Assessment of Multifunctional Activity of a Postbiotic Preparation Derived from Lacticaseibacillus paracasei Postbiotic-P6.

Postbiotics possess various functional activities, closely linked to their source bacterial strains and preparation methods. Therefore, the functional activities of postbiotics need to be evaluated through in vitro and in vivo methods. This study aims to prepare a postbiotic and explore its antihemolytic, anti-inflammatory, antioxidant, and antibacterial activities. Specifically, a postbiotic preparation named PostbioP-6 was prepared by intercepting 1-5 kDa of Lacticaseibacillus paracasei Postbiotic-P6 fermentation broth. The results demonstrate that PostbioP-6 exhibited notable biological activities across multiple assays. It showed significant antihemolytic activity, with a 4.9-48.1% inhibition rate at 10-50% concentrations. Anti-inflammatory effects were observed both in vitro, where 8-40% PostbioP-6 was comparable to 259.1-645.4 µg/mL diclofenac sodium, and in vivo, where 3.5 and 4.0 µL/mL PostbioP-6 significantly reduced neutrophil counts in inflamed zebrafish (p < 0.05). Antioxidant properties were evident through increased reducing power (OD700 increased from 0.279 to 2.322 at 1.25-12.5% concentrations), DPPH radical scavenging activity (38.9-92.4% scavenging rate at 2.5-50% concentrations), and hydroxyl radical scavenging activity (4.66-10.38% scavenging rate at 0.5-4% concentrations). Additionally, PostbioP-6 demonstrated antimicrobial activity against two Gram-positive bacteria, eight Gram-negative bacteria, and one fungus. Furthermore, PostbioP-6 significantly inhibited the increase in peroxide value and malondialdehyde content in cookies, highlighting its potential application in food preservation. In conclusion, we prepared a novel postbiotic, termed PostbioP-6, which proved to have prominent anti-hemolytic, anti-inflammatory, antioxidant, and broad-spectrum antimicrobial activities. The multifunctional properties of PostbioP-6 position it as a potentially effective functional food supplement or preservative. In the future, further research is necessary to elucidate the precise mechanisms of action, identify the active components, and validate its biological activities in animal models or clinical trials.

Real-time fluorescence lifetime imaging system with a 32 x 32 0.13microm CMOS low dark-count single-photon avalanche diode array.

A compact real-time fluorescence lifetime imaging microscopy (FLIM) system based on an array of low dark count 0.13microm CMOS single-photon avalanche diodes (SPADs) is demonstrated. Fast background-insensitive fluorescence lifetime determination is achieved by use of a recently proposed algorithm called 'Integration for Extraction Method' (IEM) [J. Opt. Soc. Am. A 25, 1190 (2008)]. Here, IEM is modified for a wider resolvability range and implemented on the FPGA of the new SPAD array imager. We experimentally demonstrate that the dynamic range and accuracy of calculated lifetimes of this new camera is suitable for widefield FLIM applications by imaging a variety of test samples, including various standard fluorophores covering a lifetime range from 1.6ns to 16ns, microfluidic mixing of fluorophore solutions, and living fungal spores of Neurospora Crassa. The calculated lifetimes are in a good agreement with literature values. Real-time fluorescence lifetime imaging is also achieved, by performing parallel 32 x 16 lifetime calculations, realizing a compact and low-cost FLIM camera and promising for bigger detector arrays.

Differences in proteomic profiles of milk fat globule membrane in yak and cow milk.

Milk fat globule membrane (MFGM) is an important milk component which is rich in bioactive proteins. In this work, the isobaric tags for relative and absolute quantitation (iTRAQ) proteomic approach was used to investigate the differences in the MFGM proteins between yak and cow milk. Over 450 proteins were identified between the yak and cow MFGM. The MFGM proteins with significant differences were compared based on the relative abundance. Proteins such as Glycosylation-dependent cell adhesion molecule 1 (GlyCAM1), CD59 molecule and lactadherin, were identified having a much higher abundance (4.6-10.1 fold) in yak MFGM than cow MFGM. These proteins are thought to have biological functions such as the antimicrobial and antitumor effects. This may be due to the need that yak produces high nutritive milk including high levels of bioactive compounds in order to resist the extreme high altitude environment.

