Glass capillary column chromatography of mixed derivatives of primary prostaglandins, 6-keto prostaglandin F1α and thromboxane B2

This work describes the use of glass capillary columns (GCC) in the rapid concurrent analysis of primary prostaglandins (PGs) (e. g. PGE₂, PGE₂, PGF_2α) and other functionally significant metabolites of arachidonic acid (AA) such as TXB₂ and 6-keto PGF_1α. The use of a new multistep mixed derivatization approach that generates the methyl esters of n-butylboronate, pentafluorobenzyloxime, trimethylsilyl ether derivatives of these compounds remarkably simplifies the GC profiling of the three main pathways of AA metabolism (PGs, thromboxane, and prostacyclin). Furthermore, isomeric species giving very similar or identical mass spectral patterns can be easily identified by their relative retention times on high efficiency capillary columns.     

Determination of uranium in marine sediment pore waters by isotope dilution inductively coupled plasma mass spectrometry

Inductively coupled plasma mass spectrometry (ICP-MS) was used in an isotope dilution mode to assay small-volume (0.25 ml) sediment pore waters for their uranium contents, using ²³⁶U as the spike. The only pretreatment required was a simple dilution by a factor of 20, which gave sufficient volume for three replicate analyses per sample. Rapid and accurate results were obtained for a variety of samples and standards, ranging in concentration from 0.05 to 10 ng U ml^?1. A suite of 30 samples can be analysed in less than 6 h by this method. The relative standard deviation was better than 1.9%, with a detection limit, based on 3σ background, of 2 pg U ml^?1 in solution (40 pg ml^?1 in samples). Sea water is a difficult matrix for ICP-MS and thus the method is generally suitable for uranium determinations in many other sample solutions.     

Nano-LC-MS/MS for the quantitation of prostanoids in immune cells

Prostanoids, derivatives of arachidonic acid, are involved in inflammation and immune reactions. To understand the role of prostanoids produced by diverse immune cells, a highly sensitive quantitation method for prostaglandin E₂ (PGE₂), prostaglandin D₂ (PGD₂), 6-keto prostaglandin F_1α (6-keto PGF_1α), prostaglandin F_2α (PGF_2α), and thromboxane B₂ (TXB₂) by means of nano-liquid chromatography-tandem mass spectrometry has been developed. It was validated according to the guidelines of the Food and Drug Administration (FDA) in terms of linearity, precision, accuracy, recovery, stability, and lower limit of quantitation (LLOQ). The LLOQ were 25 pg/mL in the injected solution (75 fg on column (o.c.)) for PGE₂ and PGD₂ and 37.5 pg/mL (112.5 fg on column) for 6-keto PGF_1α, PGF_2α, and TXB₂, respectively. It was successfully applied to murine mast cells isolated from paws after zymosan injection and to CD4⁺ and CD8⁺ T lymphocytes from blood of sensitized versus non-sensitized mice in context of a delayed type hypersensitivity model. About 5,000 (T cells) to 40,000 (mast cells) cells were sufficient for quantitation. In the mast cells, the production of PGE₂ increased at a significantly higher extent than the synthesis of the other prostanoids. The T lymphocytes did not show any difference in prostanoid production, no matter whether they were obtained from sensitized mice or non-sensitized mice.     

Fluorogenic reaction between adenine derivatives and chloroacetaldehyde and its application to the determination of 9-(2-chloro-6-fluorobenzyl)adenine in human plasma

A sensitive (1 ng ml^?1) liquid chromatographic method with fluorescence detection is described for the determination of arprinocid and analogous compounds in human plasma. The method is based on chemical derivatization with chloroacetaldehyde to form highly fluorescent derivatives. Extraction of the drug from plasma and separation of the derivative from the reaction mixture are accomplished by solid-phase extraction with two silica cartridges. The effects of pH, solvent, and concentration of the reagents on the efficiency of derivatization were studied. The assay in plasma was validated in the 1–50 ng ml^?1 range. The fluorescent derivatives were synthesized and fully characterized. The derivatives were also found to be efficient as energy acceptors in the oxalate ester/hydrogen peroxide chemiluminescent system.     

