Ciprofloxacin (CFX) is a quinolone-based wide-spectrum antimicrobial compound used to treat several bacterial infections. It was patented in the 1980s and has since been a major drug against urinary tract infections, anthrax, STDs, and other serious infections [1]. It is highly effective against Gram-negative bacteria, with an MIC90 value of less than 1 g/L against most strains such as Hemophilus spp., Salmonella spp., Neisseria spp., etc. [2]. However, environmental contamination from pharmaceutical products like CFX is increasing at an alarming rate [[3], [4], [5]]. Recent studies showed that waterways and rivers near pharmaceutical manufacturing sites are contaminated with various drugs and chemicals [6]. The problem is not confined to industrial sites but also affects sewage released from hospitals and households, containing non-metabolized pharmaceuticals. Out of several drug compounds released into the environment, wide-spectrum antimicrobial agents like CFX, manufactured in bulk due to market demand, are likely to be found in higher concentrations in the environment. A study in 2009 found that an industrial effluent sample collected from a combination of 90 pharmaceutical manufacturers detected more than 10 drug molecules, with CFX holding the highest concentration at 6.5 mg/L [7]. These findings raise concerns about the consequences of antimicrobial drug contamination. According to a study conducted on different continents, the Antibiotic Resistance Predicted No Effect Concentration (ABR PNEC) was exceeded by CFX. The ABR PNEC suggests that a given antimicrobial agent must not exceed 100 ng/L, whereas for ecotoxicology, the PNEC is 1200 ng/L. However, CFX exceeded these limitations by 58 % and 16 %, respectively [8]. Consequently, antibiotic resistance against CFX increased, which is concerning for the health sector as it is one of the best drugs for treating severe infections. Antibiotic resistance and ecotoxicity are serious issues that need to be addressed decisively [9]. Therefore, a robust and accurate early detection system for CFX is needed.

Normally, the detection of pharmaceutical contaminants is carried out ex situ using the HPLC method [10]. Despite providing more accurate and reliable results, this method is not entirely feasible due to high operating expenses and personnel requirements, especially in underdeveloped countries where the problem is prevalent. Therefore, a portable, highly specific, and cost-effective solution is needed to detect CFX contamination. Considering these requirements, several types of low-cost biosensor solutions have been tested successfully to detect CFX [[11], [12], [13]]. Buglak and colleagues investigated the recognition patterns of antibodies against fluoroquinolones, particularly focusing on CFX and clinafloxacin as immunogens [14]. They also developed models that could predict how well these antibodies would bind to different fluoroquinolones. Similarly, Huang and colleagues developed ELISA kit that targets CFX using rabbit monoclonal antibodies. The developed ELISA kit demonstrated utility as a monitoring tool to determine the illicit addition of CFX in food products [15]. Due to their high affinity, selectivity, and cost-effectiveness, aptamer-based biosensors are other alternatives to immunosensors [16]. Aptamers are nucleotides, either DNA or RNA-based, with only one strand that can fold into a specific structural conformation [17]. Depending on the composition and sequence order, aptamers fold into different conformations, making them specific to a certain target molecule [18]. Since aptamers were introduced, the SELEX (systematic evolution of ligands by exponential enrichment) method has been widely used to assess their affinity for various target molecules, from small compounds to larger proteins. However, the SELEX process is resource-intensive in terms of time and cost [19]. It typically requires over 15 cycles to identify the optimal binding aptamer, with each cycle lasting up to 24 h [20]. Moreover, the expense of synthesizing the random sequences in the library adds to its inefficiency. Therefore, adopting newer, in silico-based methodologies could cut costs and save a lot of time. Bioinformatics tools and algorithms have revolutionized biology and drug discovery, extending their impact to the study of aptamer interactions with target molecules [21]. Aptamer docking experiments elucidate binding mechanisms, aiding in aptamer refinement. The concept of in silico aptamer maturation shows promise, particularly with the anticipated influence of artificial intelligence [22]. In our study, we employed a combined computational and experimental approach. After isolating the aptamer using computational methods, we conducted CV and EIS to validate its functionality. By immobilizing the DNA aptamer onto a gold electrode and conducting multiple readings, we confirmed biosensor stability and selectivity.