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1.
Bioresour Technol ; 296: 122323, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31698224

RESUMO

Effects of solid-state fermentation on rapid drying and spoilage prevention of potato pulp were evaluated. Pectin hydrolyzing and antibacterial ability of pectinase-secreting Aspergillus aculeatus and Bacillus subtilis were compared. A. aculeatus grew better in potato pulp, with highest pectinase yield of 342.71 ±â€¯5.09 U/mL and rapid pH reduction to 3.76 ±â€¯0.01. Next generation sequencing showed that the abundance of genera Candida and Enterobacter, which probably caused undesirable fermentation and spoilage, were significantly reduced after inoculation with A. aculeatus. In addition, fermentation with A. aculeatus significantly reduced water holding capacity from 16.63 ±â€¯0.36 g/g to 7.78 ±â€¯0.12 g/g, which resulted in lower viscosity and water binding capacity, and concomitantly significantly decreased moisture content from 76.05 ±â€¯0.24% to 12.95 ±â€¯0.19% after filtration and airflow drying. These results suggested that solid-state fermentation might be a promising technology for efficient processing and utilization of potato pulp.

2.
Food Chem ; 305: 125441, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31494499

RESUMO

Samples of granular corn starch were treated with 1,4-α-glucan branching enzyme (GBE) for 20 h using three different methods. These GBE modification methods all increased glycosidic linkage ratio, cyclic glucan content, and proportion of short chains while reducing weight mean molecular weight. The in vitro digestion rates of the modified starches were suppressed. Among these methods, a novel two-stage modification method comprising a 10-h GBE treatment, gelatinization, and a second 10-h GBE treatment, produced samples with the lowest in vitro digestibility. The rapidly digestible starch content was 34.2% lower than that of the control and 18.0% lower than that of the product of one-stage modification with the same duration. Fine structure characterization showed that more cluster structures were proved during the two-stage modification. This two-stage method suppressed the digestibility of corn starch and increased the substrate concentration, showing great potential for the industrial processing of slowly-digestible starchy foods.

3.
Biotechnol Lett ; 2019 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-31792661

RESUMO

OBJECTIVES: Analyze the thermostability, mode of action, and product specificity of a maltooligosaccharide-forming amylase from Bacillus stearothermophilus STB04 (Bst-MFA) from the biochemical and structural point of view. RESULTS: Using three-dimensional co-crystal structure of Bst-MFA with acarbose as a guide, experiments were performed to analyze the thermostability, mode of action and product specificity of Bst-MFA. The results showed that the Ca2+-Ca2+-Ca2+ metal triad of Bst-MFA is responsible for its high thermostability. Multiple substrate binding modes, rather than one productive binding mode determined by non-reducing end recognition, are in accordance with an endo-type mode of action. Significant interactions between subsites - 5 and - 6 and glucosyl residues at the non-reducing end explain the maltopentaose (G5) and maltohexaose (G6) specificity of Bst-MFA. CONCLUSIONS: Bst-MFA is a thermostable enzyme that preferentially produces G5 and G6, with an endo-type mode. The understanding of structure-function relationships provides the foundation for future efforts to the modification of Bst-MFA.

4.
Int J Biol Macromol ; 2019 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-31751711

RESUMO

The maltooligosaccharide-forming amylases (MFAses) degrade starch into maltooligosaccharides which potentially benefit human diet and grow popular in food processing, but little has been studied about their product specificity and structures. We focused on this topic and provide evidence through an X-ray crystal structure of the maltotetraose (G4)-forming amylase from Pseudomonas saccharophila STB07 (MFAps), as well as co-crystal structures of MFAps with G4 and with pseudo-maltoheptaose (pseudo-G7) determined at up to 1.1 Å resolution. G4 and pseudo-G7 occupy active cleft subsites -4 to -1 and -4 to +3 respectively. Binding induces conformational changes in the active sites except Asp193, working as the base catalyst. Comparison of the MFAps structure with those of other α-amylases revealed obvious differences in the loop structures providing dominant interactions between protein and substrate in the non-reducing side of the active sites cleft. These structures at the non-reducing end may govern the G4 specificity of MFAps and also be relevant to its exo-type action pattern.

