Recent applications of paper-based point-of-care devices for biomarker detection.

Detection of biomarkers is essential for diagnosis and prognosis of diseases for healthcare teams to provide timely appropriate treatments. Reliable, inexpensive, and sensitive devices are desirable to serve this purpose. Paper-based analytical devices (PADs) have gained enormous attention during the past decade as point-of-care devices for biomarker detection due to their simplicity, portability, and biocompatibility. Importantly, the devices highly comply with the WHO "ASSURED" concept (i.e. Affordable, Sensitive, Specific, User-friendly, Rapid and Robust, Equipment free, and Deliverable to end-users). Thus, PADs could be sustainably alternative tools for biomarker detection, especially in resources-constraint areas where budget, skillful operators, and health infrastructure are limited. First, this review introduces overviews of biomarkers, their detection and brief history of PADs. Second, description of common fabrication and detection techniques in PADs are provided. Then, it highlights recent state of the art technologies and applications of PADs for various biomarker detection published during 2018 to May 2019. Applications for tumor marker, cardiac and coronary heath disease, and metabolic disorder biomarker detection are tabulated and selected examples are described in details. Finally, it discusses the current challenges, advances, and novel applications of PADs. The review would be valuable for healthcare workers using PADs as cost-effective point-of-care devices for biomarker detection and for researchers working on future developments of the devices.

Volatile components and sensory characteristics of Thai traditional fermented shrimp pastes during fermentation periods.

Headspace-volatile components and sensory characteristics, including color, Maillard reaction products and free amino acid profiles, of two types of Thai traditional fermented shrimp paste, Kapi Ta Dam and Kapi Ta Deang, were investigated during the fermentation periods up to 6 months. The results showed that the colors of both products were changed with a decrease in CIELAB values over the fermentation period, except for yellowness of Kapi Ta Deang. Essential amino acids such as lysine and leucine and non-essential amino acids such as glutamic acid and alanine were found to be predominant free-amino acids in the products. After headspace volatile component extraction of the product was carried out using a SPME fiber coated with DVB/CAR/PDMS and analyzed by GC-MS, the main compounds responsible for distinct volatiles in the products were N-containing compounds, especially pyrazines which give roasted nutty odor. The results of sensory evaluation from panelists also suggest that fermentation period had an effect on sensory characteristics of the fermented shrimp pastes. Moreover, the sensory perceptions of the products would associate with their color, the Maillard reaction products, amino acid profiles and volatile compounds.

Characterization and in vitro biological activities of Thai traditional fermented shrimp pastes.

In this work, chemical and biological characteristics of two types of Thai fermented shrimp paste, Kapi Ta Dam and Kapi Ta Deang, at different fermentation periods and their raw materials were investigated. Kapi had low water activity and high proteins with high glutamic acid and lysine. Both Kapis, which had different sources, showed similar characteristics. The number of lactic acid bacteria in the products increased during the early stages of fermentation. Free α-amino acid contents in the products increased with the fermentation time. The water extracts from Kapi products showed strong antioxidative activities against ABTS(+) radical, and ACE inhibitory activity but they did not exhibit antimicrobial activity against Staphylococcus aureus, Bacillus cereus, Escherichia coli and Salmonella Typhimurium. Biological activities in Kapi could be developed by fermentation process, enzymatic hydrolysis of proteins and non-enzymatic browning reactions. Kapi could, thus, serve as a potential source of natural bioactive substances.

Influence of Some Hydrocolloids and Sterilization Conditions on the Physical Properties of Texture-Modified Foods Developed for the Swallow Training of Dysphagia Patients.

This research aimed to develop jelly soup for dysphagia patients at the International Dysphagia Diet Standardization Initiative (IDDSI) Framework levels 4 (puree) and 5 (minced and moist), who require swallow training to regain normal swallowing ability due to neurological issues. The study comprised three main parts: (1) an investigation of hydrocolloid types and concentrations for texture-modified foods to aid dysphagia patients during training; (2) a study of sterilization conditions and ascorbic acid's impact on physical properties (e.g., texture, viscosity, color) of the texture-modified foods; and (3) an evaluation of changes in physical, chemical, and microbial properties of the product during storage. Results revealed that the ideal recipe involved using pork bone broth with 1% κ-carrageenan for texture modification, which closely matched the properties of hospital jelly samples in terms of hardness, adhesiveness, and viscosity. Sterilization at 110 °C for 109 min effectively eliminated microorganisms without affecting the product's appearance or texture, albeit causing a slight increase in brownness. Adding ascorbic acid helped to prevent the Maillard reaction but reduced the gel strength of the sample and induced milk protein denaturation, leading to aggregation. During storage at room temperature for 9 weeks, the product became browner and less firm. Notably, no bacteria were detected throughout this period. In conclusion, this heating process is suitable for producing jelly soup to support swallow training for dysphagia patients with neurological problems. It offers invaluable assistance in their daily training to regain normal swallowing function.

