Ronald Bartzatt

Dansylation of hydroxyl and carboxylic acid functional groups

Fluorescent labeling of primary and secondary amines using dansyl chloride has been widely used in the past. Its application provides an extremely sensitive means to detect amine functional groups to amounts of less than 1 microg of material. This work describes a method for the dansylation of hydroxyl (-OH) and carboxylic acid (-COOH) functional groups. This technique is demonstrated with ethanol, gamma hydroxy butyric acid (GHB), benzoic acid, and p-chloroaniline. Sensitivity of detection for all compounds are microgram or microliter. For the compounds ethanol and GHB which are liquids at room temperature, as little as 1 microl quantity can be detected. Benzoic acid and p-chloroaniline which are solids at room temperature can be detected at levels of 1 microg. Fast thin layer chromatography was accomplished using acetone as the resolving solvent, which resulted in good differentiation of analytes for R(f) measurement. The dansylation reaction performed similarly at pH 11, 10 and 9.6 and uses 2 molar Na(2)CO(3).

Tuberculosis infections of the central nervous system.

The causative agent of Tuberculosis meningitis is Mycobacterium tuberculosis, which is the bacteria that causes pulmonary tuberculosis. Proliferating into the central nervous system occurs from other sites of infection within the body. Brain damage can result from the infection that may lead to abnormal behavior, mental impairments, motor type paralysis, and seizures. Tuberculosis infections of the central nervous system are a serious and often fatal disease predominantly impacting young children, and is thought to be the most devastating form of the disease. Isoniazid is the only first line bactericidal agent that easily crosses the blood-brain barrier and achieves concentrations in cerebrospinal fluid similar to those in serum. Rifampicin, ethambutol, and streptomycin all penetrate into the cerebrospinal fluid poorly, and even in the setting of meningeal inflammation. As much as one-third of the current worlds population may be infected with tuberculosis. Tuberculosis infection of the central nervous system is a serious type of extrapulmonary proliferation of this disease . In developing countries, it has high predominance in children. Pathological manifestations of cerebral tuberculosis occur, of which the most common is tuberculous meningitis, followed by tuberculoma, tuberculous abscess, cerebral miliary tuberculosis, tuberculous encephalopathy, tuberculous encephalitis, and tuberculous arteritis. Brain abscesses of Mycobacterium tuberculosis can induce seizures and coma leading to death and complicated due to multiorgan failure. Rapid diagnosis and early intervention is vital for successful outcome for patients. Further studies are required to understand the proliferation of tuberculosis meningitis in addition to the elucidation of new therapeutic drugs for the successful clinical treatment of this deadly disease.

Synthesis, Structural Analysis and Antibacterial Activity of a Butyl Ester Derivative of Ampicillin

Background: Ester groups placed on medicinal agents have been found to increase lipophilicity (more positive log p value). Alterations in a partition coefficient may affect medicinal activity. Formation of an ester group such as -C(O)OCH2CH2CH2CH3 from a carboxyl group [-C(O)OH] can be efficiently accomplished utilizing diazoalkanes and produces a prodrug-like structure of ampicillin (a bipartate drug carrier). Formation of an ester group may produce beneficial changes in medicinal parameters such as log P, log BB, and molecular polar surface area (TPSA). Methods: An ester of ampicillin was synthesized by replacing the carboxyl group with a butyl ester group. The ester formed was synthesized utilizing diazobutane, which is a gas at room temperature and reacts with the acidic hydrogen of the carboxylic acid. The ampicillin ester was dissolved into LB agarose media at levels of 0.228 and 0.180 mg/ml, and its antibacterial activity was compared to that of normal ampicillin. Medicinal parameters such as log P, solubility, polar surface area, log BB, molecular dipole, Clog P, intestinal absorption and the ‘Rule of 5’ were determined. Results: The ampicillin ester consisted of yellow crystals that were stable for >12 weeks at 0°C. The ester has reduced hydrogen bonding capacity and increased lipophilic character (by log P) compared to the parent structure. Each concentration of the ester tested induced 100% growth inhibition of ampicillin-susceptible but streptomycin-resistant Escherichia coli bacteria, and >30% growth inhibition of an ampicillin-resistant E. coli. This ester compound of ampicillin showed zero violations of the ‘Rule of 5’, which indicates good bioavailability and good bioactivity. Intestinal absorption indicated by TPSA value showed moderate absorption activity for both parent and ester compounds. The lipophilic substituent constant for the butyl ester is +0.941, which indicates increased lipophilicity. The ester group improves log BB and dermal permeability of ampicillin. Conclusion: The butyl ester of ampicillin inhibited the growth of susceptible and resistant E. coli. Favorable values for parameters such as TPSA, log P, log BB, and zero violations of the ‘Rule of 5’ suggest good bioavailability and good bioactivity.