5.
Appl Microbiol Biotechnol ; 103(23-24): 9433-9442, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31676918

RESUMO

The maltooligosaccharide-forming amylase from Bacillus stearothermophilus STB04 (Bst-MFA) randomly cleaves the α-1,4 glycosidic linkages of starch to produce predominantly maltopentaose and maltohexaose. The three-dimensional co-crystal structure of Bst-MFA with acarbose highlighted the stacking interactions between Trp139 and the substrate in subsites - 5 and - 6. Interactions like this are thought to play a critical role in maltopentaose/maltohexaose production. A site-directed mutagenesis approach was used to test this hypothesis. Replacement of Trp139 by alanine, leucine, or tyrosine dramatically increased maltopentaose production and reduced maltohexaose production. Oligosaccharide degradation indicated that these mutants also enhance productive binding of the substrate aglycone, leading to a high maltopentaose yield. Therefore, the aromatic stacking between Trp139 and substrate is suggested to control product specificity and the oligosaccharide cleavage pattern.

6.
Artigo em Inglês | MEDLINE | ID: mdl-31654380

RESUMO

By sequence comparison, the majority of 1,4-α-glucan-branching enzymes (GBEs) consist of an N-terminal carbohydrate-binding domain, a TIM-barrel catalytic domain, and a C-terminal all-beta domain. Among these structures, the GBE from Geobacillus thermoglucosidans STB02 uniquely has a highly charged 26-amino-acid C-terminal extension, whose functional roles are the least understood. In this research, the functional significance of the C-terminal domain in GBE from G. thermoglucosidans STB02 and its extension were assessed using a C-terminal deletion analysis. Mutants lacking of more than 7 residues of the C-terminal all-beta domain could not be detected in lysates of their Escherichia coli expression strains, suggesting that an intact all-beta domain is required for structural stability. In contrast, truncation of the C-terminal extension resulted in greater stability and solubility than the wild type, as well as a lower sensitivity to the presence of added metal ions. Comparison of this mutant with the wild type suggests that the interaction of metal ions with the C-terminal extension influences performance of this enzyme.

7.
J Control Release ; 313: 1-13, 2019 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-31622690

RESUMO

Non-invasive means of insulin administration circumvent some of the inconveniences of injections. Oral administration in particular is convenient, pain-free, and allows favorable glucose homeostasis, but is subject to chemical instability, enzymatic degradation, and poor gastrointestinal absorption. Natural polymeric nanoparticles have emerged as a promising oral delivery system for peptide therapeutics due their safety, biocompatibility, and stability. In this study, self-assembled nanocomposites from chitosan (CS) and insulin-loaded, zein-carboxymethylated short-chain amylose (IN-Z-CSA) nanocomposites were synthesized to improve oral bioavailability of insulin. The optimized IN-Z-CSA/CS0.2% nanocomposites exhibited an average size of 311.32±6.98 nm, a low polydispersity index (0.227±0.01), a negative zeta potential (43.77±1.36 mV), an encapsulation efficiency of 89.6±0.9%, and a loading capacity of 6.8±0.4%. The IN-Z-CSA/CS0.2% nanocomposites were stable in storage conditions. The transepithelial permeability of the N-Z-CSA/CS0.2% nanocomposites was 12-fold higher than that of insulin. Cellular uptake studies revealed that the IN-Z-CSA/CS0.2% nanocomposites were internalized into Caco-2 cells by both endocytosis and a paracellular route. Additionally, in pharmacological studies, orally administered IN-Z-CSA/CS0.2% nanocomposites had a stronger hypoglycemic effect with a relative bioavailability of 15.19% compared with that of IN-Z-CSA1.0% nanocomposites. Furthermore, cell toxicity and in vivo tests revealed that the IN-Z-CSA/CS0.2% nanocomposites were biocompatible. Overall, these results indicate that the IN-Z-CSA/CS0.2% nanocomposites can improve oral bioavailability of insulin and are a promising delivery system for insulin or other peptide/protein drugs.

