Process analytical technology as in-process control tool in semi-continuous manufacturing of PLGA/PEG-PLGA microspheres.

Nowadays, among 3rd generation drug delivery systems, biodegradable polymeric based long-acting injectable depot has achieved tremendous success in clinical application. So far, there have been two dozen of commercial products of Poly (lactic-co-glycolic acid) microspheres available in the market. Recently, continuous manufacturing concept has been successfully applied on oral solid formulation from buzzword to reality. However, the polymeric injectable microspheres are still stayed at batch manufacturing phase due to the lack of understanding of knowledge matrix. In this study, micro-mixer as a plug-and-play emulsification modules, Raman spectroscopy and focused beam reflectance measurement as real-time monitoring modules are integrated into a novel semi-continuous manufacturing streamline to provides more efficient upscaling flexibility in microspheres production. In this end to end semi-continuous manufacturing process, amphiphilic block polymer monomethoxy-poly (ethylene glycol) modified PLGA (mPEG-PLGA) was used for encapsulating Gallic acid. Additionally, with guarantee of good robustness, the correlation relationship between critical process parameters, critical material attributes and critical quality attributes were investigated. The time-space evolution process and mechanism for formation of PEG-PLGA microsphere with particular morphology were elaborated. Altogether, this study firstly established semi-continuous manufacturing streamline for PLGA/PEG-PLGA microspheres, which would not only lower the cost of production, narrow process variability and smaller equipment/environmental footprint but also applied in-process control (IPC) and QbD principle on complicated production process of microspheres. Therefore, this study build confidence in the industrial development of PLGA/PEG-PLGA microspheres and establish best practice standards, which might be a quantum leap for developing PLGA microspheres in the future.

Efficacy and Mechanism of Chinese Medicinal Compounds and Monomers from Chinese Medicinals against Colorectal Cancer： A Review / 中国实验方剂学杂志

Colorectal cancer （CRC） is one of the refractory malignant tumors of the digestive system worldwide. With limitations， the available clinical therapies are usually suspended or show unsatisfactory effect. Therefore， it is urgent to find and develop new candidate drugs specifically targeting the cancer with ideal efficacy， low toxicity， and low cost， and the solutions can be found in traditional Chinese medicine （TCM） which has a long history. In TCM， sovereign， ministry， assistant， and guiding medicinals are selected based on the syndrome differentiation， and it has shown remarkable efficacy on CRC in recent years. In particular， Chinese medicinal compounds and monomers from Chinese medicinals which have been applied in clinical settings are advantageous in the treatment of CRCs， as they improve the quality of life， alleviate clinical symptoms and toxic and side effects of chemotherapy， and prolong the survival of patients. Therefore， we retrieved the English and Chinese articles with "CRC"， "TCM"， "compound" and "monomer" as keywords， and summarized the progress in the treatment of CRC with Chinese medicinal compounds and monomers from Chinese medicinals from four aspects of "replenishing Qi and invigorating spleen"， "clearing heat and removing toxin"， "nourishing liver and kidney"， and "tonifying Qi and nourishing blood". However， Chinese medicine features multiple components， multiple targets， and multiple pathways， and in-depth research should be carried out on the application of Chinese medicinal compounds and monomers from Chinese medicinals in the treatment of CRC， in an attempt to minimize the pain and side effects and maximize the therapeutic effect. This study is expected to provide new insight into the treatment of CRC and a reference for further research on the efficacy and mechanism of Chinese medicine.

Optimization and Evaluation of the Quercus infectoria Galls Thermosensitive In Situ Gel for Rectal Delivery.

Quercus infectoria galls (QIGs) have a long history of treating ulcerative colitis (UC). The aqueous extract of QIG has an anti-UC effect. However, QIG's enema is easy to leak, and the action time and dose of the drug cannot be controlled well. Thus, QIG is inconvenient to use. This study aims to screen and prepare an optimized thermosensitive in situ gel with slow release and retention. Taking the transition sol-gel temperature (T sol-gel) as the investigation index, the Box-Behnken design response surface method (BBD-RSM) was used to optimize the dosages of Poloxamer 407 (P407), Poloxamer 188 (P188), and hydroxypropyl methyl cellulose (HPMC). Moreover, three formulations were selected, and the in vitro release rates were further optimized. The optimized rates of P407, P188, and HPMC were 24.07%, 1.22%, and 0.60%, respectively, and T sol-gel was 32.8°C ± 0.4°C. The cumulative release of gallic acid in the gel conformed to the first-order kinetic equation, and gallic acid was released entirely within 24 h. In addition, the morphological and chemical characterization of thermosensitive in situ gel demonstrated that excipients did not affect the characteristic functional groups of QIG and that the surface of the QIG gel had a porous and loose structure. Rheological methods showed that the QIG thermosensitive in situ gel was fluid at low temperature and semisolid at gelation temperature. Therefore, the prepared gel was sensitive to temperature and had slow-release, local retention properties.

