrated that the encapsulation could maintain the active conformational structure of α-amylase after immobilization procedure with excellent bioactivity. Further, apart from merits of framework shielding, it offers mass transfer limitation for macromolecular substrate (like starch) which was determined and expressed as effectiveness factor (η) by using Michaelis–Menten enzyme kinetics. At the end, recycling studies were carried out for α-amylase MOF-sponge which showed 32% residual activity after six consecutive cycles. It is believed that this strategy has great potential in the field of continuous biotransformation (catalysis), biosensing and biomedicine application. A facile and highly efficient one pot synthesis method for α-amylase MOF-sponge by rapid and straightforward method. Unlabelled Image • α-Amylase embedded ZIF-67 fabricated onto melamine sponge by rapid and straightforward method. • α-Amylase MOF-sponge was analysed and characterized by FT-IR, XRD, SEM and TGA. • The melamine sponge didn't limit mass transfer of substrate even after coating of amylase MOF on it. • In reusability of α-amylase MOF-sponge, 32% residual activity was retained after six cycles. [ABSTRACT FROM AUTHOR]

An organic–inorganic hybrid glucoamylase nanoflower was prepared in single pot by simple, facile and highly efficient method. The stepwise formation of enzyme-embedded hybrid nanoflowers and influence of experimental parameters viz. pH of solution mixture, enzyme and copper ion concentration on the activity of prepared hybrid nanoflowers were systematically investigated. The self-assembled hybrid glucoamylase nanoflowers were synthesized by mixing aqueous solution of copper sulphate (200 mM) with PBS (pH 7.5, 5 mM) containing glucoamylase (1 mg/mL) in 24 h at room temperature. These prepared nanoflowers were further characterized by FT-IR, SEM and XRD. The hybrid nanoflowers exhibited 204% enhanced activity recovery and two folds improvement in thermal stability in terms of half-life (in the range of 50–70 °C) with respect to the free form. The hybrid glucoamylase nanoflowers retained 70% residual activity after eight successive cycles indicating their excellent durability. Additionally, the nanoflowers retained up to 91% residual activity upto 25 days of storage. Moreover, the conformational changes occurred in glucoamylase structure after preparing hybrid nanoflowers were evaluated by FT-IR spectroscopy data tools. [ABSTRACT FROM AUTHOR]

Abstract The catalytic efficiency of enzyme is mainly dependent on its structural conformation and its stability. In this work, laccase, the copper containing enzyme was stabilized with the help of 20 different natural amino acids by non-covalent interactions. It was seen that the positively charged amino acids viz arginine, histidine and lysine induced the favourable conformation changes in laccase which led to the enhancement in catalytic activity. Further, the thermal stability of laccase-amino acid conjugates was tested in terms of the deactivation energy (E d) and half-life (t 1/2) which showed superior thermal stability in the temperature range of 45–65 °C with respect to free form. In Michaelis-Menten kinetics studies, positively charged amino acid conjugates exhibited higher V max than free laccase. To understand the mechanism of binding of amino acids to laccase and thereby induced conformational changes in laccase, FT-IR data analysis tools and intrinsic fluorescence spectroscopy method were used. At the end, laccase-amino acid conjugates have been employed for enzymatic decolourization of dye solutions and the decolourization was found to be enhanced as compared to that by unconjugated form. Highlights • Positively charged amino acid induced hyper activation of laccase by non-covalent interaction. • In kinetics studies, positively charged amino acid conjugate exhibited higher V max than free form. • FT-IR data analysis revealed the structural conformational changes induced by amino acids. • Laccase-amino acid conjugates have been employed for enzymatic decolourization of dye solutions. [ABSTRACT FROM AUTHOR]