Hardware implementation algorithm and error analysis of high-speed fluorescence lifetime sensing systems using center-of-mass method.

A new, simple, high-speed, and hardware-only integration-based fluorescence-lifetime-sensing algorithm using a center-of-mass method (CMM) is proposed to implement lifetime calculations, and its signal-to-noise-ratio based on statistics theory is also deduced. Compared to the commonly used iterative least-squares method or the maximum-likelihood-estimation-based, general purpose fluorescence lifetime imaging microscopy (FLIM) analysis software, the proposed hardware lifetime calculation algorithm with CMM offers direct calculation of fluorescence lifetime based on the collected photon counts and timing information provided by in-pixel circuitry and therefore delivers faster analysis for real-time applications, such as clinical diagnosis. A real-time hardware implementation of this CMM FLIM algorithm suitable for a single-photon avalanche diode array in CMOS imaging technology is now proposed for implementation on field-programmable gate array. The performance of the proposed methods has been tested on Fluorescein, Coumarin 6, and 1,8-anilinonaphthalenesulfonate in water/methanol mixture.

Fluorescence lifetime biosensing with DNA microarrays and a CMOS-SPAD imager.

Fluorescence lifetime of dye molecules is a sensitive reporter on local microenvironment which is generally independent of fluorophores concentration and can be used as a means of discrimination between molecules with spectrally overlapping emission. It is therefore a potentially powerful multiplexed detection modality in biosensing but requires extremely low light level operation typical of biological analyte concentrations, long data acquisition periods and on-chip processing capability to realize these advantages. We report here fluorescence lifetime data obtained using a CMOS-SPAD imager in conjunction with DNA microarrays and TIRF excitation geometry. This enables acquisition of single photon arrival time histograms for a 320 pixel FLIM map within less than 26 seconds exposure time. From this, we resolve distinct lifetime signatures corresponding to dye-labelled HCV and quantum-dot-labelled HCMV nucleic acid targets at concentrations as low as 10 nM.

Hardware implementation and calibration of background noise for an integration-based fluorescence lifetime sensing algorithm.

A new integration based fluorescence lifetime imaging microscopy (FLIM) called IEM has been proposed to implement lifetime extraction [J. Opt. Soc. Am. A25, 1190 (2008)]. A real-time hardware implementation of the IEM FLIM algorithm suitable for single photon avalanche diode arrays in nanometer-scale CMOS technology is now proposed. The problems of reduced pixel readout bandwidth and background noise are studied and a calibration method suitable for FPGA implementation is introduced. In particular, the relationship between signal-to-noise ratio and background noise is considered based on statistics theory and compared with a rapid lifetime determination method and maximum-likelihood estimator with-without background correction. The results are also compared with Monte Carlo simulations giving good agreement. The performance of the proposed methods has been tested on monoexponential decay experimental data. The high flexibility, wide range, and hardware friendliness make IEM the best candidate for system-on-chip integration to our knowledge.

On-chip, time-correlated, fluorescence lifetime extraction algorithms and error analysis.

A new, simple, and hardware-only fluorescence-lifetime-imaging microscopy (FLIM) is proposed to implement on-chip lifetime extractions, and their signal-to-noise-ratio based on statistics theory is also deduced. The results are compared with Monte Carlo simulations, giving good agreement. Compared with the commonly used iterative least-squares method or the maximum-likelihood-estimation- (MLE-) based, general purpose FLIM analysis software, our algorithm offers direct calculation of fluorescence lifetime based on the collected photon counts stored in on-chip counters and therefore delivers faster analysis for real-time applications, such as clinical diagnosis. Error analysis considering timing jitter based on statistics theory is carried out for the proposed algorithms and is also compared with MLE to obtain optimized channel width or measurement window and bit resolution of the time-to-digital converters for a given accuracy. A multi-exponential, pipelined fluorescence lifetime method based on the proposed algorithms is also introduced. The performance of the proposed methods has been tested on mono-exponential and four-exponential decay experimental data.