The colorimetric determination of nitrate anion in aqueous and solid samples utilizing an aromatic derivative in acidic solvent

Ingestion of nitrates has been associated with cancer activity and other health disorders in humans. Presented here are colorimetric methods to detect and quantify nitrates from solids and liquids. A rapid spot test protocol determines nitrate anion to an amount of less than 0.10 µg. Applying a standard curve allows determination of in many samples by spectrometer. Assay by standard curve is sensitive to less than 1.00 mg/mL. In a strongly acidic mixture a brilliant blue color develops upon the reaction of nitrate anion with the secondary amine diphenylamine. An absorbance spectrum determined for the nitrated diphenylamine shows a broad absorbance peak at 597 nm and molar absorptivity of approximately 61 400 L/mole cm. The derivative that is formed is stable for more than 1 h. Inorganic salts such as phosphates, carbonates, sulfates, azides, and chlorides do not interfere with determination of nitrates. Groundwater, pond water, and dust particles were analyzed for nitrates. These methods are rapid, sensitive, relatively inexpensive, do not require complex instrumentation, and are useful for solid and liquid samples. The acidic solvent will dissolve sample components not soluble in water and results are easily discerned by eye. The reagent may easily be transported.

The interaction of surfactants with the chloroplast thylakoid membrane at sub-solubilizing concentrations

Abstract The interactions of nonionic, anionic and zwitterionic surfactants with the chloroplast thylakoid membrane of wheat ( Triticum aestivum ) were examined. Measurements of the surface tension of the solution and behavior of the membrane material during centrifugation were used to identify solubilizing concentrations of the surfactants. Studies at sub-solubilizing concentrations indicated that the primary interaction between the surfactant and the thylakoid membrane may involve adsorption at the membrane / solution interface rather than insertion of surfactant molecules into the membrane. This hypothesis was supported by a lack of increase in the fluorescence yield of chlorophyll at sub-solubilizing surfactant concentrations, indicating that interactions between the photosynthetic pigment-protein complexes are not extensively disrupted. Difference spectroscopy revealed that long-wavelength (greater than 690 nm) chlorophyll molecules are perturbed by the surfactant. We conclude that at sub-solubilizing concentrations, surfactants specifically interact with exposed Photosystem I components on the surface of the thylakoid membrane. It appears that use of sub-solubilizing concentrations of surfactants has the potential to provide much useful information about the structure and organization of the thylakoid membrane.

Small Molecule Hydrazide Agents to Inhibit Growth and Proliferation of Mycobacterium Tuberculosis

Four novel drug designs for the treatment of Mycobacterium tuberculosis are analyzed and shown to prevent the growth and proliferation of this dangerous bacteria. All four agents, designated A, B, C, and D, are hydrazide type compounds, where D has three hydrazide functional groups. Agents B and C have a halogenated aromatic ring substituent, while A contains a pyridine ring. Pharmaceutical properties such as Log P, polar surface area, and violations of the Rule of 5 are determined for all agents. The Polar surface area for these four agents ranged from 55.121 A2 to 165.363 A2 and Log P values for A, B, C, and D were determined at -0.916, 0.95, 0.974, and -4.921, respectively. Drug designs A, B, and C show zero violations of the Rule of 5, where D exhibits only one violation, which are outcomes describing favorable bioavailability. Values of polar surface area for A, B, and C affirm an intestinal absorption of greater than 60% as well as the potential for crossing the blood brain barrier for targeting bacterial meningitis of the central nervous system. Interaction with Mycobacterium tuberculosis was monitored over a 14 day interval with agents at known concentration. Agents A, B, C, and D elicited more than 60% inhibition of bacterial growth by day 14 at concentrations of as little as 30 micrograms/ milliliter. All agents reduced bacteria survival to less than 60% by day 7 of culture. The inhibition of bacterial growth induced by agents A, B, C, and D was comparable to that of isoniazid. K-means cluster analysis of descriptors determined isoniazid most similar to agents A, B, and C. Other characteristics of these small hydrazide compounds render supportive evidence for an efficacious clinical application.

Design of Ciprofloxacin Derivatives that Inhibit Growth of Methicillin Resistant Staphylococcus aureus (MRSA) and Methicillin Susceptible Staphylococcus aureus (MSSA)

Three derivatives of ciprofloxacin (compound B, C, and D) were constructed utilizing microwave synthesis methodology (compound D) or diazoalkane reaction in nonaqueous solvent (compounds B and C). The final structures of the derivatives featured an ester group in place of the original carboxyl group of the ciprofloxacin. These ester groups contained aliphatic single carbon (compound B), two carbon length (compound C), or three carbon length propyl ester group (compound D). The ester groups strongly affected the molecular properties of the parent ciprofloxacin. As the size of the ester group increased the formula weight, molar volume, and number of rotatable bonds increased. The Log P for these compounds were -0.701, -0.441, -0.065, 0.437 for ciprofloxacin, B, C, and compound D, respectively. Numerical values of dermal permeability coefficient (Kp) increased rapidly as length of the ester carbon chain increased. The immediate consequence of Kp increase is an increased skin penetration rate based on dose and time span of administration. Polar surface area for ciprofloxacin is 74.569 Angstroms(2), but decreases to 63.575 Angstroms(2) for all three derivatives. All three derivatives of ciprofloxacin showed zero violations of the Rule of 5, indicating these drugs would have favorable bioavailability. Compounds A, B, C, and D were placed into tissue culture with methicillin resistant and susceptible Staphylococcus aureus (MRSA and MSSA, respectively) to determine levels of bacterial growth inhibition. All compounds induced greater than 60 % inhibition of MSSA at concentrations as low as 15.63 micrograms/milliliter. All four compounds induced greater than 80 % inhibition of MRSA at concentratins as low as 15.63 micrograms/milliliter. Development of novel drug designs will benefit the clinical treatment of dangerous infections of MSSA and MRSA.

