Recent advances in microbial synthesis of free heme.

Heme is an iron-containing porphyrin compound widely used in the fields of healthcare, food, and medicine. Compared to animal blood extraction, it is more advantageous to develop a microbial cell factory to produce heme. However, heme biosynthesis in microorganisms is tightly regulated, and its accumulation is highly cytotoxic. The current review describes the biosynthetic pathway of free heme, its fermentation production using different engineered bacteria constructed by metabolic engineering, and strategies for further improving heme synthesis. Heme synthetic pathway in Bacillus subtilis was modified utilizing genome-editing technology, resulting in significantly improved heme synthesis and secretion abilities. This technique avoided the use of multiple antibiotics and enhanced the genetic stability of strain. Hence, engineered B. subtilis could be an attractive cell factory for heme production. Further studies should be performed to enhance the expression of heme synthetic module and optimize the expression of heme exporter and fermentation processes, such as iron supply. KEY POINTS: • Strengthening the heme biosynthetic pathway can significantly increase heme production. • Heme exporter overexpression helps to promote heme secretion, thereby further promoting excessive heme synthesis. • Engineered B. subtilis is an attractive alternative for heme production.

Biportal Endoscopic Spinal Cord Stimulation Paddle Lead Implantation: Technical Note and Preliminary Clinical Results.

OBJECTIVE: This study aims to elucidate a novel, minimally invasive surgical technique using a biportal endoscope for the implantation of spinal cord stimulation (SCS) paddle leads and to report the preliminary results of its clinical application. MATERIALS AND METHODS: The perioperative data of patients who underwent the biportal endoscopic SCS paddle lead implantation in our department were collected; the surgical procedure was delineated, and the clinical outcomes were assessed. RESULTS: From February 2022 to December 2023, six patients underwent biportal endoscopic SCS paddle lead implantation. The median follow-up time was nine months (range one to three months). The median intraoperative blood loss was 30 mL (range 25-50 mL), and the median operative time was 87.5 minutes (range 75-110 minutes). One patient experienced severe neck pain during the operation, whereas the other five patients experienced no surgical complications. One patient was found to have a slight lead migration three months after surgery, which did not affect the therapeutic effect. The median visual analogue scale (VAS) of the surgical area was 0.5 (range 0-2), 2.5 (range 1-4), and 0.5 (range 0-1) during the operation and one day and one week after the operation, respectively. The median VAS of the six patients' primary disease was 8 (range 7-9) before surgery and 2.5 (range 1-4) at the last postoperative follow-up (pain reduction ≥50%). CONCLUSION: Paddle lead systems for SCS can be implanted successfully using a biportal endoscopic technique.

Screening and identification of genes involved in ß-alanine biosynthesis in Bacillus subtilis.

ß-alanine is the only naturally occurring ß-amino acid, which is widely used in medicine, food, and feed fields, and generally produced through synthetic biological methods based on engineered strains of Escherichia coli or Corynebacterium glutamicum. However, the ß-alanine biosynthesis in Bacillus subtilis, a traditional industrial model microorganism of food safety grade, has not been thoroughly explored. In this study, the native l-aspartate-α-decarboxylase was overexpressed in B. subtilis 168 to obtain an increase of 842% in ß-alanine production. A total of 16 single-gene knockout strains were constructed to block the competitive consumption pathways to identify a total of 6 genes (i.e., ptsG, fbp, ydaP, yhfS, mmgA, and pckA) involved in ß-alanine synthesis, while the multigene knockout of these 6 genes obtained an increased ß-alanine production by 40.1%. Ten single-gene suppression strains with the competitive metabolic pathways inhibited revealed that the inhibited expressions of genes glmS, accB, and accA enhanced the ß-alanine production. The introduction of heterologous phosphoenolpyruvate carboxylase increased the ß-alanine production by 81.7%, which was 17-fold higher than that of the original strain. This was the first study using multiple molecular strategies to investigate the biosynthetic pathway of ß-alanine in B. subtilis and to identify the genetic factors limiting the excessive synthesis of ß-alanine by microorganisms.

Flavihumibacter fluminis sp. nov., a novel thermotolerant bacterium isolated from river silt.

