Evaluation of the Suitability of Kinetic Chromogenic LAL Assay for Determination of Endotoxin Levels in Heparin Sodium Injection

 Determination of the levels of endotoxins in injectable products has always been one of the concerns of regulatory authorities and manufacturers. Since a number of pharmaceuticals interfere with the LAL test to some degree, overcoming the inhibition or enhancement properties of a product is required as part of the validation of the LAL test for use in the final release testing of parenteral products. In this study, interference profile of Heparin injection in quantitative chromogenic LAL test was evaluated and the method of overcoming was investigated and validated. The results indicate that dilution as the most widely used technique for overcoming interference could not eliminate LAL interference in the aforementioned medicinal product. The inhibitory nature of heparin occurs due to its anticoagulant properties and can be overcome by using divalent cations such as magnesium. Three concentrations of magnesium chloride were evaluated for elimination of heparin’s inhibitory effect. All three concentrations studied (5, 10 and 25 mM) could effectively eliminate the inhibitory effects of heparin. Hence, one-way analysis of variance was used to determine statistically significant differences between these three concentrations. The results of ANOVA statistical method showed the optimal recovery of spiked endotoxin was at a concentration of 10 mM of magnesium chloride. In consequence, chromogenic LAL test using 10 mM of magnesium chloride as diluent could be a validated method of choice for heparin LAL assay.Highlights Bacterial endotoxins are important contaminants associated with injectable pharmaceuticals.Kinetic chromogenic LAL assay was used as the method to determine endotoxin levels in heparin injections.Selectivity, linearity and repeatability of the endotoxin chromogenic method was validated

A Novel Induction Strategy Based on Temperature and pH Optimization to Improve the Yield of Recombinant Streptokinase in Escherichia coli

Introduction:Streptokinase (SK) is used for thrombolytic therapy of acute myocardial infarction. Recombinant fermentation is the most common and cost-effective procedure in the production SK. Nutrients composition, as well as such fermentation variables as temperature and pH, can affect the production level of protein-based drugs expressed in E. coli. In the present study, optimized recombinant streptokinase (r-SK) production in E. coli in the form of inclusion body (IB), the effect of induction temperature and extracellular pH were evaluated.Methods and Results:To maximize the expression level of r-SK three different induction temperatures were used to obtain the optimal temperature for r-SK production. The effect of extracellular pH on the level of r-SK production was also studied and the optimal pH was determined. To this purpose, 8 fermentation processes were applied and followed by: Cell disruption, Isolation of IBs, determination of target protein (SDS-PAGE, Western-blot). The biological activity of r-SK obtained from optimal conditions was determined in comparison with reference standard using gel clot method.The highest amount of expression level and IBs formation were performed by combining two temperatures: 42˚C for the first two hours of induction followed by 39˚C at pH 6. The expression level of r-SK under these conditions shows an 11.6% increase in comparison with the control. Conclusions:Choosing appropriate cultivation conditions can increase IB formation and use its advantages to produce higher quantities of r-SK with appropriate biological activity

Preparation and Characterization of pH-sensitive Doxorubicin Grafted Hyaluronic Acid Nano-micelles

Background: This study was done to achieve a new drug delivery system delivering two different chemotherapy molecules to the target tissue simultaneously. Significance: Co-delivery of chemotherapy medicines provides synergistic effects leading to lowering the dose of administered drugs and side effects. Methods: Doxorubicin (DOX) was introduced to water-soluble hyaluronic acid (Hyal) using 1-amino-3,3-diethoxy-propane (ADEP), as a pH-sensitive linker, to develop a new hydrophobic structure, i.e. Hyal-ADEP-DOX. The conjugates were grafted in three ratios (7.5%, 12.5%, and 20%) to Hyal and characterized by Fourier transform infrared (FT-IR) and proton nuclear magnetic resonance (1HNMR). Cannabidiol (CBD) was physically loaded in the core of nano-micelles.Results: Prepared nano-micelles were analyzed for critical micelle concentration (CMC) particle size stability and drug release before and after loading the CBD. Hyal-ADEP-DOX-12.5 was the optimized ratio and had a mean diameter of 50 nm before loading the CBD and 120 nm after loading (observed by transmission electron microscopy). The release results showed that DOX releases slowly in physiological pH, and CBD has a burst release at acidic pH from Hyal-ADEP-DOX-12.5. These properties altogether make Hyal-ADEP-DOX nano-micelles, as a stimuli-sensitive nano-carrier, a potential candidate for hydrophobic anticancer agents’ co-delivery.Conclusion: The grafted DOX exhibited pH-sensitive release behavior, i.e. while 22.8% of the drug was released after 24 h at pH 5.5, and 5% was released after 24 h at pH 7.4. Hyal-ADEP-DOX due to its low CMC value, colloidal stability, slow drug release in physiological pH, and burst release in acidic pH Hyal-ADEP-DOX, is an excellent candidate as a carrier to co-deliver hydrophobic therapeutic agents to tumor tissues