Determination of ultra-trace levels of cobalt by ion chromatographic separation and chemiluminescence detection

A luminol chemiluminescence detection/flow injection analysis technique coupled with ion chromatography (IC) has been examined for the selective determination of cobalt (II) at pg ml^?1 levels. A barium chloride solution was used as an eluent in the IC to separate cobalt(II) from interferents. When a 100-μ1 sample injection volume was used, the detection limit was 1.0 pg ml^?1 cobalt; the minimum detectable amount of cobalt was 100 fg. The calibration graph was linear above 10 pg ml^?1 and the linear dynamic range extended over six orders of magnitude. The relative standard deviation for ten replicate measurements of 30 pg ml^?1 cobalt was 3.8%. The results of the analysis of a synthetic sample corresponding to a boiling-water reactor coolant and some commercially available copper(II) standard solutions are given.     

Capillary electrophoresis‐laser‐induced fluorescence detection of rat brain catecholamines with microwave‐assisted derivatization

In this study, a rapid and sensitive method is described for the catecholamines detection in rat brain. CE with LIF detection for the determination of FITC derivatized catecholamines (dopamine, epinephrine, and norepinephrine) was demonstrated. Conventional water bath and microwave‐assisted derivatization methods were employed and a significant reduction in the derivatization time from 2 h for the conventional water bath at room temperature (ca. 25°C) to 2 min for the microwave‐assisted derivatization was achieved. Online sample concentration of field‐amplified sample stacking (FASS) method was employed to achieve higher sensitivities (the detection limits obtained in the normal injection mode ranged from 2.6 to 4.5 ng L^?1 and in the FASS mode ranged from 22 to 34 pg L^?1). Furthermore, this microwave‐assisted derivatization CE–LIF method successfully determined catecholamines in rat brain with as low as 100 ng L^?1 (FASS mode) to 10 μg L^?1 (normal injection mode). This CE–LIF method provided better detection ability when compared to the best reports on catecholamines analyses.     

Evaluation of electrochemical hydride generation for the determination of arsenic and selenium in sea water by graphite furnace atomic absorption with in situ concentration

A continuous flow electrochemical hydride generation technique coupled with in situ concentration in a graphite furnace has been developed for determination of As and Se in seawater. Lead is used as cathode material for the production of arsine and hydrogen selenide. The efficiency of generation of arsine from As(III) is 86 ± 6%, that from As(V) ranges from 73% to 86%. The efficiency of generation of hydrogen selenide from Se(IV) is 60 ± 5% and from Se(VI) is 30 ± 5%. The hydrides are trapped in an iridium-palladium coated graphite furnace prior to atomization. Absolute detection limits and concentration detection limits of 84 pg (3s_blank) and 84 pg ml⁻¹ for determination of As using 1 ml sample volume and 75 pg (3s_blank) and 7.5 pg ml⁻¹ for determination of Se using 10 ml sample volumes are obtained, respectively. The precision of replicate measurement for the analysis of reference materials at the 1.3 μg l⁻¹ level for As(III) (0.8 ng absolute mass level) and at the 0.042 μg l⁻¹ level for Se(IV) (0.42 ng absolute mass level) is better than 4% and 23% (relative standard deviation, RSD), respectively. The RSD values quoted above for Se include errors introduced by the sample preparation procedure.     

Mass spectra of halogenated esters 5—Chloromethyl esters of aliphatic C2?C12 n-carboxylic acids and their monochlorinated derivatives

A study has been made of the mass spectral fragmentation upon electron impact of aliphatic C₂? C₁₂ chloromethyl esters and all their 66 monochlorinated derivatives. The fragmentation pathways of the parent chloromethyl esters were elucidated with the aid of the 1st FFR metastable ions. A McLafferty rearrangement gives the base peak in the C₆? C₁₁ parent esters and in almost all the 4-chloro and ω-chloro isomers. The subsequent loss of HCl gives a very characteristic peak of the chloromethyl esters and their (3-ω)-chloro derivatives at m/z 72, [C₃H₄O₂]⁺. The 2-chloro isomers have the corresponding chlorine-containing fragment ion at m/z 106/108. The mass spectra of 2-, 3-, 4-, 5- and ω-chloro isomers give the characteristic fragment ions, the mass spectra of the other isomers being very similar.     