A yellow, Gram-stain-positive, strictly aerobic, thermotolerant, non-motile and rod-shaped bacterial strain, designated RY-1T, was isolated from a silt sample of Fuyang River, Wuqiang County, Hengshui City, Hebei Province, PR China. Cells showed oxidase- and catalase-positive activities. Growth occurred at 20-45 °C (optimum, 37 °C) and pH 6.0-8.0 (optimum, pH 7.0), and in the presence of 0-1.5 % (w/v) NaCl (optimum, 0%). A phylogenetic tree based on 16S rRNA gene sequences revealed that strain RY-1T formed a phylogenetic lineage with Flavihumibacter members within the family Chitinophagaceae. A comparison of 16S rRNA gene sequences showed that strain RY-1T was most closely related to Flavihumibacter cheonanensis WS16T (98.6 %), Flavihumibacter sediminis CJ663T (97.7 %) and Flavihumibacter solisilvae 3-3T (97.6 %). The genome size of strain RY-1T was 4.71 Mb, and the DNA G+C content was 44.3  %. The average nucleotide identity, digital DNA-DNA hybridization and average amino acid identity values between strain RY-1T and reference strains were all lower than the threshold values for species delineation. Strain RY-1T contained menaquinone-7 and iso-C15 : 0, iso-C17 : 0 3-OH and iso-C15 : 1G as the sole respiratory isoprenoid quinone and major cellular fatty acids (≥5 %), respectively. The major polar lipids consisted of phosphatidylethanolamine, three unidentified aminolipids and four unidentified lipids. According to the results of phenotypic, phylogenetic and chemotaxonomic characteristics, strain RY-1T represents a novel species of the genus Flavihumibacter, for which the name Flavihumibacter fluminis sp. nov. is proposed. The type strain is RY-1T (=GDMCC 1.2775T=JCM 34870T).

Improved biosynthesis of heme in Bacillus subtilis through metabolic engineering assisted fed-batch fermentation.

BACKGROUND: Heme is an iron/porphyrin complex compound, widely used in the health care, food, and pharmaceutical industries. It is more advantageous and attractive to develop microbial cell factories to produce heme by fermentation, with lower production costs and environmentally more friendly procedures than those of the traditional extraction based on animal blood. In this study, Bacillus subtilis, a typical industrial model microorganism of food safety grade, was used for the first time as the host to synthesize heme. RESULTS: The heme biosynthetic pathway was engineered as four modules, the endogenous C5 pathway, the heterologous C4 pathway, the uroporphyrinogen (urogen) III synthesis pathway, and the downstream synthesis pathway. Knockout of hemX encoding the negative effector of the concentration of HemA, overexpression of hemA encoding glutamyl-tRNA reductase, and knockout of rocG encoding the major glutamate dehydrogenase in the C5 pathway, resulted in an increase of 427% in heme production. Introduction of the heterologous C4 pathway showed a negligible effect on heme biosynthesis. Overexpression of hemCDB, which encoded hydroxymethylbilane synthase, urogen III synthase, and porphobilinogen synthase participating in the urogen III synthesis pathway, increased heme production by 39%. Knockouts of uroporphyrinogen methyltransferase gene nasF and both heme monooxygenase genes hmoA and hmoB in the downstream synthesis pathway increased heme production by 52%. The engineered B. subtilis produced 248.26 ± 6.97 mg/L of total heme with 221.83 ± 4.71 mg/L of extracellular heme during the fed-batch fermentation in 10 L fermenter. CONCLUSIONS: Strengthening endogenous C5 pathway, urogen III synthesis pathway and downstream synthesis pathway promoted the biosynthesis of heme in B. subtilis. The engineered B. subtilis strain has great potential as a microbial cell factory for efficient industrial heme production.

Pharmacodynamic comparison of acarbose tablets in Chinese healthy volunteers under chewing and swallowing conditions.