Study on Anti-colorectal Cancer Mechanism of Gallic Acid Based on JAK/STAT Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the effects of gallic acid （GA） on human colon cancer HCT-116 and Caco-2 cell activities， intracellular Janus kinase （JAK）/signal transducer and activator of transcription factor （STAT） signaling pathway， and the expression of anti-apoptotic protein B-cell lymphoma-2 （Bcl-2） and pro-apoptotic protein B-cell lymphoma-2-associated X protein （Bax）， so as to explore its underlying molecular mechanism. MethodFollowing the classification of cells into GA group， blank group， and 5-fluorouracil （5-FU， 0.05 g·L-1） group， the HCT-116 and Caco-2 cells were treated with GA （0.02， 0.05， 0.1， 0.15， 0.2 g·L-1） for 12， 24， 48， and 72 h， respectively， and the cell proliferation inhibition rats were determined by cell counting kit-8 （CCK-8） assay to select the GA concentration that effectively inhibited proliferation. The colony formation ability was detected by crystal violet staining and the migration of cells by scratch test. The level of reactive oxygen species （ROS） was measured using a fluorescent probe （DCFH-DA）. The expression of interleukin-6 （IL-6） and tumor necrosis factor-α （TNF-α） in cell supernatant were determined using the enzyme-linked immunosorbent assay （ELISA） kits. The expression levels of JAK2， phosphorylated （p）-JAK2， STAT3， p-STAT3， Bcl-2， and Bax were assayed by Western blot. ResultCCK-8 assay showed that after 12， 24， 48， and 72 h of treatment， GA （0.02， 0.05， 0.1， 0.15， 0.2 g·L-1） inhibited the proliferation of HCT-116 and Caco-2 cells in a dose- and time-dependent manner， and the inhibition rates were higher than those in the blank control group. Compared with the 5-FU group， GA （0.2 g·L-1） enhanced the inhibition of cell proliferation in a time-dependent manner. Compared with the blank control group， GA （0.1， 0.15， and 0.2 g·L-1） significantly decreased the number of cell colonies （P<0.01）， increased the inhibition rate of cell colony formation （P<0.01）， diminished the scratch healing rate （P<0.05， P<0.01）， elevated the fluorescence intensity of intracellular ROS （P<0.01）， and down-regulated the expression of IL-6 and TNF-α in the supernatant （P<0.01） in a dose-dependent manner. Compared with the 5-FU group， GA （0.2 g·L-1） decreased the scratch healing rate （P<0.01）， enhanced the fluorescence intensity of intracellular ROS （P<0.01）， and down-regulated the levels of IL-6 and TNF-α in cell supernatant （P<0.01）. According to Western blot analysis， compared with the blank control group， GA （0.1， 0.15， 0.2 g·L-1） obviously lowered the expression of p-JAK2， p-STAT3， Bcl-2， p-JAK2/JAK2， p-STAT3/STAT3， and Bcl-2/Bax （P<0.01） and raised Bax protein expression （P<0.05， P<0.01） in a dose-dependent manner. Compared with the 5-FU group， GA （0.2 g·L-1） down-regulated the expression of p-JAK2， p-STAT3， Bcl-2， p-JAK2/JAK2， p-STAT3/STAT3， and Bcl-2/Bax （P<0.05， P<0.01） and up-regulated the expression of Bax protein （P<0.05， P<0.01）. ConclusionGA significantly inhibits the proliferation of HCT-116 and Caco-2 cells， which may be related to the increased accumulation of intracellular ROS， down-regulation of inflammatory factors IL-6 and TNF-α， p-JAK2 and p-STAT3 protein expression in JAK/STAT signaling pathway， and Bcl-2， and up-regulation of Bax.