Reverse-FIA with spectrophotometric detection method for determination of vitamin C

Simple reverse flow injection analysis (rFIA) manifold with spectrophotometric detection was developed for an indirect determination of ascorbic acid. Parameters such as stability, accuracy and precision were established for the method and evaluated statistically to assess the applications of the method. Ascorbic acid in this procedure accelerates dediazoniation reaction of formed diazonium ions; hence its quantity can be determined by monitoring the derivatization product from coupling unreacted diazonium ion with phenol to give an azo dye (coupling reaction). The rFIA design was based on the injection of sodium nitrite into an acidic p-aminoacetophenon carrier stream in which diazonium ion was formed. This ion was inhibited by ascorbic acid stream before coupling with the phenol-Na₂CO₃ stream. Under optimum conditions, ascorbic acid acts in accordance with the Beer??s law at two concentration ranges 0.4?C6.5 ??g ml^?1 (R = 0.9995) and 7.0?C20.0 ??g ml^?1 (R = 0.9949), with detection limits of 0.25 ??g ml^?1. The developed method was applied to the determination of vitamin C in pharmaceutical formulations which produced satisfactory results compared with the standard methods reported in the British Pharmacopoeia.     

Polarography of disodium pentacyanonitrosylferrate(II): Part 2. Determination at Therapeutic Levels in Serum,Plasma, and Whole Blood

Disodium pentacyanonitrosylferrat(II) (sodium nitroprusside) is determined at therapeutic (ng ml^?1) levels in plasma, serum and blood with conventional and high-performance differential pulse polarography (d.p.p. and h.p.d.p.p.) at a dropping mercury electrode or a static mercury drop electrode. Serum or plasma (3 ml) is treated with perchloric acid containing 1 mg ml^?1 potassium hexacyanoferrate(II), centrifuged for 10 min and subjected to polarography. For spiked serum, calibration graphs are linear over the range 30–1000 ng ml^?1 sodium nitroprusside, regardless of the polarographic technique; the estimated detection limit is 15 ng ml^?1 (5 × 10^?8 M). Calculated therapeutic levels range from 100 to 1000 ng ml^?1. Similar results were obtained for spiked plasma. A similar procedure is suitable for whole blood and was used to study the in-vitro degradation of sodium nitroprusside (200 ng ml^?1) on incubation at 37°C. The in-vitro loss is rapid (t_{$\text{1}\text{2}$} ≈ 6 min) but meaningful in-vivo levels can be obtained when the blood is collected in a 0.9% sodium chloride solution at 0°C. Thiocyanate, the main metabolite of nitroprusside, and thiosulphate, which is a potential antidote for cyanide, do not interfere.     

An efficient method for the determination of enantiomeric purity of racemic amino acids using micellar chromatography,a green approach

A simple, sensitive and green micellar liquid chromatographic method (RP-HPLC) was developed for enantioseparation of four racemic amino acids, namely, (RS)-selenomethionine, (RS)-methionine, (RS)-cysteine and (RS)-penicillamine. An aqueous solution of sodium dodecyl sulfate and Brij-35 was prepared and used as mobile phase for HPLC analysis. Activated esters of (S)-ibuprofen, (S)-ketoprofen and (S)-levofloxacin were synthesized by reacting them with N-hydroxybenzotriazole. These esters were characterized by UV, IR, ¹HNMR, HRMS and elemental analysis. These chiral reagents (activated esters) were used for the synthesis of diastereomeric derivatives of the chosen amino acids. The diastereomeric derivatives were separated on a C₁₈ column by micellar liquid chromatography. Chromatographic conditions were optimized by varying concentration of surfactant in aqueous solution, and by varying the concentration and pH of the buffer. The green assessment score was calculated for the developed method (78, an excellent green method score). In addition, density functional theory calculations were performed, using Gaussian 09 rev. A.02 and hybrid density functional B3LYP with a 6-31G* basis set program, in order to develop lowest energy optimized structures of diastereomeric derivatives. The method was validated according to International Conference on Harmonization guidelines and the retention factor (k), selectivity factor (α), resolution factor (R_S) and limit of detection (0.295 ng ml⁻¹) and limit of quantification (0.896 ng ml⁻¹) were calculated.     

GC-MS Discrimination of Citrulline from Ornithine and Homocitrulline from Lysine by Chemical Derivatization: Evidence of Formation of N5-Carboxy-ornithine and N6-Carboxy-lysine