WHAT IS KNOWN AND OBJECTIVE: Acarbose can efficiently block glucose absorption in the intestine as an alpha-glucosidase inhibitor. It is currently manufactured in several oral dosage forms, with the most common types being tablets and chewable tablets. The acarbose tablet (Glucobay® , 50 mg, Bayer) package insert gives instructions for either directly swallowing or chewing then swallowing. This study compared the pharmacodynamic effects of a single formulation of acarbose tablets under these two different administration routes. METHODS: This randomized, crossover study enrolled 24 healthy subjects who were instructed to chew (C group) or swallow (S group) the acarbose tablet. Glucose levels were monitored in subjects for up to 4 h following administration of 75 g of sucrose to establish a baseline firstly, after which subjects in the C and S groups were administered 50- or 100- mg of acarbose along with 75 g of sucrose. Then, subjects entered a 1-week washout period before being crossed over to the alternate dosing route. RESULTS AND DISCUSSION: Compared with the S group, the C group had a lower maximum concentration of serum glucose (Cmax ) and areas under the concentration-time curve (AUC0-2 , AUC0-1.5 ). In addition, the maximum reduction in serum glucose (ΔCmax ) and the reduction in the AUC (AUEC0-1.5 ) were both increased in the S group. This occurred at both the 50 mg and 100 mg dosages. These results indicate that fluctuations in blood glucose were lower following chewing of the acarbose tablet. Both administration routes exhibited similar safety and tolerance profiles. WHAT IS NEW AND CONCLUSION: In summary, chewing acarbose tablets appears to induce a superior glycaemic-controlling effect compared with swallowing them directly, at least with a single dose. It will be important to inform both clinicians and patients about these differences between the two administrations so that informed clinical decisions can be made, as numerous patients with diabetes are inclined to directly swallow acarbose tablets for convenience.

Pharmacokinetic comparison of nalbuphine with single injection and patient-controlled analgesia mimic method in healthy Chinese volunteers.

WHAT IS KNOWN AND OBJECTIVE: Nalbuphine is a mu (µ) receptor partial antagonist/kappa (κ) receptor agonist analgesic and can be administered as a single injection or using patient-controlled analgesia (PCA) in the clinical setting. However, differences in the pharmacokinetics of the two administration methods are unclear. Here, a clinical trial was performed to compare the pharmacokinetic characteristics and superiority of nalbuphine with a single-injection or PCA-mimic method to provide a reference for the selection of an appropriate administration method. METHODS: Twenty healthy individuals were divided into two groups and injected with 10 mg nalbuphine intravenously using a single-injection or a PCA-mimic method (2 mg once for five times with a 30-min interval). Blood samples were collected, and safety was investigated. The liquid chromatography-tandem mass spectrometry was adopted to determine the concentration of nalbuphine in plasma. RESULTS AND DISCUSSION: The maximum concentration (Cmax ) and area under concentration-time curve (AUC0-t ) values of nalbuphine in the single-injection and PCA groups were as follows: Cmax , 81.3 ± 24.7 and 39.8 ± 6.4 ng/ml, respectively; moreover, AUC0-t , 110.3 ± 19.5 and 128.3 ± 23.0 h ng/ml, respectively. The effective analgesic concentration durations (EACDs) for the two administration methods were 1.39 ± 0.64 and 1.96 ± 0.91 h, respectively. Nalbuphine was well tolerated, and improvements were observed in the PCA group. WHAT IS NEW AND CONCLUSION: Compared with those in the single-injection group, the AUC0-t and EACDs in the PCA group were similar, whereas Cmax was decreased significantly. Therefore, the PCA method was more suitable for the clinical application of nalbuphine injection owing to the superiority of lower concentration fluctuation and the improved safety profile.

Pharmacokinetics and tolerability of intravenous posaconazole in healthy Chinese volunteers: a randomized, open-label and single-dose study.

Posaconazole is a triazole antifungal drug with strong antifungal effect. The pharmacokinetics, safety, and tolerability were evaluated following the intravenous administration of posaconazole injection. A total of 36 healthy adults were enrolled in the parallel-designed clinical trial, and the subjects received single doses of posaconazole injection (100, 200 and 300 mg). Posaconazole concentrations in plasma were determined with liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. The levels of posaconazole in plasma increased proportionally between 100 and 300-mg dose, but AUC showed a more-than-dose-proportional increase. Besides, decreased Vd and CL were observed, along with the increased posaconazole dosage. Posaconazole was well tolerated at all dose levels, and the adverse events were not dose dependent. No clinically significant changes in electrocardiograms were observed.

Pharmacokinetic profile of ivabradine hemisulfate sustained-release tablets administered in Chinese healthy volunteers: An open-label, randomized, single-dose, three-period crossover study.