Study on regulation of ethanol extract of Turkish Galls on JAK 2/STAT3 signaling pathway in colorectal cancer cells / 中国药房

OBJECTIVE To study the regulatory mechanism of ethanol extract of Turkish Galls on proliferation and migration of colorectal cancer cells HCT- 116 and Caco- 2 based on janus kinase 2（JAK2）/signal transducer and activator of transcription 3 （STAT3）signaling pathway. METHODS CCK-8 method was used to detect the effects of 0.05，0.1，0.2，0.3，0.4 and 0.5 mg/mL ethanol extract of Turkish Galls on the proliferation of HCT- 116 and Caco- 2 cells after treated for 12，24，48 and 72 h. After treated with 0.1，0.3，0.5 mg/mL ethanol extract of Turkish Galls for 24 h，the migrations of HCT- 116 and Caco- 2 cells were detected by scratch test ；the level of reactive oxygen species （ROS）was detected by fluorescent probe method. The levels of interleukin-6（IL-6）and tumor necrosis factor-α（TNF-α）in cell supernatant were detected by ELISA . The phosphorylations of JAK2 and STAT 3 as well as the expressions of B-cell lymphoma- 2（Bcl-2）and Bcl- 2 associated protein X （Bax）were detected by Western blot assay. RESULTS Compared with blank control ，0.05，0.1，0.2，0.3，0.4 and 0.5 mg/mL ethanol extract of Turkish Galls could significantly inhibit cell proliferation after treated for 12，24，48，72 h（P＜0.05）. After treated with 0.1，0.3 and 0.5 mg/mL ethanol extract of Turkish Galls for 24 h，the scratch healing rate of 2 kinds of cells ，the levels of IL- 6 and TNF-α in the cell supernatant ，the phosphorylation of JAK 2 and STAT 3 as well as the expression of Bcl- 2 protein were all significantly decreased （P＜0.05）；the level of ROS and protein expression of Bax were increased significantly （P＜0.05）. CONCLUSIONS The ethanol extract of Turkish Galls can inhibit the proliferation and migration of HCT- 116 and Caco- 2 cells. The mechanism may be related with down-regulation of protein expression of Bcl- 2 and up-regulation of protein expression of Bax by increasing the accumulation of intracellular ROS ，down-regulating the expressions of inflammatory factors IL- 6 and TNF-α and the phosphorylation of JAK2 and STAT 3 in JAK 2/STAT3 signaling pathway.

A review of the phytochemical, pharmacological, pharmacokinetic, and toxicological evaluation of Quercus Infectoria galls.

ETHNOPHARMACOLOGICAL RELEVANCE: Quercus Infectoria galls (QIG) have a long history of use in traditional Chinese medicine and traditional Uyghur medicine for the treatment of diarrhea, hemorrhage, skin disease, and many other human ailments. Medicinal applications of QIG have become increasingly popular in Greece, Asia Minor, Syria, and Iran. AIM OF THE REVIEW: The present paper reviewed the ethnopharmacology, phytochemistry, analytical methods, biological activities, metabolism, pharmacokinetics, toxicology, and drug interactions of QIG to assess the ethnopharmacological uses, explore its therapeutic potential, and identify future opportunities for research. MATERIALS AND METHODS: Information on QIG was gathered via the Internet (using Google Scholar, Baidu Scholar, Elsevier, ACS, Pubmed, Web of Science, CNKI, and EMBASE) and libraries. Additionally, information was also obtained from local books and PhD and MS dissertations. RESULTS: QIG has played an important role in traditional Chinese medicine. The main bioactive metabolites of QIG include tannins, phenolic acids, flavonoids, triterpenoids, and steroids. Scientific studies on the QIG extract and its components have shown its wide range of pharmacological activities, such as cholinesterase- and monoamine oxidase-inhibitory, antitumor, anti-hypertension, antidiabetic, antimicrobial, insecticidal, antiparasitic, antioxidant, and anti-inflammatory. CONCLUSIONS: The ethnopharmacological, phytochemical, pharmacological, and analytical methods of QIG were highlighted in this review, which provides information for future studies and commercial exploration. QIG has a huge potential for pharmaceutical and nutraceutical applications. Moreover, comprehensive toxicity studies of this plant must be conducted to ensure its safety. Additional investigations are recommended to transmute the ethnopharmacological claims of this plant in folklore medicines into scientific rationale-based information. Research on pharmacokinetics studies and potential drug interactions with standard-of-care medications is still limited, which calls for additional studies particularly on humans. Further assessments and clinical trials should be performed before it can be integrated into medicinal practices.