Derivatization of amino acids by 2 M HCl/CH₃OH (60 min, 80 °C) followed by derivatization of the intermediate methyl esters with pentafluoropropionic anhydride (PFPA) in ethyl acetate (30 min, 65 °C) is a useful two-step derivatization procedure (procedure A) for their quantitative measurement in biological samples by gas chromatography-mass spectrometry (GC-MS) as methyl ester pentafluoropropionic (PFP) derivatives, (Me)_m-(PFP)_n. This procedure allows in situ preparation of trideutero-methyl esters PFP derivatives, (d₃Me)_m-(PFP)_n, from synthetic amino acids and 2 M HCl/CD₃OD for use as internal standards. However, procedure A converts citrulline (Cit) to ornithine (Orn) and homocitrulline (hCit) to lysine (Lys) due to the instability of their carbamide groups under the acidic conditions of the esterification step. In the present study, we investigated whether reversing the order of the two-step derivatization may allow discrimination and simultaneous analysis of these amino acids. Pentafluoropropionylation (30 min, 65 °C) and subsequent methyl esterification (30 min, 80 °C), i.e., procedure B, of Cit resulted in the formation of six open and cyclic reaction products. The most abundant product is likely to be N⁵-Carboxy-Orn. The second most abundant product was confirmed to be Orn. The most abundant reaction product of hCit was confirmed to be Lys, with the minor reaction product likely being N⁶-Carboxy-Lys. Mechanisms are proposed for the formation of the reaction products of Cit and hCit via procedure B. It is assumed that at the first derivatization step, amino acids form (N,O)-PFP derivatives including mixed anhydrides. At the second derivatization step, the Cit-(PFP)₄ and hCit-(PFP)₄ are esterified on their C¹-Carboxylic groups and on their activated N^ureido groups. Procedure B also allows in situ preparation of (d₃Me)_m-(PFP)_n from synthetic amino acids for use as internal standards. It is demonstrated that the derivatization procedure B enables discrimination between Cit and Orn, and between hCit and Lys. The utility of procedure B to measure simultaneously these amino acids in biological samples such as plasma and urine remains to be demonstrated. Further work is required to optimize the derivatization conditions of procedure B for biological amino acids.     

Colorimetric Determination of Propofol in Bulk form,Dosage Form and Biological Fluids

ABSTRACT

Propofol is coupled with 2, 6-dichloroquinone-4-chlorimide (DCQ) in a reaction buffered at pH 9.6 to give a colored product having an analytically useful maximum at 635 nm. The factors affecting the color generation were optimized and incorporated in the procedure. The reacted propofol has a molar absorptivity of 3.9 × 10^?4 L mol^?1 cm^?1, and Beer's law is obeyed for concentrations 1-5 μg ml^?1 with detection limit 0.25 μg ml^?1. The method was found applicable to biological fluids (plasma and urine) spiked with propofol at concentration levels 1-5 μg ml^?1 for plasma and 1-5 μg 0.5 ml^?1 urine (less sensitivity is obtained with urine volumes above 0.5 ml) with detection limits 0.28 μg ml^?1 for plasma and 0.4 μg 0.5 ml^?1 urine. The average recovery for the commercial preparation (1% w/v propofol emulsion intravenous injection for infusion) was 99.54% with an RSD of 1.05%. The method was validated by an adopted HPLC method. The results obtained by the HPLC method for the commercial preparation were statistically compared with the proposed method and evaluated at the 95% confidence limits.     

Magnetic electrochemiluminescent Fe3O4/CdSe-CdS nanoparticle/polyelectrolyte nanocomposite for highly efficient immunosensing of a cancer biomarker

Magnetic electrochemiluminescent Fe₃O₄/CdSe–CdS nanoparticle/polyelectrolyte nanostructures have been synthesized and used to fabricate an electrochemiluminescence (ECL) immunosensor for the detection of carcinoembryonic antigen (CEA). CEA is a protein used as a biomarker for several cancers; particularly, to monitor response to treatment in colon and rectal cancer patients. The nanocomposites can be easily separated and firmly attached to an electrode owing to their excellent magnetic properties. This represents a promising advantage for bioassay applications. More importantly, the nanostructures exhibit intense and stable ECL emissions in neutral solution, which makes them ideal for ECL immunosensing. The 3‐aminopropyltriethoxysilane (APS) polyelectrolyte shell on the nanostructure surface not only enhances the intensity and stability of the ECL signal, but also acts as a crosslinker for immunosensor fabrication. A CEA antibody immobilized onto a nanocomposite/APS/electrode with gold nanoparticles comprises the ECL immunosensor. The principle of ECL detection for CEA is based on a change in steric hindrance after immunoreaction, which leads to a decrease in ECL intensity. A wide detection range (0.064 pg ml^?1–10 ng ml^?1) and low detection limit (0.032 pg ml^?1) are achieved. The immunosensor is highly sensitive and selective, and exhibits excellent stability and good reproducibility. It thus has great potential for clinical protein detection. In particular, this approach uses a novel class of bifunctional nanocomposites that display both intense ECL and excellent magnetism, which renders them suitable for a large range of bioassay applications.     