Preparation and Characterization of Different Multilayer Alginate Microcapsules

Introduction: Microencapsulation technology is a valuable technique for protection and delivery of materials which cannot be administered alone due to their low solubility, volatility and etc. A biopolymer alginate has been used most commonly as microcapsule forming material. In order to increase the mechanical stability and safety of alginate microcapsules, multilayered microcapsules are prepared. Alginate-poly l-lysine alginate, is the oldest multilayered microcapsule. Introducing another polycationic polymer which has similar properties to poly l-lysine (PLL) and also is  much more cost effective , so it might be a valuable suggestion. The aim of this study was to compare linear (LPEI) and branched (BPEI) form of polyethylene imine, with the oldest and rather expensive one, PLL, and also, we have compared sodium cellulose sulfate (NCS) as an anionic layer with sodium alginate in outer layer of multilayered microcapsules to investigate the effect of different covering layers in microcapsule’s cytotoxicity. Methods and Results: In this study by using electrostatic bead generator different types of microcapsules, APA, ALA and ABA were produced. Shape, size, surface morphology, mechanical stability and cytotoxicity of microcapsules were evaluated using optical microscope, SEM, explosion test and MTT assay respectively. According to shape and size evaluation, multilayered microcapsules with different cationic layer concentrations (0.01, 0.03 and 0.06 W/V %) were spherical, with a diameter range of 500- 900 μm. SEM images showed uniform and smooth surfaces. Explosion test revealed that applying cationic solutions with 0.03% and 0.01% concentration resulted in higher mechanical stability for ALA and APA in comparison to ABA (P<0.05), while mechanical strength induced by cationic solutions with 0.06% concentration were not statistically different in all three groups of microcapsules (P>0.05). MTT assay on HepG2 cell line was performed using microcapsules ALA, ALN, APA and APN and showed no statistically significant difference in cell viability for the all types. Conclusions: According to our results, LPEI as a covering layer for alginate microcapsules showed the same properties as PLL. Therefore it could be introduced as a cost effective alternative to PLL in fabrication of multilayered microcapsules

Preparation and Optimization of Vancomycin hydrochloride Encapsulated Multivesicular Liposomes for Sustained Locoregional Delivery

Introduction: Osteomyelitis is a destructive inflammatory condition of the bone that is usually caused by a wide range of microorganisms especially Staphylococcus aureus. Considering the downsides of systemic antibiotic therapies as well as conventional local drug delivery systems such as using polymethylmethacrylate, this study aimed to develop, characterize and optimize vancomycin hydrochloride loaded multivesicular liposomes (MVLs) as a proper therapeutic option for the treatment of osteomyelitis. Methods and Results: A 23 full factorial design technique was applied to determine the effects of three variables (lipid to drug ratio, triolein content and cholesterol to phospholipid ratio) on the encapsulation efficiency and release profile of vancomycin hydrochloride loaded MVLs to optimize the final formulation. Further characterization was performed on the optimized formula by evaluating the morphology, size and storage stability. The average drug encapsulation efficiency and the mean diameter of the optimized formulation was 54.7 ± 0.3% and 9.019 ± 0.26 µm, respectively with a span value of 0.188. Additionally, the spherical and multivesicular nature of MVLs was visible using optical microscopy (x400). The optimized formula showed an in vitro sustained release characteristic with proper stability and insignificant change in size, morphology and EE% for 30 days at 4°C. Conclusion: This study suggests that vancomycin hydrochloride loaded MVLs might have the potential to be used in the treatment of chronic osteomyelitis as a biocompatible drug carrier with a high antibiotic entrapment capacity as well as controlled drug release

Compcrypt–lightweight ANS-based compression and encryption

Compression is widely used in Internet communication to save communication time and bandwidth. Recently invented by Jarek Duda asymmetric numeral system (ANS) offers an improved efficiency and a close to optimal compression. The ANS algorithm has been deployed by major IT companies such as Facebook, Google and Apple. Compression by itself does not provide any security (such as confidentiality or authentication of transmitted data). An obvious solution to this problem is an encryption of compressed bitstream. However, it requires two algorithms: one for compression and the other for encryption. In this work, we investigate natural properties of ANS that allow to incorporate authenticated encryption using as little cryptography as possible. We target low-level security communication such as transmission of data from IoT devices/sensors. In particular, we propose three solutions for joint compression and encryption (compcrypt). All of them use a pseudorandom bit generator (PRGB) based on lightweight stream ciphers. The first solution applies state jumps controlled by PRGB. The second one employs two ANS algorithms, where compression switches between the two. The switch is controlled by a PRGB bit. The third compcrypt modifies the encoding function of ANS depending on PRGB bits. Security and efficiency of the proposed compcrypt algorithms are evaluated

Preparation and Characterization of pH-sensitive Doxorubicin Grafted Hyaluronic Acid Nano-micelles