Modification of Konjac Biodegradable Material Using Deacetylation and Reinforcement Process for Its Applications in Food Packaging.

Common konjac flour, especially of low grade, is a waste material produced in large quantities during purification of konjac glucomannan (KGM). It contains impurities, particularly oxalate salts, which irritate and may cause kidney stones. Konjac flour has glucomannan as a main component. Glucomannan is characterized by low crystallinity, high thermostability, and the ability to form a strong gel. Subsequently, glucomannan has good potential for the production of biodegradable material. However, its high-water affinity limits its use in packaging. The deacetylated by thermal forming process and reinforced konjac flour with 15% and 20% of microcrystalline cellulose showed improved water absorption and thermal properties of the specimen. Moreover, the thermal forming process resulted in the reduction of soluble oxalate. Therefore, due to the conditions used in this experiment, the material will be stronger, more waterproof properties, and more highly resistant to temperatures, so it is suitable to be used as a packaging that is environmentally friendly.

Effect of Ultrasonic-Assisted Enzymatic Hydrolysis on Functional Properties and Antioxidant Activity of Eri Silkworm Pupa Protein Isolate.

Philosamia ricini (Eri silkworm) pupa protein isolate (EPI) was utilized to prepare pupa protein hydrolysate (EPIH) through enzymatic hydrolysis. Additionally, the isolate underwent ultrasonic treatment at 20 kHz to become ultrasound pretreated EPI (EPIU), which was then enzymatically hydrolyzed to obtain ultrasound pretreated protein hydrolysate (EPIUH). The physicochemical properties of these samples were investigated, including molecular weight, solubility, foaming and emulsion properties, water- and oil-holding capacity, antioxidant activity, and color. When compared to EPI (used as the control), EPIU exhibited a high degree of hydrolysis at 20 minutes (DH = 29.24%). At a total process time of 20 minutes, the degree of hydrolysis for EPIH, EPIU, and EPIUH was found to be 13%, 29%, and 41%, respectively. SDS-PAGE analysis indicated no difference in molecular weight between EPI and EPIU (11-75 kDa). However, the molecular weight profiles of EPIH and EPIUH were reduced (8-45 kDa), resulting in changes in protein functionalities. The high DH value contributed to the enhancement of antioxidant activity, solubility, emulsion capacity, emulsion stability, and foam capacity of the protein isolate at pH 7. Furthermore, the ultrasonic pretreatment of the protein hydrolysate increased the lightness of the protein powder by reducing the enzyme activity of the polyphenol oxidase (PPO). These results suggest that ultrasonic pretreatment of the protein hydrolysate could be applied to improve the properties of Eri silkworm pupa protein for use in the food and beverage industry, such as protein-rich beverages or salad dressings.

Rapid separations of nile blue stained microorganisms as cationic charged species by chip-CE with LIF.

Rapid detection of microorganisms by alternative methods is desirable. Electromigration separation methods have the capability to separate microorganisms according to their charge and size and laser-induced fluorescence (LIF) detection have single-cell detection capability. In this work, a new combined separation and detection scheme was introduced using chip-based capillary electrophoresis (chip-CE) platform with LIF detection. Three microorganisms Escherichia coli, Staphylococcus aureus, and Candida albicans were selected as representatives of Gram-positive bacteria, Gram-negative bacteria, and fungi. While their cells carry an overall negative charge in neutral to alkaline pH, staining them with nile blue (NB) provided highly sensitive LIF detection with excitation and emission wavelengths at 635 nm and 685 nm, respectively, and at the same time, the overall charge was converted to positive. Electrolyte pH and concentration of polyethylene oxide (PEO) significantly affected the resolution of the microorganisms. Their optimal separation in the 14 mm separation channel was achieved in less than 30 s (R(s) > 5.3) in an electrolyte consisting of 3.94 mM Tris, 0.56 mM boric acid, 0.013 mM ethylenediaminetetraacetic acid disodium salt dihydrate (pH 10.5), and 0.025% PEO, with injection/separation voltages of +1000/+1000 V. The separation mechanism is likely employing contributions to the overall cationic charge from both the prevalently anionic membrane proteins and the cationic NB. Importantly, the resulting cationic NB-stained cells exhibited excellent separation selectivity and efficiency of ∼38000 theoretical plates for rapid separations within 30-40 s. The results indicate the potential of chip-CE for microbial analysis, which offers separations of a wide range of species with high efficiency, sensitivity, and throughput.