We aimed to determine the pharmacokinetics and safety of three single oral doses (5, 10 and 15 mg) of ivabradine hemisulfate sustained-release tablets in healthy Chinese volunteers. A total of 12 volunteers (six males and six females) were randomized to receive a single oral dose of ivabradine hemisulfate sustained-release tablets 5, 10 or 15 mg, with a 1-week washout between periods. Blood samples were collected at regular intervals from 0 to 48 h after drug administration, and the concentrations of ivabradine and N-desmethyl ivabradine were determined by HPLC-tandem mass spectrometry. Pharmacokinetic parameters were estimated by non-compartmental analysis. After administering single doses of 5, 10 and 15 mg, the mean maximum concentration (Cmax ) levels of ivabradine were 4.36, 7.29 and 12.62 ng/mL, and the mean area under the curve from time 0 to 48 h (AUC0-48 ) values were 55.66, 101.16 and 182.09 h·ng/mL, respectively. The mean Cmax levels of N-desmethyl ivabradine were 1.05, 2.03 and 3.16 ng/mL, and the mean AUC0-48 values were 20.61, 39.44 and 65.72 h·ng/mL, respectively. The median time of maximum concentration (Tmax ) levels of ivabradine and N-desmethyl ivabradine were 5 h for all three doses tested. The pharmacokinetic properties of ivabradine hemisulfate sustained-release tablets were linear at doses from 5 to 15 mg. Ivabradine hemisulfate sustained-release tablet appears to be well tolerated in these healthy volunteers.

[Effect of key-gene modification on uridine biosynthesis in Bacillus subtilis].

OBJECTIVE: We studied several crucial factors influencing the uridine biosynthesis in Bacillus subtilis, including mutations of phosphoribosylpyrophosphate synthetase (PRPP synthetase) (prs) and carbamyl phosphate synthetase (pyrAA/pyrAB), and overexpression of heterologous 5'-nucleotidase (sdt1). METHODS: According to the inferred allosteric sites, we introduced point mutation into coding sequences of prs and pyrAB. The mutated prs gene was integratedly expressed in the xylR locus of the chromosome and the pyrAB gene was modified in-situ. The sdt1 gene was overexpressed in the saB locus of the chromosome. The effect of the genetic modification on uridine biosynthesis was characterized by the analysis of uridine, cytidine and uracil in the fermentation broth. RESULTS: The mutations of Asn120Ser, Leu135Ile, Glu52Gly or Val312Ala on PRPP synthase resulted in an increase of uridine production by 67% and 96%, respectively. The mutations of Ser948Phe, Thr977Ala and Lys993Ile on carbamyl phosphate synthase resulted in a 182% increase of uridine yield to 6.97 g/L. The overexpression of heterologous 5'-nucleotidase resulted in a 17% increase of uridine yield to 8.16 g/L. CONCLUSION: The activity and regulation mechanism of PRPP synthase and carbamyl phosphate synthase was an important factor to limit the excessive synthesis of uridine. Asn120Ser and Leu135Ile mutations of PRPP synthase and Ser948Phe, Thr977Ala and Lys993Ile mutations of carbamyl phosphate synthase will facilitate the biosynthesis of uridine. The additional Glu52Gly and Val312Ala mutations of PRPP synthase were beneficial for uridine biosynthesis. The reaction from UMP to uridine also limited the biosynthesis of uridine in B. subtilis.

Metabolic and genetic factors affecting the productivity of pyrimidine nucleoside in Bacillus subtilis.

BACKGROUND: Cytidine and uridine are produced commercially by Bacillus subtilis. The production strains of cytidine and uridine were both derivatives from mutagenesis. However, the exact metabolic and genetic factors affecting the productivity remain unknown. Genetic engineering may be a promising approach to identify and confirm these factors. RESULTS: With the deletion of the cdd and hom genes, and the deregulation of the pyr operon in Bacillus subtilis168, the engineered strain produced 200.9 mg/L cytidine, 14.9 mg/L uridine and 960.1 mg/L uracil. Then, the overexpressed prs gene led to a dramatic increase of uridine by 25.9 times along with a modest increase of cytidine. Furthermore, the overexpressed pyrG gene improved the production of cytidine, uridine and uracil by 259.5%, 11.2% and 68.8%, respectively. Moreover, the overexpression of the pyrH gene increasesd the yield of cytidine by 40%, along with a modest augments of uridine and uracil. Lastly, the deletion of the nupC-pdp gene resulted in a doubled production of uridine up to 1684.6 mg/L, a 14.4% increase of cytidine to 1423 mg/L, and a 99% decrease of uracil to only 14.2 mg/L. CONCLUSIONS: The deregulation of the pyr operon and the overexpression of the prs, pyrG and pyrH genes all contribute to the accumulation of pyrimidine nucleoside compounds in the medium. Among these factors, the overexpression of the pyrG and pyrH genes can particularly facilitate the production of cytidine. Meanwhile, the deletion of the nupC-pdp gene can obviously reduce the production of uracil and simultaneously improve the production of uridine.