Traditional Uyghur medicine Quercus infectoria galls water extract triggers apoptosis and autophagic cell death in colorectal cancer cells.

BACKGROUND: The water extract of Quercuse infectoria galls (QIG) is the active ingredient of Uyghur medicine Xipayi Kui Jie'an (KJA) which has promising therapeutic effects on Ulcerative Colitis (UC) as an alternative medicine. Considering the relationship between UC and the development of colorectal cancer (CRC), the present work aims to explore the direct anti-CRC activity of QIG extract. METHODS: CCK8 assay and flow cytometry were used to detect cytotoxicity and apoptosis. Transmission electron microscopy (TEM), flow cytometry, laser confocal and western blotting were performed to examine autophagy. We also adopted Reactive Oxygen Assay kit, as well as transwell and wound healing tests to study the underlying mechanism of QIG against CRC cells. RESULTS: First, we found that QIG extract could suppress the viability of CRC cells and trigger caspases-dependent apoptosis. Subsequently, we proved for the first time that QIG extract also triggered autophagic cell death in CRC cells, which together with apoptosis contributed to the cytotoxic effect on CRC cells. Further investigation revealed that QIG-induced cytotoxicity associated with intracellular ROS accumulation which could suppress the AKT/mTOR signaling pathway, and then induce autophagy and inhibit cell growth. Besides, Erk signaling pathway was also involved in the process of autophagic cell death. Moreover, QIG extract also influenced EMT process and inhibited CRC cell migration. CONCLUSION: Altogether, this study provides a basis for the utilization of QIG as an alternative medicine for CRC prevention and treatment.

[Process of gallnut suppository preparation].

The main objective was to research the process of gallnut suppository preparation with its water extract as the main drug, and evaluate its irritation to rectal mucosa. gallnut extract was obtained by decocting method, and its suppository preparation was obtained by fusion method with semi-synthetic aliphatic esters and rose flower oil as the matrix. Weight difference and in vitro melting time limit of the suppository were assayed and UV-Vis was used to determine the contents of polyphenols, tannin and saccharide. The irritation to colon mucosa was evaluated after successive administration of 14 days to New Zealand white rabbits. Finally, the prescription compositions were determined: semi-synthetic aliphatic esters and rose flower oil with the ratio of 2:1 as the proper matrix, with the drug loading of 54%. The prepared suppository was brown, conical and smooth. The weight difference was (1.43±0.03) g, with an average melting time limit of (17±2) min. The Contents of Polyphenols, tannic and polysaccharide were 332.4, 245.0, 3.3 mg•g-1 respectively in each suppository. The results also showed that the continuous administration had no irritation to rectal mucosa. It can be concluded that the suppository was an acceptable administrate form, whose preparation process was easily controlled, and with no irritation to rectum mucosa.

Investigation into the toxicity of traditional Uyghur medicine Quercus infectoria galls water extract.

OBJECTIVE: Quercus infectoria galls (QIG) is being widely used in Traditional Uyghur Medicine. To gather preclinical safety information for the aqueous extract of QIG, a toxicity study was performed. METHODS: Subject animals were randomized, and divided into exposure and control groups. In the acute toxicity phase, three different doses--5, 7.5, and 10 g/kg, respectively--were administered via enema to imprinting control region (ICR) mice. An experiment using the maximum tolerance dose (MTD) i.e.10 g/kg was also performed. Data were gathered for 14 days, and study parameters were clinical signs, body weight, general behavior, adverse effects and mortality. At the day 14, major organs of the subjects were examined histologically. Chronic toxicity was also evaluated in Wistar rats for over 180 consecutive days. The rats were divided into three groups with different doses of 0.2 g/kg, 0.8 g/kg, and 2 g/kg, QIG. Furthermore, observations were carried out in rabbits to investigate if there were signs of irritation. RESULTS: In comparison to control group, acute, chronic toxicity and mortality were not significantly increased in exposure group. CONCLUSION: Study result suggests that the aqueous extract of QIG is unlikely to have significant toxicity and that clinical trials may proceed safely.