Application of an acid proteinase from Monascus purpureus to reduce antigenicity of bovine milk whey protein.

An acid proteinase from Monascus purpureus No. 3403, MpuAP, was previously purified and some characterized in our laboratory (Agric Biol Chem 48:1637-1639, 1984). However, further information about this enzyme is lacking. In this study, we investigated MpuAP's comprehensive substrate specificity, storage stability, and prospects for reducing antigenicity of whey proteins for application in the food industry. MpuAP hydrolyzed primarily five peptide bonds, Gln(4)-His(5), His(10)-Leu(11), Ala(14)-Leu(15), Gly(23)-Phe(24) and Phe(24)-Phe(25) in the oxidized insulin B-chain. The lyophilized form of the enzyme was well preserved at 30-40°C for 7 days without stabilizers. To investigate the possibility of reducing the antigenicity of the milk whey protein, enzymatic hydrolysates of the whey protein were evaluated by inhibition ELISA. Out of the three main components of whey protein, casein and α-lactalbumin were efficiently degraded by MpuAP. The sequential reaction of MpuAP and trypsin against the whey protein successfully degraded casein, α-lactalbumin and ß-lactoglobulin with the highest degree of hydrolysis. As a result, the hydrolysates obtained by using the MpuAP-trypsin combination showed the lowest antigenicity compared with the single application of pepsin, trypsin or pepsin-trypsin combination. Therefore, the overall result suggested that the storage-stable MpuAP and trypsin combination will be a productive approach for making hypoallergic bovine milk whey protein hydrolysates.

Antifungal activities of ethanolic extract from Jatropha curcas seed cake.

Phorbol ester extraction was carried out from Jatropha curcas seed cake, a by-product from the bio-diesel fuel industry. Four repeated extractions from 5 g J. curcas seed cake using 15 ml of 90% (v/v) ethanol and a shaking speed of 150 rev/min gave the highest yield of phosbol esters. The ethanolic extract of J. curcas seed cake showed antifungal activities against important phytofungal pathogens: Fusarium oxysporum, Pythium aphanidermatum, Lasiodiplodia theobromae, Curvularia lunata, Fusarium semitectum, Colletotrichum capsici and Colletotrichum gloeosporiodes. The extract contained phorbol esters mainly responsible for antifungal activities. The extract could therefore be used as an antifungal agent for agricultural applications.

Toxic compound, anti-nutritional factors and functional properties of protein isolated from detoxified Jatropha curcas seed cake.

Jatropha curcas is a multipurpose tree, which has potential as an alternative source for biodiesel. All of its parts can also be used for human food, animal feed, fertilizer, fuel and traditional medicine. J. curcas seed cake is a low-value by-product obtained from biodiesel production. The seed cake, however, has a high amount of protein, with the presence of a main toxic compound: phorbol esters as well as anti-nutritional factors: trypsin inhibitors, phytic acid, lectin and saponin. The objective of this work was to detoxify J. curcas seed cake and study the toxin, anti-nutritional factors and also functional properties of the protein isolated from the detoxified seed cake. The yield of protein isolate was approximately 70.9%. The protein isolate was obtained without a detectable level of phorbol esters. The solubility of the protein isolate was maximal at pH 12.0 and minimal at pH 4.0. The water and oil binding capacities of the protein isolate were 1.76 g water/g protein and 1.07 mL oil/g protein, respectively. The foam capacity and stability, including emulsion activity and stability of protein isolate, had higher values in a range of basic pHs, while foam and emulsion stabilities decreased with increasing time. The results suggest that the detoxified J. curcas seed cake has potential to be exploited as a novel source of functional protein for food applications.

Factorial design applied to a non-aqueous capillary electrophoresis method for the separation of beta-agonists.

The aim of this work was to study the effects of both chemical and instrumental parameters on the separation of beta-agonists (clenbuterol (CLE), salbutamol (SAL) and terbutaline (TER)) by non-aqueous capillary electrophoresis (NACE) method. Due to the number of parameters involved and their interactions, factorial experimental designs (EDs) at two levels was applied to investigate the influence of experimental factors (ionic strength of the background electrolyte (BGE), organic solvent, injection time, voltage and temperature) in sets of several CE responses (resolution, (RS), number of theoretical plate (N), tailing factor (TF) and migration time (tm)). As a compromise between the four responses, the optimum condition was obtained in 18 mM ammonium acetate in methanol (MeOH):acetonitrile (ACN):glacial acetic acid (66:33:1%, v/v/v) using an injection time of 4 s, the voltage and the temperature of 28 kV and 24 degrees C, respectively. The proposed NACE permitted the baseline separation of the three beta-agonists within 10.5 min with good repeatability (%RSD < 3.5%) and linearity (r2 > 0.99). The developed method was applicable for the analysis of the beta-agonists in syrup and tablets and the NACE condition was compatible with a mass spectrometer detector.