8.
Int J Biol Macromol ; 2019 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-31614157

RESUMO

Naturally fermented and sundried tapioca starch is reportedly the traditional material for polvilho azedo and the primary ingredients of gluten-free food items in Brazil. This study aimed to investigate starch structure and expansion rate, high rate is acclaimed in baking application, changes of tapioca starch and potato starch during combinatorial fermentation and drying. The rate of expansion changed from 3.37 mL/g in native tapioca starch to 3.71 mL/g in fermented oven-dried tapioca starch and 6.97 mL/g in fermented sun-dried tapioca starch, while potato starch sample displayed lesser expansion on all treatments. Rapid viscosity analysis, size-exclusion chromatography, X-ray diffraction (XRD), and attenuated total reflectance Fourier transform infrared (ATR-FTIR) and electron paramagnetic resonance spectrometry (EPR) were performed to determine the structure and investigate its relationship with the expansion rate. Fermentation attacked amorphous area and cleaved glycosidic bonds. Sunlight exposure facilitated complex interactions, and crosslinking increased the molecular weight distribution (MWD) in fermented sun-dried potato starch and led to depolymerization in tapioca starch. EPR revealed an initial free radical distribution in both starches, and our results show that intensity changes in tapioca starch are essential for a high expansion capacity.

9.
Protein Expr Purif ; 164: 105478, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31421223

RESUMO

A gene encoding 1,4-α-glucan branching enzyme (GBE, EC 2.4.1.18) from the extremely thermophilic bacterium Rhodothermus obamensis STB05 was successfully cloned and expressed in Escherichia coli. Extracellular expression of the recombinant enzyme (R.o-GBE) was achieved with a yield of 1080 mg/L. Then it was purified and further characterized biochemically. R.o-GBE was optimally active at pH 7.0 and 65 °C. It remained stable at temperatures up to 80 °C and had a half-life at 85 °C of approximately 31 min. Far-UV circular dichroism and intrinsic fluorescence analyses revealed that high temperatures reduced its activity by changing the secondary and tertiary structure of R.o-GBE. The enzyme had broad pH stability between pH 3.0 and 11.0 at 4 °C, and preferred weakly acidic conditions at high temperatures. None of the metal ions enhanced the activity of R.o-GBE, but Ca2+ may be required for its activity. Its specific activity with amylopectin was 6651 U/mg, which is much higher than that reported for other GBEs. Its excellent thermostability, broad pH stability, and high specific activity make R.o-GBE highly suitable for industrial applications.

10.
Int J Biol Macromol ; 138: 394-402, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31325505

RESUMO

To better understand structure-function relationships, an X-ray crystal structure of the maltooligosaccharide-forming amylase from Bacillus stearothermophilus STB04 (Bst-MFA) with bound acarbose has been determined at 2.2 Å. The structure revealed a classical three-domain fold stabilized by four calcium ions, in which CaI-CaIII form an unprecedented linear metal triad in the interior of domain B. Catalytic residues are deduced to be two aspartic acids and one glutamic acid (Asp234, Glu264, Asp331), and the acarbose is bound to surrounding amino acid residues, mainly through extensive hydrogen bonds. Furthermore, analysis of the structure indicates the existence of at least 8 subsites in Bst-MFA, six glycone sites (-6, -5, -4, -3, -2, -1) and two aglycone sites (+1, +2). Subsite +3 remains to be further explored. Sugar-binding subsites contribute to further presentation of the oligosaccharide-binding mode, which explains the product specificity of Bst-MFA to some extent. In addition, we propose a mechanism by which maltooligosaccharide-forming amylases produce particular maltooligosaccharide products, a result different from that seen with typical α-amylases. Finally, the three-dimensional structure of Bst-MFA complexed with acarbose provides the basis for further studies, designed to increase product specificity.

11.
Int J Biol Macromol ; 138: 966-974, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31356941

RESUMO

In the present study, we investigated the effects of complexing temperatures (75 °C and 95 °C) and oleic acid (OA) content (2%, 6%, 10% and 14%, w/w, dry basis of starch) on the structure and in vitro digestibility of maize starch-OA complexes. The resistant starch content, complexing index, thermal transition temperatures, enthalpy change and relative crystallinity of the complexes prepared at 75 °C were higher than those of corresponding complexes prepared at 95 °C. Additionally, starch-10% OA complexes prepared at 75 °C had the highest resistant starch content (18.82%), complexing index (78.83%) and relative crystallinity (14.62%). Fourier transform infrared spectroscopy revealed that the intermolecular interactions between starch and OA were affected by the complexing temperatures and OA content. Raman spectroscopy indicated that the starch-10% OA complexes prepared at 75 °Ð¡ had a greater short-range molecular order than other complexes. The V-type complexes were further confirmed as V6III polymorph structures according to the spectra obtained by nuclear magnetic resonance spectroscopy. Therefore, we conclude that complexing temperatures and OA content greatly changed the fine structure of the complexes, which further affected their digestibility.

