Stabilization of zein nanoparticles with tween-80 and fucoidan for encapsulation of eugenol via a nozzle simulation chip.

Eugenol (EU), a natural bioactive compound found in various plants, offers numerous health benefits, but its application in the food and pharmaceutical industry is limited by its high volatility, instability, and low water solubility. Therefore, this study aimed to utilize the surface coating technique to develop zein-tween-80-fucoidan (Z-T-FD) composite nanoparticles for encapsulating eugenol using a nozzle simulation chip. The physicochemical characteristics of the composite nanoparticles were examined by varying the weight ratios of Z, T, and FD. Results showed that the Z-T-FD weight ratio of 5:1:15 exhibited excellent colloidal stability under a range of conditions, including pH (2-8), salt concentrations (10-500 mmol/L), heating (80 °C), and storage (30 days). Encapsulation of EU into Z-T-FD nanoparticles (0.5:5:1:15) resulted in an encapsulation efficiency of 49.29 ± 1.00%, loading capacity of 0.46 ± 0.05%, particle size of 205.01 ± 3.25 nm, PDI of 0.179 ± 0.006, and zeta-potential of 37.12 ± 1.87 mV. Spherical structures were formed through hydrophobic interaction and hydrogen bonding, as confirmed by Fourier transform infrared spectroscopy and molecular docking. Furthermore, the EU-Z-T-FD (0.5:5:1:15) nanoparticles displayed higher in vitro antioxidant properties (with DPPH and ABTS radical scavenging properties at 75.28 ± 0.16% and 39.13 ± 1.22%, respectively), in vitro bioaccessibility (64.78 ± 1.37%), and retention rates under thermal and storage conditions for EU compared to other formulations. These findings demonstrate that the Z-T-FD nanoparticle system can effectively encapsulate, protect, and deliver eugenol, making it a promising option for applications in the food and pharmaceutical industries.

Nanoencapsulation and delivery of bioactive ingredients using zein nanocarriers: approaches, characterization, applications, and perspectives.

Zein has garnered widespread attention as a versatile material for nanosized delivery systems due to its unique self-assembly properties, amphiphilicity, and biocompatibility characteristics. This review provides an overview of current approaches, characterizations, applications, and perspectives of nanoencapsulation and delivery of bioactive ingredients within zein-based nanocarriers. Various nanoencapsulation strategies for bioactive ingredients using various types of zein-based nanocarrier structures, including nanoparticles, nanofibers, nanoemulsions, and nanogels, are discussed in detail. Factors affecting the stability of zein nanocarriers and characterization methods of bioactive-loaded zein nanocarrier structures are highlighted. Additionally, current applications of zein nanocarriers loaded with bioactive ingredients are summarized. This review will serve as a guide for the selection of appropriate nanoencapsulation techniques within zein nanocarriers and a comprehensive understanding of zein-based nanocarriers for specific applications in the food, pharmaceutical, cosmetic, and agricultural industries. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-023-01489-6.

A nozzle simulation chip toward high-throughput formation of curcumin-loaded zein nanoparticles with tunable properties.

Size of nanoparticle (NP) is a crucial factor in determining its applicability to various fields. This study aimed to develop a nozzle chip for the scalable formation of self-assembled curcumin-loaded zein NPs with tunable properties. A four-factor (zein concentration in dispersed phase, ethanol concentration in continuous phase, flow rate ratio, and total flow rate), three-level Box-Behnken design on the measured responses (particle size and polydispersity index [PDI]) was established. The particle size and PDI, ranging from 194.43 to 420.51 nm, and 0.089 to 0.219, respectively, were readily controlled by adjusting four factors. Under the optimal conditions of 6% zein, 0% EtOH, the flow rate ratio of 7, and a total flow rate of 8 mL/min targeting higher production rate, the particle size of 306.02 ± 1.52 nm (mean ± standard deviation) and the PDI of 0.135 ± 0.001 were obtained. High throughput for zein NP production (86.4 g/day) was reached, which was 200 and 960 times higher than using microfluidic and electrospraying techniques, respectively. Curcumin-loaded zein NPs under the abovementioned experimental conditions were successfully prepared via the nozzle chip with the encapsulation efficiency of 64.29% ± 0.29%, a loading capacity of 3.06% ± 0.01%, enhanced stability, and improved in vitro antioxidant properties. Curcumin was primarily released from zein NPs in the simulated intestinal phase. This study demonstrated that the property of self-assembled zein NPs can be tuned by altering the operating parameters using the nozzle simulation chip. The results suggest that this approach has potential for use in the food and pharmaceutical industries, particularly for curcumin encapsulation. PRACTICAL APPLICATION: The fabricated nozzle chip is a promising technology to obtain zein nanoparticles (NPs) with enhanced productivity and narrow particle size distribution. It can be easily adopted to spray drying process. Besides, the nozzle chip shows the potential for the large-scale production of bioactive loaded zein NPs in the food or pharmaceutical industries.

