4,4'-Azobis(4-cyanovaleric acid): Stability-Indicating HPLC Analysis and Drug Delivery Applications

4,4'-Azobis(4-cyanovaleric acid), short for ACVA, CAS No. 2638-94-0. Its molecular formula is C12H16N4O4, molecular weight 280.28. It is a white crystalline powder under normal temperature and pressure, relative density is about 1.23g/cm³. 4,4'-Azobis(4-cyanovaleric acid) is an important organic compound, mainly used as a polymer initiator, can be used in polyvinyl chloride, polyacrylonitrile, polyvinyl alcohol, synthetic fiber optics and other polymers, but also can be used in plastics and synthetic rubber foaming agent. In addition, it is a commonly used water-soluble radical initiator for the preparation of bentonite clay, modification of inorganic nanoparticle-coated polymers, and as a source of free radicals in kinetic surveys, an effective photosensitizer in the near-ultraviolet region, and a scavenger of oxygen in aqueous media. In terms of safety, 4,4'-Azobis(4-cyanovaleric acid) is a hazardous material, hazard class 4.1, highly flammable, and needs to be ventilated, low-temperature and dry when stored, and transported in accordance with relevant regulations.

Development and Validation of Stability-Indicating HPLC Methods

The stability of drugs is the one of the basic requirements, which is closely related to the efficiency of pharmacological action and safety of therapy. Changes in the structure of a chemical compound and physicochemical properties may lead to a decrease in the therapeutic value of the drug through disturbances in the absorption process or a decrease in pharmacological activity or even increase in its toxicity. Therefore, the chemical stability of pharmaceutical substances is a significant problem that affects the safety and efficiency of the pharmaceutical preparation. The initiators of radical reactions are azo compounds, including 2,2′-azobisisobutyronitrile (AIBN) and 4,4'-Azobis(4-cyanovaleric acid) (ACVA), which are widely used in forced stress procedures. Although AIBN is used in organic synthesis usually at temperatures between 60 °C and 80 °C, its oxidation is carried out at a much lower temperature of 40 °C, as oxidation conditions should simulate long-term degradation and allow for the isolation of labile degradation products, such as hydroperoxides that are destroyed at higher temperatures. The aim of this study was to evaluate the oxidation of LOM and BAL under the influence of azo initiator of radical reactions of 4,4'-Azobis(4-cyanovaleric acid) and hydrogen peroxide, and to compare both of the processes.[1]

The oxidation process of BAL and LOM under the influence of an azonitrile radical initiator, i.e., ACVA at 40, 50, and 60 °C, and hydrogen peroxide were investigated. Additionally, the oxidation of BAL under the influence of KMnO4 in an acidic environment was assessed. The oxidation process was estimated while using newly developed HPLC methods that were validated. The oxidation reactions in the presence of 4,4'-Azobis(4-cyanovaleric acid), H2O2, and KMnO4 followed the kinetics of the second-order reactions. Based on the conducted research and literature data, it was found that the FQs oxidation process is the fastest as a result of KMnO4 in an acidic environment. The profile of the resulting degradation products of BAL is similar to the profile that was obtained under 4,4'-Azobis(4-cyanovaleric acid). The profiles of LOM degradation products produced under the influence of KMNO4 and ACVA were different. Whereas under the influence of hydrogen peroxide, the most frequently used in drug stability studies, only one product was formed. It seems that the use of other oxidants in stress tests, apart from H2O2, is fully justified and should be recommended due to the obtained results. In addition to the value of scientific knowledge, the research undertaken is of practical importance for the technology of the drug form, the aim of which is to obtain stable, safe, and effective drugs.

Drug Delivery System Based on 4,4′-Azobis (4-cyanovaleric Acid)

In recent decades, drug delivery system (DDS) based nanomaterials have gained numerous anticipated achievements, which have increasingly become an essential strategy for diagnosis and disease therapy in the biomedical field. Here, we designed an Fe3O4@CS nanoparticle that achieves time-dependent control of drug release by adjusting the AMF exposure time. CS-modified Fe3O4 as a drug transport carrier was firstly prepared by alkaline co-precipitation, followed by a two-step amide reaction to covalently graft drug molecular onto the Fe3O4-CS surface by using a heat-sensitive molecule 4,4'-Azobis(4-cyanovaleric acid) as a linker group. The prepared MNPs (Fe3O4@CS-NH2) allow for subsequent modification and drug loading. Then, to introduce excellent magneto-thermal response functionality, the azo compound ACVA was chosen as a thermal-sensitive switch, which is gradually being developed as a triggering agent for stimulation-responsive drug delivery systems. Here, some of the carboxylic acid of 4,4'-Azobis(4-cyanovaleric acid) firstly formed an amide bond with the prepared Fe3O4@CS-NH2 nanoparticles, and the product was named as Fe3O4@CS-ACVA. Subsequently, the remaining carboxylic acid of ACVA formed another amide bond with the amine groups on TB, and the final particles were named Fe3O4@CS-ACVA-TB.[2]

In summary, CS-modified magnetic nanoparticles (Fe3O4@CS), with a high magnetization (54.0 emu/g), were successfully synthesized by the co-precipitation method. Subsequently, the temperature-sensitive molecule 4,4'-Azobis(4-cyanovaleric acid) and the amino-based model drug toluidine blue were modified on the surface of Fe3O4@CS and were used to illustrate that the nanocarriers could control drug release. In vitro hydrothermal release studies showed that in the absence of AMF, most parts of the TB would be effectively enclosed within the nanocarriers at lower ambient temperatures (23 or 37 °C) due to the molecular bonding of ACVA, and the ACVA could remain unbroken for a longer period (at least 9 days in 37 °C). However, with the increase in incubation temperature (57 °C), the 4,4'-Azobis(4-cyanovaleric acid) would become unstable, resulting in the slow release of TB immobilized by ACVA. Moreover, the results of kinetic fitting of hydrothermal release data showed that TB released from nanoparticles followed first-order kinetics (R2 > 0.99) and the Korsemeyer–Peppas model (R2 > 0.99, n < 0.5).

References

[1]Żuromska-Witek B, Żmudzki P, Szlósarczyk M, Maślanka A, Hubicka U. Development and Validation of Stability-Indicating HPLC Methods for the Estimation of Lomefloxacin and Balofloxacin Oxidation Process under ACVA, H2O2, or KMnO4 Treatment. Kinetic Evaluation and Identification of Degradation Products by Mass Spectrometry. Molecules. 2020 Nov 11;25(22):5251. doi: 10.3390/molecules25225251. PMID: 33187198; PMCID: PMC7697971.

[2]Yin W, Nziengui Raby RB, Li Y, Li Z, Sun M, Huang Z. An Alternating Magnetic Field-Controlled Drug Delivery System Based on 4,4'-Azobis (4-cyanovaleric Acid)-Functioned Fe3O4@Chitosan Nanoparticles. Bioengineering (Basel). 2023 Jan 18;10(2):129. doi: 10.3390/bioengineering10020129. PMID: 36829623; PMCID: PMC9952477.