Dual‐column capillary microextraction (CME) combined with electrothermal vaporization inductively coupled plasma mass spectrometry (ETV‐ICP‐MS) for the speciation of arsenic in human hair extracts

In this work, dual‐column capillary microextraction (CME) system consisting of N‐(2‐aminoethyl)‐3‐aminopropyltrimethoxysilane (AAPTS)‐silica coated capillary (C1) and 3‐mercaptopropyl trimethoxysilane (MPTS)‐silica coated capillary (C2) was developed for sequential separation/preconcentration of arsenite [As(III)], arsenate [As(V)], monomethylarsonic acid [MMA(V)] and dimethylarsinic acid [DMA(V)] in the extracts of human hair followed by electrothermal vaporization inductively coupled plasma mass spectrometry (ETV‐ICP‐MS) detection with iridium as permanent modifier. Various experimental parameters affecting the dual‐column microextraction of different As species had been investigated in detail. It was found that at pH 9, As(V) and MMA could be quantitatively retained by C1 and only As(III) could be quantitatively retained by C2. With the aid of valve switching, As(V)/MMA(V) retained on C1 and As(III) retained on C2 could be sequentially desorbed by 10 µl of 0.01 mol l^?1 HNO₃ [for As(V)], 0.1 mol l^?1 HNO₃ [for MMA(V)] and 0.2 mol l^?1 HNO₃‐3% thiourea (m/v) [for As(III)], respectively, the eluents were immediately introduced into the Ir‐coated graphite tubes for further ETV‐ICP‐MS detection. With two‐step ETV pyrolysis program, Cl^? in the sample matrix could be in situ removed, and the total As in the human hair extracts or digested solution could be interference‐free, determined by ETV‐ICP‐MS. DMA(V) in the human hair extracts was obtained by subtraction of total As in the human hair extracts from other three As species. Under the optimized conditions, the detection limits (3 σ) of the method were 3.9 pg ml^?1 for As(III), 2.7 pg ml^?1 for As(V), 2.6 pg ml^?1 for MMA(V) and 124 pg ml^?1 for total As with the relative standard deviations less than 7.0% (C = 0.1 ng ml^?1, n = 7), and the enrichment factor was 286, 262 and 260 for As(III), As(V) and MMA(V), respectively. The developed method was successfully applied for the speciation of arsenic in the extracts of human hair. Copyright © 2009 John Wiley & Sons, Ltd.     

Electroanalytical Determination of Carbaryl in Natural Waters on Boron Doped Diamond Electrode

The anodic voltammetric behavior of carbaryl on a boron‐doped diamond electrode in aqueous solution is reported. The results, obtained by square‐wave voltammetry at 0.1 mol L^?1 Na₂SO₄ and pH 6.0, allow the development of a method to determine carbaryl, without any previous step of extraction, clean‐up, preconcentration or derivatization, in the range 2.5–30.0×10^?6 mol L^?1, with a detection limit of 8.2±0.2 μg L^?1 in pure water. The analytical sensitivity of this electrochemical method diminished slightly, from 3.07 mA mmol^?1 L to 2.90 mA mmol^?1L, when the electrolyte was prepared with water samples collected from two polluted points in an urban creek. In these conditions, the recovery efficiencies obtained were around 104%. The effect of other pesticides (fenthion and 4‐nitrophenol) was evaluated and found to exert a negligible influence on carbaryl determination. The square‐wave voltammetric data obtained for carbaryl were typical of an irreversible electrode process with mass transport control. The combination of square‐wave voltammetry and diamond electrodes is an interesting and desirable alternative for analytical determinations.     

Total synthesis of PGF2α and 6,15-diketo-PGF1α and formal synthesis of 6-keto-PGF1α via three-component coupling

The asymmetric total synthesis of PGF_2α and 6,15-diketo-PGF_1α and formal synthesis of 6-keto-PGF_1α from a common key intermediate are described. The key intermediate, which has a chiral cyclopentane backbone possessing suitable functional groups with required stereochemistry for both side chains, was prepared from (R)-4-silyloxy-2-cyclopentenone through a three-component coupling reaction. The Wittig reaction, Nozaki-Hiyama-Kishi (NHK) coupling and cross metathesis completed the synthesis of PGF_2α, 6,15-diketo-PGF_1α and 6-keto-PGF_1α.     