Synergistic enhancement of cancer therapy using a combination of ceramide and docetaxel.

Ceramide (CE)-based combination therapy (CE combination) as a novel therapeutic strategy has attracted great attention in the field of anti-cancer therapy. The principal purposes of this study were to investigate the synergistic effect of CE in combination with docetaxel (DTX) (CE + DTX) and to explore the synergy mechanisms of CE + DTX. The 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and combination index (CI) assay showed that simultaneous administration of CE and DTX with a molar ratio of 0.5:1 could generate the optimal synergistic effect on murine malignant melanoma cell (B16, CI = 0.31) and human breast carcinoma cell (MCF-7, CI = 0.48). The apoptosis, cell cycle, and cytoskeleton destruction study demonstrated that CE could target and destruct the microfilament actin, subsequently activate Caspase-3 and induce apoptosis. Meanwhile, DTX could target and disrupt the microtubules cytoskeleton, leading to a high proportion of cancer cells in G2/M-phase arrest. Moreover, CE plus DTX could cause a synergistic destruction of cytoskeleton, which resulted in a significantly higher apoptosis and a significantly higher arrest in G2/M arrest comparing with either agent alone (p < 0.01). The in vivo antitumor study evaluated in B16 tumor-bearing mice also validated the synergistic effects. All these results suggested that CE could enhance the antitumor activity of DTX in a synergistic manner, which suggest promising application prospects of CE + DTX combination treatment.

Improving Surfactin Production in Bacillus subtilis 168 by Metabolic Engineering.

Surfactin is widely used in the petroleum extraction, cosmetics, biopharmaceuticals and agriculture industries. It possesses antibacterial and antiviral activities and can reduce interfacial tension. Bacillus are commonly used as production chassis, but wild-type Bacillus subtilis 168 cannot synthesise surfactin. In this study, the phosphopantetheinyl transferase (PPTase) gene sfp* (with a T base removed) was overexpressed and enzyme activity was restored, enabling B. subtilis 168 to synthesise surfactin with a yield of 747.5 ± 6.5 mg/L. Knocking out ppsD and yvkC did not enhance surfactin synthesis. Overexpression of predicted surfactin transporter gene yfiS increased its titre to 1060.7 ± 89.4 mg/L, while overexpression of yerP, ycxA and ycxA-efp had little or negative effects on surfactin synthesis, suggesting YfiS is involved in surfactin efflux. By replacing the native promoter of the srfA operon encoding surfactin synthase with three promoters, surfactin synthesis was significantly reduced. However, knockout of the global transcriptional regulator gene codY enhanced the surfactin titre to 1601.8 ± 91.9 mg/L. The highest surfactin titre reached 3.89 ± 0.07 g/L, with the yield of 0.63 ± 0.02 g/g DCW, after 36 h of fed-batch fermentation in 5 L fermenter. This study provides a reference for further understanding surfactin synthesis and constructing microbial cell factories.

Nitrate budget of a terrestrial-to-marine continuum in South China: Insights from isotopes and a Markov chain Monte Carlo model.

Anthropogenic nitrate (NO3-) production has been increasing and is exported to the ocean via river networks, causing eutrophication and ecological damage. While studies have focused on river NO3- pollution, what has been lacking is the quantification of the sources of NO3- in coastal rivers. This study applied the dual isotopes (δ15N/δ18O-NO3-) to quantify the sources and their fluxes of NO3- in two inflow rivers of the Qinzhou Bay. By adding our results to the NO3- source apportionment in Qinzhou Bay, we, for the first time, established the NO3- budgets of the terrestrial-to-marine continuum in both high- and low-flow seasons. We quantitatively showed the direct and indirect roles (e.g., the stimulation of nitrification by sewage ammonium-NH4+) of terrestrial sources in driving the high NO3- loading in the estuary. The results highlighted the necessity to consider coastal rivers and estuary as a whole, which could shed light on the effective reduction of NO3- pollution in coastal environments.