Analysis of an ampicillin propyl ester prodrug which inhibits the growth of Escherichia coli

An ampicillin prodrug was synthesized by utilizing the chemical reaction of ampicillin with diazopropane (CH3CH2CHN2) in an organic solvent. The result is esterification of the carboxylic acid functional group. The ampicillin prodrug is a solid that forms yellow crystals which are soluble in water and LB agarose media. The ampicillin prodrug was stable for more than 10 weeks when stored at 0.0 °C. The prodrug has reduced hydrogen‐bonding capability compared with the unmodified structure of ampicillin. Evaluation of the logP parameter (the octanol/water partition coefficient) indicates that the ampicillin prodrug (logP=1.773) has increased lipophilic characteristics relative to the unmodified ampicillin structure (logP=1.06). The lipophilic substituent constant for the esterification of the carboxylic acid is 0.713, a positive value which indicates that the substituent has a lipophilic nature. The ampicillin prodrug was solubilized into LB agarose media at a concentration of 0.228 mg/ml, and was found to induce 100% growth inhibition of an ampicillin‐susceptible and streptomycin‐resistant Escherichia coli strain (designated DH1), and induced greater than 30% growth inhibition of an ampicillin‐resistant E. coli strain (designated PKK). Synthesis of this prodrug utilizing a diazoalkane was highly efficient, with no undesirable by‐products being formed.

Sulfonamide Agents for Treatment of Staphylococcus MRSA and MSSA Infections of the Central Nervous System

For drugs that target the central nervous system (CNS) the penetration of the blood-brain barrier (BBB) is a necessity, whereas for other drugs it is an unwanted side-effect. Infections by Staphylococcus aureus (SA) of the CNS causes brain abscesses and inflammation of the tissue enclosing the brain and spinal cord. In this study four sulfonamide agents are compared to streptomycin for growth inhibition of SA bacteria in vitro. All sulfonamide agents were synthesized using microwave irradiation techniques. All drugs tested show substantial growth inhibition of MRSA and MSSA types of Staphylococcus aureus. Important pharmacological properties of streptomycin and sulfonamide drugs are determined to evaluate the efficacy for CNS targeting in clinical treatment of MSSA and and MRSA infections. Although streptomycin demonstrated considerable inhibition of MSSA and MRSA bacteria, the Log P value of -5.28 and polar surface area (PSA) of 331.4 Angstroms(2) effectively inhibits its penetration into the CNS. The Log BB for streptomycin at -4.756 also indicates a very unlikely candidate to treat SA infections of the CNS. All sulfonamide agents showed significant growth inhibition of MRSA and MSSA while having useful values of Log P and PSA for penetrating the BBB. In addition the Log BB values for each sulfonamide agent indicates these agents will enter the CNS to halt SA infections. The Log BB values for sulfonamide drugs 2, A, B, and C are -1.658, -0.109, -0.109, and -0.243, respectively. Values for Log P were 2.549, 3.27, 3.47, and 2.63, respectively; with PSA values at 137.8 A(2), 40.98 A(2), 40.98 A(2), and 49.36 A(2), respectively. The small size of the sulfonamide agents and other physicochemical properties strongly support the contention they can be effectively applied to treat MRSA and MSSA infections of the brain and spinal cord.

Applying pattern recognition methods to analyze the molecular properties of a homologous series of nitrogen mustard agents

The purpose of this research was to analyze the pharmacological properties of a homologous series of nitrogen mustard (N-mustard) agents formed after inserting 1 to 9 methylene groups (-CH2-) between 2-N(CH2CH2Cl)2 groups. These compounds were shown to have significant correlations and associations in their properties after analysis by pattern recognition methods including hierarchical classification, cluster analysis, nonmetric multi-dimensional scaling (MDS), detrended correspondence analysis, K-means cluster analysis, discriminant analysis, and self-organizing tree algorithm (SOTA) analysis. Detrended correspondence analysis showed a linear-like association of the 9 homologs, and hierarchical classification showed that each homolog had great similarity to at least one other member of the series—as did cluster analysis using paired-group distance measure. Nonmetric multi-dimensional scaling was able to discriminate homologs 2 and 3 (by number of methylene groups) from homologs 4, 5, and 6 as a group, and from homologs 7, 8, and 9 as a group. Discriminant analysis, K-means cluster analysis, and hierarchical classification distinguished the high molecular weight homologs from low molecular weight homologs. As the number of methylene groups increased the aqueous solubility decreased, dermal permeation coefficient increased, Log P increased, molar volume increased, parachor increased, and index of refraction decreased. Application of pattern recognition methods discerned useful interrelationships within the homologous series that will determine specific and beneficial clinical applications for each homolog and methods of administration.