Intelligent films based on pectin, sodium alginate, cellulose nanocrystals, and anthocyanins for monitoring food freshness.

To address consumer-level food waste, and pollution from commercial plastics, we developed intelligent films using sodium alginate (SA), pectin (PC), cellulose nanocrystals (CNCs), and anthocyanins extracted from red cabbage (RCA). We also investigated two methods of reinforcing these films - cross-linking (CL), and the addition of CNCs. Both together and separately, these methods improved SA/PC films' mechanical properties and thermal stability. The optimal SA/PC/CNCs/RCA/CL films exhibited pH-dependent color-response properties and high water resistance. These were then tested as colorimetric freshness indicators for shrimp samples, both through seepage and the monitoring of volatile compounds. The colors of the indicators changed from lilac to dark green to greenish-yellow after storage at 25 °C for 72 h, whereas at 4 °C, they changed much more slowly over the same time period. This demonstrated the excellent potential of such films to reduce food waste by providing real-time warnings of pH variation resulting from spoilage.

Social partnering alters sleep in fear-conditioned Wistar rats.

Social support, when provided following a traumatic experience, is associated with a lower incidence of stress-related psychiatric disorders. Our hypothesis was that providing a social interaction period with a naive conspecific would improve sleep architecture in response to cued fear conditioning in Wistar rats. Rats were randomly assigned to either the socially isolated or socially partnered groups. Rats assigned to the socially isolated group were individually housed following electrode implantation and fear conditioning. Rats assigned to the socially partnered group were initially paired-housed, and then one rat from each pair was randomly chosen for sleep electrode implantation and fear conditioning. Rats from both groups were habituated to a recording chamber, and baseline sleep was recorded over 22 hours. One day later (Training Day), they were fear-conditioned to 10 presentations of a tone (800 Hz, 90 dB, 5 sec) co-terminating with a mild electric foot shock (1.0 mA, 0.5 sec), at 30-sec intervals. While rats in the socially isolated group were left undisturbed in their home cage for 30-min, socially partnered rats interacted for 30 minutes with their non-stressed rat partner immediately after fear conditioning and while the auditory tones were presented on Days 1 and 14. The results indicated that social interaction increased sleep efficiency in partnered rats compared to isolated rats following the fear conditioning procedure. This was due to an increase in the amount of rapid eye movement sleep (REMS) during the light phase. Evaluation of REMS microarchitecture revealed that the increase in REMS was due to an increase in the number of single REMS episodes (siREMS), which represented a more consolidated REMS pattern. A surprising finding was that partnered rats had a greater number of sequential REMS episodes (seqREMS) at Baseline, on the Training Day and on Day 1 when compared to isolated rats. The greater number of seqREMS episodes in partnered rats may be due to the partnering procedure and not fear conditioning, as the effect was also seen at Baseline. Thus it appears that while the partnering procedure may have given rise to a fragmented REMS pattern, social partnering promoted a greater consolidation of REMS in response to the fear conditioning procedure.

Adolescent social defeat alters N-methyl-D-aspartic acid receptor expression and impairs fear learning in adulthood.