Understanding the variable metal concentrations in estuarine oysters Crassostrea hongkongensis: A biokinetic analysis.

Understanding the metal concentrations in oysters is important because of its relevance to human health and biomonitoring. However, metal concentrations in oysters are highly variable in nature and not well explained by metal exposure. This study examined the metal contamination in farm oysters Crassostrea hongkongensis grown in Qinzhou Bay, south China. Cadmium (Cd), zinc (Zn), nickel (Ni), and copper (Cu) concentrations in the oysters varied between 7.9 and 72.2, 282-17003, 0.37-47.7 and 37-4012 µg g-1, respectively, showing large metal variability among different individuals. Oyster metal concentrations decreased with increasing body size and significantly higher levels were observed in wet season. Low salinity and slower oyster growth due to inferior growth conditions could be responsible for the elevated metal concentrations in the wet season. Biokinetic modeling showed that the coupling of ingestion rate and growth can cause 2.8-4.2 folds differences in the oyster Cd and Zn concentrations, respectively, suggesting the significant role of oyster bioenergetics in contributing to the metal variability. Modeling data revealed that Cd and Zn concentrations in oyster tissues reach maximum levels when oysters have their lowest growth efficiency. This suggests that any factors influencing the energy budget in oysters could simultaneously alter their metal concentrations, which might be the reason why oyster metal concentrations are so variable in the natural environment.

A novel model based on CEEMDAN, IWOA, and LSTM for ultra-short-term wind power forecasting.

The randomness and instability of wind power bring challenges to power grid dispatching. Accurate prediction of wind power is significant to ensure the stable development of power grid. In this paper, a new ultra-short-term wind power forecasting model based on complete ensemble empirical mode decomposition with adaptive noise (CEEMDAN) and long-short-term memory (LSTM) network optimized by improved whale optimization algorithm (IWOA) is proposed. Firstly, CEEMDAN is applied to decompose the power history data into several intrinsic mode functions (IMFs) and a residual (RS) to reduce the complexity and unsteadiness of the original data. Then the partial autocorrelation method is used to analyze and select the input variables of each IMF and the residual. Finally, the IWOA-LSTM prediction model is established, and the parameters of LSTM are optimized by using the improved whale optimization algorithm. Each IMF and the residual are predicted respectively. The prediction results are superimposed to obtain the final wind power prediction value. The hybrid model is applied to the ultra-short-term wind power prediction of a wind farm in northern China. The prediction results of comparison experiments with other 11 models prove the effectiveness of the proposed model.

A novel PM2.5 concentrations probability density prediction model combines the least absolute shrinkage and selection operator with quantile regression.

PM2.5 has a significant negative impact on human health and atmospheric quality, and accurate prediction of its concentration is necessary. When using common point prediction models for PM2.5 concentration prediction, the influence of various uncertainties on PM2.5 concentrations makes the prediction results suffer from poor accuracy. To address this issue, this paper proposes the quantile regression neural network (QRNN) model based on the least absolute shrinkage and selection operator (LASSO), combined with kernel density estimation (KDE) for probabilistic density prediction of PM2.5 concentrations. The model uses LASSO regression to select the influential factors, and then the quartiles of daily PM2.5 concentrations calculated by the QRNN model are imported into the KDE model to obtain the probability density predictions of PM2.5 concentrations. In the paper, empirical analyses are carried out with the cities of Beijing and Jinan in China as well as six other datasets, and the prediction performance of the model is assessed by using evaluation criteria in both point prediction and interval prediction. The simulation reveals that the predictive performance of the LASSO-QRNN-KDE model is well, and the model is not only effective in filtering high-dimensional data, but also has a higher accuracy compared to common research models. In addition, the model is able to describe the uncertainty of PM2.5 concentration fluctuations and carry more information on the variation of PM2.5 concentrations, which can provide a novel and excellent PM2.5 concentration prediction tool for relevant policy makers.

Carbon price forecasting based on modified ensemble empirical mode decomposition and long short-term memory optimized by improved whale optimization algorithm.