Comparison of UV spectrophotometric method and high performance liquid chromatography for the analysis of flunarizine and its application for the dissolution test.

This study aimed to develop a simple UV spectrophotometric method for the analysis and the dissolution test of flunarizine in capsules. The UV absorbance was both measured directly and by the first derivative measurements at 254 and 268 nm, respectively. The developed methods were validated for their linearity, accuracy, precision, limit of detection (LOD) and limit of quantitation (LOQ) in comparison with the reported HPLC method. The UV spectrophotometric method illustrated excellent linearity (r2 > 0.9999) in the concentration range of 6-24 microg/mL. Precision (%R.S.D. < 1.50) and recoveries were good (%R > 99.62). The LOD of direct UV and first derivative measurements were 0.09 and 0.84 microg/mL, respectively, and the LOQ were 0.26 and 2.55 microg/mL, respectively. Results from the assay of flunarizine in capsules by the UV spectrophotometric methods, both direct and first derivative measurements were not significantly different from those of the HPLC method (P > 0.05). Additionally, the method was successfully used for the dissolution test of flunarizine capsule and was found to be reliable, simple, fast, and inexpensive.

Recent applications of microchip electrophoresis to biomedical analysis.

Many separation methods have been developed for biomedical analysis, including chromatographic (e.g. high performance liquid chromatography (HPLC) and gas chromatography (GC)) and electrophoretic methods (e.g. gel electrophoresis and capillary electrophoresis (CE)). Among these techniques, CE provides advantages in terms of high separation efficiency, simplicity, low sample and solvent volume consumption, short analysis time and applicability to a wide range of biomedically important substances. Microchip electrophoresis (ME) is a miniaturized platform of CE and is now considered as a simpler and more convenient alternative, which has demonstrated potential in analytical chemistry. High-throughput, cost-effective and portable analysis systems can be developed using ME. The current review describes different separation modes and detectors that have been employed in ME to analyze various classes of biomedical analytes (e.g. pharmaceuticals and related substances, nucleic acids, amino acids, peptides, proteins, antibodies and antigens, carbohydrates, cells, cell components and lysates). Recent applications (during 2010-2014) in these areas are presented in tables and some significant findings are highlighted.

Extraction of antioxidant and ACE inhibitory peptides from Thai traditional fermented shrimp pastes.

Antioxidant and angiotensin converting enzyme (ACE) inhibitory peptides were extracted and isolated from two different types of Thai traditional fermented shrimp pastes, Kapi Ta Dam (Kp-B6) and Kapi Ta Deang (Kp-R6). Compounds with masses less than 500Da were found to be predominantly presented in both extracts. Following fractionation with sequential anion exchange chromatography and solid phase extraction (C18 matrix), three dipeptides were identified. Ser-Val and Ile-Phe were shown to exhibit ACE inhibitory activity with IC50 values of 60.68±1.06 and 70.03±1.45µM, respectively. Trp-Pro was shown to have high 2,2'-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) radical scavenging activity (EC50 17.52±0.46µM). These results indicate that Thai traditional fermented shrimp pastes are potential sources of bioactive peptides possessing ACE inhibitory and antioxidant activities.

Detoxification and anti-nutrients reduction of Jatropha curcas seed cake by Bacillus fermentation.

Jatropha curcas seed cake is a by-product generated from oil extraction of J. curcas seed. Although it contains a high amount of protein, it has phorbol esters and anti-nutritional factors such as phytate, trypsin inhibitor, lectin and saponin. It cannot be applied directly in the food or animal feed industries. This investigation was aimed at detoxifying the toxic and anti-nutritional compounds in J. curcas seed cake by fermentation with Bacillus spp. Two GRAS (generally recognized as safe) Bacillus strains used in the study were Bacillus subtilis and Bacillus licheniformis with solid-state and submerged fermentations. Solid-state fermentation was done on 10 g of seed cake with a moisture content of 70% for 7 days, while submerged fermentation was carried out on 10 g of seed cake in 100 ml distilled water for 5 days. The fermentations were incubated at the optimum condition of each strain. After fermentation, bacterial growth, pH, toxic and anti-nutritional compounds were determined. Results showed that B. licheniformis with submerged fermentation were the most effective method to degrade toxic and anti-nutritional compounds in the seed cake. After fermentation, phorbol esters, phytate and trypsin inhibitor were reduced by 62%, 42% and 75%, respectively, while lectin could not be eliminated. The reduction of phorbol esters, phytate and trypsin inhibitor was related to esterase, phytase and protease activities, respectively. J. curcas seed cake could be mainly detoxified by bacterial fermentation and the high-protein fermented seed cake could be potentially applied to animal feed.