Repeated social defeat of adolescent male rats results in adult mesocortical dopamine hypofunction, impaired working memory, and increased contextual anxiety-like behavior. Given the role of glutamate in dopamine regulation, cognition, and fear and anxiety, we investigated potential changes to N-methyl-D-aspartic acid (NMDA) receptors following adolescent social defeat. As both NMDA receptors and mesocortical dopamine are implicated in the expression and extinction of conditioned fear, a separate cohort of rats was challenged with a classical fear conditioning paradigm to investigate whether fear learning is altered by adolescent defeat. Quantitative autoradiography was used to measure 3H-MK-801 binding to NMDA receptors in regions of the medial prefrontal cortex, caudate putamen, nucleus accumbens, amygdala and hippocampus. Assessment of fear learning was achieved using an auditory fear conditioning paradigm, with freezing toward the auditory tone used as a measure of conditioned fear. Compared to controls, adolescent social defeat decreased adult NMDA receptor expression in the infralimbic region of the prefrontal cortex and central amygdala, while increasing expression in the CA3 region of the hippocampus. Previously defeated rats also displayed decreased conditioned freezing during the recall and first extinction periods, which may be related to the observed decreases and increases in NMDA receptors within the central amygdala and CA3, respectively. The alteration in NMDA receptors seen following adolescent social defeat suggests that dysfunction of glutamatergic systems, combined with mesocortical dopamine deficits, likely plays a role in the some of the long-term behavioral consequences of social stressors in adolescence seen in both preclinical and clinical studies.

Respiratory modulation of lingual muscle activity across sleep-wake states in rats.

In obstructive sleep apnea (OSA) patients, inspiratory activation (IA) of lingual muscles protects the upper airway from collapse. We aimed to determine when rats' lingual muscles exhibit IA. In 5 Sprague-Dawley and 3 Wistar rats, we monitored cortical EEG and lingual, diaphragmatic and nuchal electromyograms (EMGs), and identified segments of records when lingual EMG exhibited IA. Individual segments lasted 2.4-269 s (median: 14.5 s), most (89%) occurred during slow-wave sleep (SWS), and they collectively occupied 0.3-6.1% of the total recording time. IA usually started to increase with a delay after SWS onset and ended with an arousal, or declined prior to rapid eye movement sleep. IA of lingual EMG was not accompanied by increased diaphragmatic activity or respiratory rate changes, but occurred when cortical EEG power was particularly low in a low beta-1 frequency range (12.5-16.4 Hz). A deep SWS-related activation of upper airway muscles may be an endogenous phenomenon designed to protect the upper airway against collapse.

Evidence that adrenergic ventrolateral medullary cells are activated whereas precerebellar lateral reticular nucleus neurons are suppressed during REM sleep.

Rapid eye movement sleep (REMS) is generated in the brainstem by a distributed network of neurochemically distinct neurons. In the pons, the main subtypes are cholinergic and glutamatergic REMS-on cells and aminergic REMS-off cells. Pontine REMS-on cells send axons to the ventrolateral medulla (VLM), but little is known about REMS-related activity of VLM cells. In urethane-anesthetized rats, dorsomedial pontine injections of carbachol trigger REMS-like episodes that include cortical and hippocampal activation and suppression of motoneuronal activity; the episodes last 4-8 min and can be elicited repeatedly. We used this model to determine whether VLM catecholaminergic cells are silenced during REMS, as is typical of most aminergic neurons studied to date, and to investigate other REMS-related cells in this region. In 18 anesthetized, paralyzed and artificially ventilated rats, we obtained extracellular recordings from VLM cells when REMS-like episodes were elicited by pontine carbachol injections (10 mM, 10 nl). One major group were the cells that were activated during the episodes (n = 10). Their baseline firing rate of 3.7±2.1 (SD) Hz increased to 9.7±2.1 Hz. Most were found in the adrenergic C1 region and at sites located less than 50 µm from dopamine ß-hydroxylase-positive (DBH(+)) neurons. Another major group were the silenced or suppressed cells (n = 35). Most were localized in the lateral reticular nucleus (LRN) and distantly from any DBH(+) cells. Their baseline firing rates were 6.8±4.4 Hz and 15.8±7.1 Hz, respectively, with the activity of the latter reduced to 7.4±3.8 Hz. We conclude that, in contrast to the pontine noradrenergic cells that are silenced during REMS, medullary adrenergic C1 neurons, many of which drive the sympathetic output, are activated. Our data also show that afferent input transmitted to the cerebellum through the LRN is attenuated during REMS. This may distort the spatial representation of body position during REMS.