12.
BMC Neurol ; 19(1): 122, 2019 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-31185936

RESUMO

BACKGROUND: Krabbe disease (also known as globoid cell leukodystrophy) cause by a deficiency of the enzyme ß-galactocerebrosidase (galactosylceramidase, GALC). The deficiency of GALC leads to accumulation of galactosylceramide and psychosine, the latter GALC substrate having a potential role in triggering demyelination. Typically, the disease has an infantile onset, with rapid deterioration in the first few months, leading to death before the age of 2 years. The late onset forms (late-infantile, juvenile, and adult forms) are rare with variable clinical outcomes, presenting spastic paraplegia as the main symptom. CASE PRESENTATION: We recruited a family with two affected individuals. The proband (Patient 1), a 25-year-old male, was presented with slow progressive symptoms, including spastic gait disturbance and vision loss since the 5th year of life. His elder sister (Patient 2), became wheelchair-bound and demented at the age of 22 years. Brain magnetic resonance imaging (MRI) showed increased signal intensity in the white matter along with the involvement of the bilateral corticospinal tracts. GALC deficiency was confirmed by biochemical analysis. DNA sequencing revealed two mutations (c.865G > C: p. G289R and c.136G > T: p. D46Y) in GALC. The clinical characteristics, brain MRI, biochemical and molecular findings led to the diagnosis of Krabbe disease. CONCLUSION: Clinical and neuroimaged signs, positive enzymatic analysis and molecular data converged to definite diagnosis in this neurodegenerative disease.


Assuntos
Galactosilceramidase/deficiência , Galactosilceramidase/genética , Leucodistrofia de Células Globoides/genética , Adulto , Idade de Início , Grupo com Ancestrais do Continente Asiático/genética , Feminino , Humanos , Imagem por Ressonância Magnética , Masculino , Mutação , Linhagem , Irmãos , Adulto Jovem
13.
Int J Biol Macromol ; 136: 460-468, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31207329

RESUMO

Inhibition of cyclodextrin glycosyltransferases (CGTases) by their product cyclodextrins limits the efficiency of cyclodextrin production. In an effort to produce variants with good activity but reduced product inhibition, six mutants were constructed at position 603 of the CGTase from Bacillus circulans STB01, which exhibits mixed-type product inhibition. In a kinetic analysis, N603I showed reduced noncompetitive inhibition while N603K, N603H and N603R showed increased noncompetitive inhibition. Unexpectedly, N603E and N603D exhibited reductions in both competitive and noncompetitive product inhibition. Noncompetitive product inhibition is closely related to the interaction between the cyclodextrin and the enzyme in maltose binding site 2 (MBS2). Structural models led to the suggestion that there is increased interaction between maltose binding sites 1 and 2 in mutants N603E and N603D, which may have led to the unexpected results. N603D exhibited a 23.9% greater cyclodextrin yield per gram of enzyme than the wild-type, suggesting it has potential for industrial use. Further reductions in product inhibition may be gained through studies of maltose binding site interactions.

14.
Int J Biol Macromol ; 134: 247-254, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31026525

RESUMO

A new biodegradable, renewable, and environmentally friendly starch-based adhesive for wood-based panels was synthesized. The synthesis was conducted by grafting polymerization of vinyl acetate (VAC) monomer onto corn starch and crosslinking polymerization with N-methylol acrylamide (NMA). Compared with the traditional starch-based wood adhesive, the water resistance of starch-based adhesive with NMA (SWA-N) was greatly improved to more than 1 MPa; this exceeds the Chinese standard by 40%. The results from various analyses, including particle size, contact angle, thermogravimetric analysis (TGA), Fourier-transform infrared (FTIR), confocal Raman microscopy (CRM) and 13C-CPMAS, indicated that such improved performance is due to increased crosslinking density and formation of complex network structure. Such complex network structure was found to inhibit excessive expansion of the adhesive during high temperature pressing and water absorption. As a result, the internal structure of the adhesive remained intact when subjected to hot pressing and placed in wet conditions.