Background: This study was done to achieve a new drug delivery system delivering two different chemotherapy molecules to the target tissue simultaneously. Significance: Co-delivery of chemotherapy medicines provides synergistic effects leading to lowering the dose of administered drugs and side effects. Methods: Doxorubicin (DOX) was introduced to water-soluble hyaluronic acid (Hyal) using 1-amino-3,3-diethoxy-propane (ADEP), as a pH-sensitive linker, to develop a new hydrophobic structure, i.e. Hyal-ADEP-DOX. The conjugates were grafted in three ratios (7.5%, 12.5%, and 20%) to Hyal and characterized by Fourier transform infrared (FT-IR) and proton nuclear magnetic resonance (1HNMR). Cannabidiol (CBD) was physically loaded in the core of nano-micelles. Results: Prepared nano-micelles were analyzed for critical micelle concentration (CMC) particle size stability and drug release before and after loading the CBD. Hyal-ADEP-DOX-12.5 was the optimized ratio and had a mean diameter of 50 nm before loading the CBD and 120 nm after loading (observed by transmission electron microscopy). The release results showed that DOX releases slowly in physiological pH, and CBD has a burst release at acidic pH from Hyal-ADEP-DOX-12.5. These properties altogether make Hyal-ADEP-DOX nano-micelles, as a stimuli-sensitive nano-carrier, a potential candidate for hydrophobic anticancer agents’ co-delivery. Conclusion: The grafted DOX exhibited pH-sensitive release behavior, i.e. while 22.8% of the drug was released after 24 h at pH 5.5, and 5% was released after 24 h at pH 7.4. Hyal-ADEP-DOX due to its low CMC value, colloidal stability, slow drug release in physiological pH, and burst release in acidic pH Hyal-ADEP-DOX, is an excellent candidate as a carrier to co-deliver hydrophobic therapeutic agents to tumor tissues

Перинатология. Настоящее и будущее

На основании обобщения многолетнего собственного опыта автора, данных литературы и результатов работы Московского центра планирования семьи и репродукции освещены актуальные проблемы перинатологии от пренатального периода до постнатальной охраны здоровья плода. Сделано заключение о необходимости и возможности создания стандартов в области лечебных мероприятий и тактики ведения родов при осложнениях.Basing on the generalization of many−year experience as well as the data of the literature and results of the work of Moscow Center for Family Planning and Reproduction, the author features the urgent issues of perinatology from prenatal period to postnatal health protection. The author concludes about the necessity and possibility to create the standards in the field of therapeutic measures and management tactics in complicated delivery

Antibacterial Effect of Hydroalcoholic Extract of Punica granatum Linn. Petal on Common Oral Microorganisms

Objectives. This study aimed to assess the effect of hydroalcoholic extract of Punica granatum Linn. (P. granatum) petal on Streptococcus sanguinis, Streptococcus mutans, Streptococcus salivarius, Streptococcus sobrinus, and Enterococcus faecalis. Materials and Methods. In this in vitro study, P. granatum extract was prepared using powdered petals and water-ethanol solvent. Antibacterial effect of the extract, chlorhexidine (CHX), and ampicillin was evaluated on brain heart infusion agar (BHIA) using the cup-plate method. By assessing the diameter of the growth inhibition zone, the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the extract were determined for the above-mentioned bacteria. Results. Hydroalcoholic extract of P. granatum petal had inhibitory effects on the proliferation of all five bacterial strains with maximum effect on S. mutans with MIC and MBC of 3.9 mg/mL. The largest growth inhibition zone diameter belonged to S. sanguinis and the smallest to E. faecalis. Ampicillin and CHX had the greatest inhibitory effect on S. sanguinis. Conclusions. Hydroalcoholic extract of P. granatum had a significant antibacterial effect on common oral bacterial pathogens with maximum effect on S. mutans, which is the main microorganism responsible for dental plaque and caries

Pseudorandom Bit Generation with Asymmetric Numeral Systems

The generation of pseudorandom binary sequences is of a great importance in numerous applications stretching from simulation and gambling to cryptography. Pseudorandom bit generators (PRBGs) can be split into two classes depending on their claimed security. The first includes PRBGs that are provably secure (such as the Blum-Blum-Shub one). Security of the second class rests on heuristic arguments. Sadly, PRBG from the first class are inherently inefficient and some PRBG are insecure against quantum attacks. While, their siblings from the second class are very efficient, but security relies on their resistance against known cryptographic attacks. This work presents a construction of PRBG from the asymmetric numeral system (ANS) compression algorithm. We define a family of PRBGs for $2^R$ ANS states and prove that it is indistinguishable from a truly random one for a big enough $R$. To make our construction efficient, we investigate PRBG built for smaller $R=7,8,9$ and show how to remove local correlations from output stream. We permute output bits using rotation and Keccak transformations and show that permuted bits pass all NIST tests. Our PRBG design is provably secure (for a large enough $R$) and heuristically secure (for a smaller $R$). Besides, we claim that our PRBG is secure against quantum adversaries