Fear conditioning fragments REM sleep in stress-sensitive Wistar-Kyoto, but not Wistar, rats.

Pavlovian conditioning is commonly used to investigate the mechanisms of fear learning. Because the Wistar-Kyoto (WKY) rat strain is particularly stress-sensitive, we investigated the effects of a psychological stressor on sleep in WKY compared to Wistar (WIS) rats. Male WKY and WIS rats were either fear-conditioned to tone cues or received electric foot shocks alone. In the fear-conditioning procedure, animals were exposed to 10 tones (800 Hz, 90 dB, 5s), each co-terminating with a foot shock (1.0 mA, 0.5s), at 30-s intervals. In the shock stress procedure, animals received 10 foot shocks at 30-s intervals, without tones. All subjects underwent a tone-only test both 24h (Day 1) and again two weeks (Day 14) later. Rapid eye movement sleep (REMS) continuity was investigated by partitioning REMS episodes into single (inter-REMS episode interval >3 min) and sequential (interval ≤ 3 min) episodes. In the fear-conditioned group, freezing increased from baseline in both strains, but the increase was maintained on Day 14 in WKY rats only. In fear-conditioned WKY rats, total REMS amount increased on Day 1, sequential REMS amount increased on Day 1 and Day 14, and single REMS amount decreased on Day 14. Alterations were due to changes in the number of sequential and single REMS episodes. Shock stress had no significant effect on REMS microarchitecture in either strain. The shift toward sequential REMS in fear-conditioned WKY rats may represent REMS fragmentation, and may provide a model for investigating the neurobiological mechanisms of sleep disturbances reported in posttraumatic stress disorder.

Strain differences in the distribution of N-methyl-d-aspartate and gamma (gamma)-aminobutyric acid-A receptors in rat brain.

AIMS: Previous studies have shown that the Wistar-Kyoto (WKY) rat strain exhibits depressive symptoms such as anhedonia, psychomotor retardation, ambivalence and negative memory bias following exposure to stress. Given the involvement of excitatory glutamate and inhibitory gamma (gamma)-aminobutyric acid (GABA) signaling pathways in influencing depressive behavior, the present study investigated strain differences in the distribution of central N-methyl-d-aspartate (NMDA) and GABA(A) receptor sites in WKY compared to their inbred counterpart, Wistar (WIS) rats. MAIN METHODS: Quantitative autoradiographic analysis was used to map the binding and distribution of NMDA and GABA(A) receptors in various brain regions in WKY and WIS rats. KEY FINDINGS: Results indicated a significant difference between the two strains. Lower NMDA receptor binding was found in the anterior cingulate cortex, caudate putmen, nucleus accumbens, CA1 region of the hippocampus and the substantia nigra pars reticulata in WKY compared to WIS rats. Conversely, higher GABA(A) receptor binding was found in the amygdala, caudate putmen, dentate gyrus, CA2 and CA3 fields of the hippocampus, periaqueductal grey and substantia nigra pars reticulata in WKY compared to WIS rats. SIGNIFICANCE: Given that these two rat strains differ in their behavioural, endocrine and neurochemical profile, the observed strain differences in NMDA and GABA(A) receptor binding suggest that these two neurotransmitter systems may be involved in the depressive and stress-sensitive phenotype of the WKY rat strain.

Long-term exposure to extremely low-frequency magnetic fields impairs spatial recognition memory in mice.