15.
Int J Biol Macromol ; 132: 759-765, 2019 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-30953720

RESUMO

1,4-α-Glucan branching enzyme (GBE, EC. 2.4.1.18), which plays a key role in the synthesis of starch and glycogen, has been overexpressed in E. coli as an intracellular enzyme by many researchers. In this study, it was found that the GBEs from Geobacillus thermoglucosidans and Rhodothermus obamensis were secreted into the culture medium when they were expressed separately, in E. coli. This occurred despite the absence of any signal peptide. In fact, although bioinformatics tools predicted that both of these proteins would localize to the cytoplasm, a high level of expression and non-classical secretion was found to achieve without addition of the inducer isopropyl ß-d-thiogalactopyranoside. Further experiments revealed that secretion was a two-step process that occurred via the periplasmic space. Results excluded the involvement of the Sec pathway or the TAT pathway. Instead, the findings indicated a relationship between cell membrane integrity and the secretion of the two GBEs, and suggested that their N-termini play an essential role in their expression and secretion.


Assuntos
Enzima Ramificadora de 1,4-alfa-Glucana/metabolismo , Escherichia coli/enzimologia , Enzima Ramificadora de 1,4-alfa-Glucana/química , Permeabilidade da Membrana Celular , Estabilidade Enzimática , Escherichia coli/citologia , Geobacillus/enzimologia , Rhodothermus/enzimologia
16.
Biosens Bioelectron ; 135: 200-207, 2019 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-31026774

RESUMO

This paper describes an integrated microfluidic SlipChip device for rapid antimicrobial susceptibility testing (AST) of bloodstream pathogens in positive blood cultures. Unlike conventional AST methods, which rely on an overnight subculture of positive blood cultures to obtain isolated colonies, this device enables direct extraction and enrichment of the bacteria from positive blood cultures by dielectrophoresis. SlipChip technology enables parallel inoculation of the extracted bacteria into nanoliter-scale broth droplets to perform multiplexed ASTs simultaneously. The nanoliter confinement in the droplets increases the effective inoculation amount of the bacteria, shortens the diffusion distance of nutrient elements and gases, and allows faster growth and proliferation rates. Entropy-based image analysis used for the characterization of bacterial susceptibility patterns eliminates the requirement for single-cell morphological analysis and fluorescence labeling. As a proof-of-concept, the susceptibility patterns of Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 6538p, and a positive blood culture containing Escherichia coli against several broad-spectrum antibiotics were determined by the SlipChip device. The on-chip AST results were well matched with those respectively reported by the broth microdilution method and a BD Phoenix Automated Microbiology System. Reliable AST results can be reported to clinicians within 3-8 h using this simple device after positive blood culture, allowing earlier proper administration of antimicrobial therapy.


Assuntos
Antibacterianos/farmacologia , Dispositivos Lab-On-A-Chip , Testes de Sensibilidade Microbiana/instrumentação , Técnicas Biossensoriais/instrumentação , Desenho de Equipamento , Escherichia coli/efeitos dos fármacos , Escherichia coli/crescimento & desenvolvimento , Infecções por Escherichia coli/sangue , Infecções por Escherichia coli/tratamento farmacológico , Humanos , Infecções Estafilocócicas/sangue , Infecções Estafilocócicas/tratamento farmacológico , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/crescimento & desenvolvimento
17.
Int J Biol Macromol ; 131: 564-571, 2019 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-30831164

RESUMO

In this work, cellulose was extracted from potato pulp, a low-efficiently utilized by-product from potato starch industry, by a combined alkali and sodium sulfite treatment. Its physicochemical properties were characterized and compared with cellulose from corn stalk. The yield and purity of cellulose from potato pulp were determined to be 24.74% and 81.34% respectively. Cellulose from potato pulp had more loosened and porous structure, lower crystallinity index and larger specific surface area (SSA) compared with cellulose from corn stalk. It also provided the highest accessibility to cellulase (5.7 mg protein/g glucan) and had the highest enzymatic digestibility (with glucose yield of 88.46%). Maximum adsorption capacity (Wmax) and equilibrium constant (K) were obtained by fitting the adsorption data with the Langmuir adsorption isotherm, suggesting that cellulose from potato pulp had the highest cellulase adsorption efficiency. The data further indicated that the presence of non-cellulosic components, especially for pectin, appeared to have a considerable effect on the hydrolysis by cellulase due to non-productive adsorption. State Key Laboratory of Food Science and Technology, Jiangnan University, 1800 Lihu Avenue Wuxi 214122, Jiangsu Province, People's Republic of China.