DPPH radical scavenging activity of a mixture of fatty acids and peptide-containing compounds in a protein hydrolysate of Jatropha curcas seed cake.

Jatropha curcas, a tropical plant, has great potential commercial relevance as its seeds have high oil content. The seeds can be processed into high-quality biofuel producing seed cake as a byproduct. The seed cake, however, has not gotten much attention toward its potential usefulness. This work was aimed to determine the antioxidant activity of different fractions of a protein hydrolysate from J. curcas seed cake and to elucidate the molecular structures of the antioxidants. Seed cake was first processed into crude protein isolate and the protein was hydrolyzed by Neutrase. The hydrolysate obtained from 1 h of Neutrase hydrolysis showed the strongest antioxidant activity against DPPH radical (2,2-diphenyl-1-picrylhydrazyl). After a purification series of protein hydrolysate by liquid chromatography, chemicals acting as DPPH radical inhibitors were found to be a mixture of fatty acids, fatty acid derivatives, and a small amount of peptides characterized by mass spectrometry and nuclear magnetic resonance (NMR) spectroscopy.

Identification of newly zeaxanthin-producing bacteria isolated from sponges in the Gulf of Thailand and their zeaxanthin production.

Sponge-associated bacteria have been found to produce a variety of bioactive compounds including natural pigments. Here, we report the molecular identification of zeaxanthin-producing sponge-associated bacteria isolated from sponges in the Gulf of Thailand and the effect of environmental factors on zeaxanthin production from a bacterium. Three colorful sponge-associated bacteria (CHOB06-6, KODA19-6, and MAKB08-4) were identified based on the 16S rDNA profile. The 16S rDNA sequence-based analyses revealed that CHOB 06-6 and MAKB 08-4 were the closest relatives to Sphingomonas phyllosphaerae FA2(T), and KODA19-6 was a relative of Shingomonas (Blastomonas) natatoria DSM 3183(T). After all bacteria were cultivated in a modified Zobell medium, S. natatoria KODA19-6 was found to produce the highest zeaxanthin at 0.62 mg/l. pH and temperature considerably affected its zeaxanthin production. Its optimal condition for zeaxanthin production was found at a pH of 7 and 30 °C. The bacterium had a maximum specific growth rate (µ(max)) of 0.06 1/h with zeaxanthin productivity (Q(p)) of 6.27 µg/l·h. Therefore, this newly zeaxanthin-producing bacterium has a potential to produce natural zeaxanthin for the food, feed, pharmaceutical, and cosmetic industries.

Capillary zone electrophoresis for enumeration of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus in yogurt.

Enumeration of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus is a priority due to their importance in yogurt production. Capillary electrophoresis (CE) of both bacteria could be achieved in 7.2 min with a resolution of 3.2 in the background electrolyte (BGE) containing 4.5mM Tris(hydroxymethyl) amminomethane (TRIS)-4.5 mM boric acid-0.1 mM ethylenediamine tetraacetate (EDTA) (TBE) buffer (pH 8.4) and 0.05% (v/v) polyethylene oxide (PEO), using a capillary of 47.5 cm (effective length) x 100 microm i.d., injection of 50 mbar x 3s followed by -5kV x 120s, a voltage and temperature of 20 kV and 25 degrees C, respectively. Appropriate amounts of PEO in the BGE, sample preparation (i.e. vortex) and introduction were key factors for their separation. A short hydrodynamic injection followed by applying reversed polarity voltage could compress the bacteria into narrow zones, which were detected as separated single peaks. Method linearity (r(2)>0.99), precision (%RSDs<9.3%), recovery (%R=91.7-106.7%) and limit of quantitation (1.0 x 10(6) colony forming unit per mL (CFU/mL)) were satisfactory. Results from the CE analysis of both bacteria in yogurt were not statistically different from those of the plate count method (P>0.05). The CE method can be used as an alternative for quantitation of L. delbrueckii subsp. bulgaricus and S. thermophilus in yogurt since it was reliable, simple, cost and labor effective and rapid, allowing the analysis of 3 samples/h (comparing to 2d/sample by plate count method).