Determination of Rhodium in Biological Materials by Electrothermal Atomic Absorption Spectrometry with a Tungsten Tube Atomizer

Abstract

Electrothermal atomic absorption spectrometry (ETAAS) of rhodium with a tungsten tube atomizer has been investigated under optimum conditions (atomization temperature; 2230 C, purge gas; Ar 480 ml min^?1 + H₂ 20 ml min^?1, and pyrolysis temperature; 590 C). The absolute characteristic mass (the mass of element giving 0.0044 abs.) of rhodium by the atomizer was 86.5 pg and the detection limit was 16.5 ng ml^?1 (3S/N). The interferences caused by large amounts of interferents were evaluated. Al, Ca, Cu, Fe, K, Mg, Na, Pb and Zn severely interfered in the AA signal of rhodium. Ammonium phosphate, ascorbic acid, palladium nitrate, copper nitrate, lanthanum nitrate, thiocyanate and thiourea, well known as matrix modifiers were tested to eliminate the severe interferences. However, by the addition of these compounds, the rhodium signal was not recovered. The standard addition method was adapted for the determination of rhodium in biological materials. The recovery of spiked-rhodium in biological materials was in the range of 97.4 to 107%.     

Combined use of cyclofructans and an amino acid ester‐based ionic liquid for the enantioseparation of huperzine A and coumarin derivatives in CE

Cyclofructans (CFs) and their derivatives have recently been proven to be efficient chiral selectors (CSs) for the enantioseparation of several analytes in CE, HPLC, and GC. In this study, the chiral separation ability of a number of native and derivatized CFs was examined in CE. Particularly, six different CFs, with different derivatization groups and cavity sizes [native CF‐6 and CF‐7, isopropyl cyclofructan‐6 (IPCF‐6), IPCF‐7, sulfated cyclofructan‐6 (SCF‐6), and SCF‐7] were used as CSs for the enantioseparation of huperzine A, warfarin, and coumachlor. Almost all of the examined CFs, except from SCF‐6 & ‐7, demonstrated relatively low and sometimes no chiral separation ability for huperzine A. In an effort to improve both resolution and efficiency, the chiral ionic liquid D‐Alanine tert butyl ester lactate (D‐AlaC₄Lac) was added into the BGE. In most of the cases, the combination of CF with D‐AlaC₄Lac resulted in an improvement in peak efficiency and/or resolution. When CF‐6 was utilized with D‐AlaC₄Lac, a resolution of 1.4 was obtained, while the use of IPCF‐6/D‐AlaC₄Lac provided a baseline enantioseparation. Although the combination of SCF‐7 and 40 mM D‐AlaC₄Lac did not affect resolution, it dramatically increased peak efficiency from 24 000 to 117 000. In the case of warfarin and coumachlor, IPCF‐6 and IPCF‐7 proved to be the most effective CSs. It is, therefore, concluded that the size of the cavity and the CF derivatization are the key parameters for the chiral separation capability. It is also clear from this study that D‐AlaC₄Lac is necessary for improved peak efficiencies and resolutions.     

Chloropyridinyl Esters of Nonsteroidal Anti-Inflammatory Agents and Related Derivatives as Potent SARS-CoV-2 3CL Protease Inhibitors

We report the design and synthesis of a series of new 5-chloropyridinyl esters of salicylic acid, ibuprofen, indomethacin, and related aromatic carboxylic acids for evaluation against SARS-CoV-2 3CL protease enzyme. These ester derivatives were synthesized using EDC in the presence of DMAP to provide various esters in good to excellent yields. Compounds are stable and purified by silica gel chromatography and characterized using ¹H-NMR, ¹³C-NMR, and mass spectral analysis. These synthetic derivatives were evaluated in our in vitro SARS-CoV-2 3CLpro inhibition assay using authentic SARS-CoV-2 3CLpro enzyme. Compounds were also evaluated in our in vitro antiviral assay using quantitative VeroE₆ cell-based assay with RNAqPCR. A number of compounds exhibited potent SARS-CoV-2 3CLpro inhibitory activity and antiviral activity. Compound 9a was the most potent inhibitor, with an enzyme IC₅₀ value of 160 nM. Compound 13b exhibited an enzyme IC₅₀ value of 4.9 µM. However, it exhibited a potent antiviral EC₅₀ value of 24 µM in VeroE6 cells. Remdesivir, an RdRp inhibitor, exhibited an antiviral EC₅₀ value of 2.4 µM in the same assay. We assessed the mode of inhibition using mass spectral analysis which suggested the formation of a covalent bond with the enzyme. To obtain molecular insight, we have created a model of compound 9a bound to SARS-CoV-2 3CLpro in the active site.