Assuntos
Celulase/química , Celulose/química , Solanum tuberosum/química , Hidrólise , Cinética , Peso Molecular , Solanum tuberosum/enzimologia , Análise Espectral
18.
Food Chem ; 283: 170-176, 2019 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-30722857

RESUMO

Maltooligosaccharide-forming amylases (MFAses) are promising tools for a variety of food industry applications because of their ability to hydrolyze starch into maltooligosaccharides. However, high thermostability is a key requirement for enzymes used in these applications. In this work, we investigated the effect of Ca2+ and Na+ on the thermostability of an MFAse from Bacillus stearothermophilus (Bst-MFAse). The results showed that Ca2+ and Na+ synergistically prolong the half-life of Bst-MFAse. The most significant improvement, which preserved 71.1% of initial activity after incubation at 80 °C for 180 min, was achieved by adding 10 mM Ca2+ and 40 mM Na+ simultaneously. The increase in Bst-MFAse thermostability imparted by the addition of Ca2+ and Na+ may be associated with an important Ca2+-Na+-Ca2+ triad structure. This study provides an effective way to enhance the thermostability of Bst-MFAse and related enzymes.


Assuntos
Amilases/metabolismo , Cálcio/farmacologia , Geobacillus stearothermophilus/enzimologia , Oligossacarídeos/biossíntese , Sódio/farmacologia , Cátions , Sinergismo Farmacológico , Estabilidade Enzimática/efeitos dos fármacos , Hidrólise , Amido/metabolismo
19.
Anal Chem ; 91(3): 1779-1784, 2019 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-30608139

RESUMO

Digital PCR is a powerful method for absolute nucleic acid quantification with unprecedented accuracy and precision. To promote the wider use and application of digital PCR, several major challenges still exist, including reduction of cost, integration of the instrumental platform, and simplification of operations. This paper describes a reusable microfluidic device that generates nanoliter droplet arrays based on step emulsification for the on-chip multiplex digital PCR of eight samples simultaneously. The device contains two glass plates that can be quickly assembled with prefilled mineral oil. Droplets are simply generated through the arrays of step emulsification nozzles driven by a single pressure controller and are self-assembled into monolayer droplet arrays in U-shaped chambers. The use of mineral oil eliminates bubble generation; thus, no overpressure is required during thermocycling. Moreover, the device can be reused many times after disassembly and a brief cleaning procedure, which significantly reduces the cost of the device per dPCR assays. The device was able to detect template DNA at concentrations as low as 10 copies/µL with a dynamic range of approximately 4 logs. We applied this device in the quantitative assessment of HER2 copy number variation, which is important for targeted therapy and prognosis of breast cancer. The performance was validated by 16 clinical samples, obtaining similar results to commercial digital PCR. We envision that this low-cost, reusable, and user-friendly device can be broadly used in various applications.

20.
J Agric Food Chem ; 66(35): 9335-9343, 2018 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-30111091

RESUMO

In this work, we use antisolvent precipitation to prepare zein/carboxymethylated short-chain amylose (CSA) complex nanoparticles for insulin encapsulation, showing that insulin-loaded zein/CSA complex nanoparticles are homogeneous, generally exhibiting sizes of <200 nm with a narrow distribution (polydispersity index < 0.100), spherical shape, and strong negative charge (-40 mV). Fourier transform infrared spectroscopy analysis reveals that the formation of the above nanoparticles is mainly driven by hydrophobic, hydrogen-bonding, and electrostatic interactions between CSA, insulin, and zein. In comparison to zein nanoparticles, zein/CSA complex nanoparticles feature much higher insulin encapsulation efficiency (45.8 versus 90.5%, respectively) and are essentially nontoxic to Caco-2 cells. Thus, this work provides new insights into the design of drug delivery systems and is expected to inspire their further development.


Assuntos
Amilose/química , Composição de Medicamentos/métodos , Insulina/química , Zeína/química , Células CACO-2 , Humanos , Ligações de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Nanopartículas/química , Tamanho da Partícula , Zeína/farmacologia
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