The accurate prediction of carbon prices poses a tremendous challenge to relevant industry practitioners and governments. This paper proposes a novel hybrid model incorporating modified ensemble empirical mode decomposition (MEEMD) and long short-term memory (LSTM) optimized by the improved whale optimization algorithm (IWOA). This model is based on the nonlinear and non-stationary characteristics of carbon price. The original carbon price is first decomposed into nine intrinsic mode functions (IMFs) and a residual using the MEEMD model. Then, the random forest method is applied to determine the input variables of each IMF and the residual, in the LSTM neural network. The carbon price is then predicted by the LSTM model optimized by the IWOA. The proposed hybrid model is applied to predict the carbon prices of Beijing, Fujian, and Shanghai to assess its effectiveness. The results reveal that the model achieved higher prediction performance than 11 other benchmark models. Our observations indicate that decomposition of carbon price can effectively improve the accuracy of prediction. Moreover, the improved LSTM model is more suitable for time series prediction. The proposed model provides a novel and effective carbon price forecasting tool for governments and enterprises.

Modular Pathway Engineering of Bacillus subtilis To Promote De Novo Biosynthesis of Menaquinone-7.

Menaquinone-7 (MK-7), a valuable vitamin K2, plays an important role in the prevention of osteoporosis and cardiovascular calcification. We chose B. subtilis 168 as the chassis for the modular metabolic engineering design to promote the biosynthesis of MK-7. The biosynthetic pathway of MK-7 was categorized into four modules, namely, the MK-7 pathway (Module I), the shikimate (SA) pathway (Module II), the methylerythritol phosphate (MEP) pathway (Module III), and the glycerol metabolism pathway (Module IV). Overexpression of menA (Module I) resulted in 6.6 ± 0.1 mg/L of MK-7 after 120 h fermentation, which was 2.1-fold that of the starting strain BS168NU (3.1 ± 0.2 mg/L). Overexpression of aroA, aroD, and aroE (Module II) had a negative effect on the synthesis of MK-7. Simultaneous overexpression of dxs, dxr, yacM, and yacN (Module III) enabled the yield of MK-7 to 12.0 ± 0.1 mg/L. Moreover, overexpression of glpD (Module IV) resulted in an increase of the yield of MK-7 to 13.7 ± 0.2 mg/L. Furthermore, deletion of dhbB reduced the consumption of the intermediate metabolite isochorismate, thus promoting the yield of MK-7 to 15.4 ± 0.6 mg/L. Taken together, the final resulting strain MK3-MEP123-Gly2-Δ dhbB with simultaneous overexpression of menA, dxs, dxr, yacM-yacN, glpD and deletion of dhbB enabled the yield of MK-7 to 69.5 ± 2.8 mg/L upon 144 h fermentation in a 2 L baffled flask.

Ceramide lipid-based nanosuspension for enhanced delivery of docetaxel with synergistic antitumor efficiency.

Ceramide (CE), a bioactive lipid with tumor suppression, has been widely used as a drug carrier and enhancer for cancer therapy. CE-based combination therapy was prone to be attractive in cancer therapy. In our previous study, the combination of CE and docetaxel (DTX) was proved to be an effective strategy for cancer therapy. To further improve the antitumor efficiency of DTX, the CE lipid-based nanosuspensions (LNS) was prepared for the delivery of DTX to exhibit synergistic therapeutic effect. The enhanced delivery and synergistic therapeutic effect of DTX-loaded CE-LNS (CE + DTX-LNS) were evaluated. CE + DTX-LNS exhibited spherical or ellipsoidal shape, uniform particle size distribution (108.1 ± 3.8 nm), sustained release characteristics and good stability in vitro. Notably, CE + DTX-LNS could effectively co-localize CE and DTX into same tumor cell and subsequently play synergistic cell damage effect compared with CE-LNS + DTX-LNS (p < 0.05). The in vivo fluorescence imaging results showed that CE + DTX-LNS could effectively prolong the in vivo circulation time and enhance the accumulation in tumor sites. Moreover, the antitumor efficacy of CE + DTX-LNS observed in B16 murine melanoma model was 93.94 ± 2.77%, significantly higher than that of CE-LNS, DTX-LNS, Duopafei® (p < 0.01) and CE-LNS + DTX-LNS (p < 0.05), respectively, demonstrating that co-delivery of CE and DTX into same tumor cell was the basis for enhanced synergistic therapeutic effect. Furthermore, histological examination of Blank-LNS showed no visible tissue toxicity compared to normal saline. Consequently, CE-LNS could effectively delivery DTX and CE + DTX-LNS exhibit synergistic inhibition of tumor growth due to the co-localization of CE and DTX. CE-LNS hold great potential to be an appropriate carrier for CE-based combination chemotherapy.