1. In the present study, we investigated the short- and long-term effects of extremely low-frequency (ELF) magnetic fields on spatial recognition memory in mice by using a two-trial recognition Y-maze that is based on the innate tendency of rodents to explore novel environments. 2. Mice were exposed to 25 or 50 Hz electromagnetic fields for either 7 (short term) or 25 days (long term) and then tested in the Y-maze. 3. The results indicated that neither short- nor long-term exposure to magnetic fields affected the locomotor activity of mice in the Y-maze. However, long-term exposure to 50 Hz fields reduced recognition of the novel arm. 4. Our findings suggest that ELF magnetic fields impair spatial recognition memory in the Y-maze depending on the field strength and/or duration of exposure.

Extremely low-frequency electromagnetic field exposure during chronic morphine treatment strengthens downregulation of dopamine D2 receptors in rat dorsal hippocampus after morphine withdrawal.

The aim of this study was to investigate the effect of extremely low-frequency electromagnetic field (ELF-EMF) exposure during morphine treatment on dopamine D2 receptor (D2R) density in the rat dorsal hippocampus following withdrawal. Rats were exposed to ELF-EMF (20 Hz, 14 mT) or sham exposed for 1h per day before injection of morphine (10mg/kg, i.p.) once daily for 12 days. The saline control group was sham exposed for the same period. Immunohistochemistry was used to detect the density of D2Rs on the 1st, 3rd and 5th morphine withdrawal days. The results showed that the density of D2Rs in sham-exposed morphine-treated rats on the 1st and 3rd days of morphine withdrawal was significantly lower than that of the saline control group. The ELF-EMF-exposed morphine group also exhibited a significantly lower density of D2Rs on the 1st and 3rd withdrawal days relative to the sham-exposed morphine group. However, the D2R density in both groups tended to recover as morphine withdrawal days increased. The results suggest that dorsal hippocampal D2Rs are sensitive to morphine withdrawal and that this is potentiated by ELF-EMF pre-exposure during morphine treatment.

Effects of extremely low-frequency electromagnetic fields on morphine-induced conditioned place preferences in rats.

In the present study, we examined the effects of extremely low-frequency (ELF) electromagnetic fields on morphine-induced conditioned place preferences in rats. During the conditioning phase (12 days), three groups of rats were placed in a sensory cue-defined environment paired with morphine (10mg/kg, i.p.) following exposure to either 20 Hz (1.80 mT) or 50 Hz (2.20 mT) or sham electromagnetic fields for 60 min/day, respectively, and were placed in another sensory cue-defined environment paired with physiological saline (1 ml/kg, i.p.) without exposure to electromagnetic fields. After finishing 12 days of conditioning, preference tests for the morphine-paired place were performed during a 10-day withdrawal period. The exposure to electromagnetic fields substantially potentiated morphine-induced place preferences in rodents, suggesting that ELF electromagnetic fields can increase the propensity for morphine-induced conditioned behaviors.

[Methods for single unit recording in behavioring morphine craving rat].

In this paper, one method was introduced, which was a combination of the cue-related morphine addiction model and a technique for obtaining chronic extracellular recordings of single unit in freely moving rats. With the combination and improvement of this technique, we have successfully applied this new method to study the neuronal activity of the hippocampus CA1 region in morphine withdrawal rats. In all, we found some more accurate and objective cellular characteristics of hippocampal neurons, and considered these characteristics as one of electrophysiological indexes of morphine addiction rats.

Telemetric recordings of single neuron activity and visual scenes in monkeys walking in an open field.

This paper describes a portable recording system and methods for obtaining chronic recordings of single units and tracking rhesus monkey behavior in an open field. The integrated system consists of four major components: (1) microelectrode assembly; (2) head-stage; (3) recording station; and (4) data storage station, the first three of which are carried by the monkey and weigh 800 g. Our system provides synchronized video and electrophysiological signals, which are transmitted by a wireless system to a distance of 50 m. Its major advantages are that neuronal recordings are made in freely moving monkeys, and well-separated action potentials with amplitude five times higher than the background noise are usually recorded and readily kept for many hours. Using this system, we were able to study "place cells" in non-human primate brains. The described methods provide a new way to examine correlations between single neuron activity and primate behaviors, and can also be used to study the cellular basis of social behaviors in non-human primates.