Hydrogen Peroxide-Resistant CotA and YjqC of Bacillus altitudinis Spores Are a Promising Biocatalyst for Catalyzing Reduction of Sinapic Acid and Sinapine in Rapeseed Meal.

For the more efficient detoxification of phenolic compounds, a promising avenue would be to develop a multi-enzyme biocatalyst comprising peroxidase, laccase and other oxidases. However, the development of this multi-enzyme biocatalyst is limited by the vulnerability of fungal laccases and peroxidases to hydrogen peroxide (H2O2)-induced inactivation. Therefore, H2O2-resistant peroxidase and laccase should be exploited. In this study, H2O2-stable CotA and YjqC were isolated from the outer coat of Bacillus altitudinis SYBC hb4 spores. In addition to the thermal and alkali stability of catalytic activity, CotA also exhibited a much higher H2O2 tolerance than fungal laccases from Trametes versicolor and Trametes trogii. YjqC is a sporulation-related manganese (Mn) catalase with striking peroxidase activity for sinapic acid (SA) and sinapine (SNP). In contrast to the typical heme-containing peroxidases, the peroxidase activity of YjqC was also highly resistant to inhibition by H2O2 and heat. CotA could also catalyze the oxidation of SA and SNP. CotA had a much higher affinity for SA than B. subtilis CotA. CotA and YjqC rendered from B. altitudinis spores had promising laccase and peroxidase activities for SA and SNP. Specifically, the B. altitudinis spores could be regarded as a multi-enzyme biocatalyst composed of CotA and YjqC. The B. altitudinis spores were efficient for catalyzing the degradation of SA and SNP in rapeseed meal. Moreover, efficiency of the spore-catalyzed degradation of SA and SNP was greatly improved by the presence of 15 mM H2O2. This effect was largely attributed to synergistic biocatalysis of the H2O2-resistant CotA and YjqC toward SA and SNP

One-Pot Hydrothermal Synthesis of mSiO₂-N-CDs with High Solid-State Photoluminescence as a Fluorescent Probe for Detecting Dopamine

An effective fluorescent probe (mSiO2-N-CDs) was prepared by embedding N-CDs into mesoporous silica via a simple one-pot hydrothermal reaction and applied to the detection of dopamine (DA). Mesoporous silica not only provided a skeleton to prevent the aggregation of N-CDs but also a medium for the centrifugal collection of N-CDs, avoiding the need for dialysis and freeze-drying. The formation process, phase composition, morphology, and luminescence properties of the composite were studied in detail. The synthesized mSiO2-N-CDs possessed spherical morphology, a smooth surface, and a diameter of approximately 150 nm. The fluorescence results indicated that mSiO2-N-CDs emitted intense blue color fluorescence at 465 nm under the optimal excitation of 370 nm. Because the mesoporous silica effectively inhibited the self-quenching caused by the aggregation of N-CDs, the quantum yield of solid mSiO2-N-CDs powder reached 32.5%. Furthermore, the emission intensity of the solid mSiO2-N-CDs remained constant for 28 days. The good sensitivity and selectivity of mSiO2-N-CDs for DA enabled the establishment of a rapid, simple, and sensitive DA detection method. The linear range was 0–50 µM and the limit of detection was calculated to be 107 nM. This method was used for the determination of DA in urine, with recovery rates ranging between 98% and 100.8%. In addition, the sensing mechanism was characterized by fluorescence lifetime decay and UV–VIS spectral analysis

Efficient Tissue Culture Protocol for Magnolia lucida (Magnoliaceae) and Confirmation of Genetic Stability of the Regenerated Plants

Magnolia lucida (Magnoliaceae) is classified as an endangered species by the International Union for Conservation of Nature. It has high commercial value owing to its attractive tree shape and flowers. We adopted an excellent genotype of M. lucida as the parent material and established a mini-cut orchard through grafting to provide trunk shoots explants over the long-term. Optimal sterilization was achieved using a combination of 75% ethanol for 30 s, one percent benzalkonium bromide for five minutes, and 0.1% mercuric chloride for five minutes. Modified Murashige and Skoog medium (ML) was the optimal medium for the growth of M. lucida. Addition of one mg/L of 6-benzyl adenine (BA) and 0.05 mg/L of α-naphthaleneacetic acid (NAA) to the medium increased the shoot induction rate to 95.56%, and the ML medium containing 0.4 mg/L BA and 0.04 mg/L NAA achieved the maximum multiplication rate (284.56%). Dark treatment for seven days, followed by continuous light treatment could better resolve the challenge of difficult rooting in M. lucida plants. Using random amplified polymorphic DNA and inter simple sequence repeat markers, we confirmed the genetic uniformity and stability of the regenerated plants. Our protocol should be helpful for the propagation and conservation of this endangered plant

Purification of YjqC and CotA from <i>B</i>. <i>altitudinis</i> SYBC hb4 spores.

<p>Purification of YjqC and CotA from <i>B</i>. <i>altitudinis</i> SYBC hb4 spores.</p

Effects of H₂O₂ concentrations on activity of YjqC and CotA from <i>B</i>. <i>altitudinis</i> hb4.

<p>(A) Variable H₂O₂ effects on peroxidase activity from the free and spore YjqC. (B) Inhibition by H₂O₂ of CotA activity toward ABTS, SA and SNP. (C) Stability of CotA incubated with different concentrations of H₂O₂ for 0–180 min. (D) Effects of H₂O₂ concentrations on enzymatic activity of spore enzymes toward SA and SNP.</p

Optimal activity of the free and spore CotA.

<p>Laccase activity toward ABTS under different pH (A) and temperature (B) conditions. Laccase activity toward SA and SNP under different pH (C) and temperature (D) conditions. Stability of laccase activity toward ABTS (E), SA and SNP (F) under optimal pH and temperature conditions.</p

Optimal activity of the free and spore YjqC.

<p>Catalase activity under different pH (A) and temperature (B) conditions. Peroxidase activity toward SA and SNP under different pH (C) and temperature (D) conditions. Stability of catalase (E) and peroxidase (F) activity under optimal pH and temperature conditions.</p

Identification of proteins extracted from spore coat fraction of <i>B</i>. <i>altitudinis</i> SYBC hb4.

<p>Identification of proteins extracted from spore coat fraction of <i>B</i>. <i>altitudinis</i> SYBC hb4.</p

Characteristics of YjqC and CotA purified from <i>B</i>. <i>altitudinis</i> SYBC hb4 spores.

<p>Characteristics of YjqC and CotA purified from <i>B</i>. <i>altitudinis</i> SYBC hb4 spores.</p

Effects of substrate concentrations on enzyme activity.

<p>Catalase activity of the free and spore YjqC under different concentrations of H₂O₂ (A). Peroxidase activity of the free and spore YjqC toward different concentrations of SA (B) and SNP (C). Laccase activity of the free and spore CotA toward different concentrations of ABTS (D), SA (E) and SNP (F).</p