相关文章推荐
考研的馒头  ·  未定义标识符FILE_INFO_BY_HAN ...·  2 月前    · 
火星上的蚂蚁  ·  An Optimized ...·  8 月前    · 
绅士的竹笋  ·  存储常见问题_容器服务 ...·  1 年前    · 
俊逸的创口贴  ·  SQL Server。按XML列的节点排序·  1 年前    · 
Code  ›  Facile synthesis of antibiotic-functionalized gold nanoparticles for colorimetric bacterial detection - PMC
pmc atb pubmed doi
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8697705/
傻傻的大脸猫
1 年前
The .gov means it’s official. Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site. The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely. As a library, NLM provides access to scientific literature. Inclusion in an NLM database does not imply endorsement of, or agreement with, the contents by NLM or the National Institutes of Health.
Learn more: PMC Disclaimer
RSC Adv. 2021 Apr 13; 11(23): 14161–14168.
Published online 2021 Apr 15. doi: 10.1039/d1ra01316e
PMCID: PMC8697705
PMID: 35423905

Facile synthesis of antibiotic-functionalized gold nanoparticles for colorimetric bacterial detection †

Charlotte N. Elliott , María Cecilia Becerra , J. Craig Bennett , Lori Graham , M. Jazmin Silvero C. , corresponding author and Geniece L. Hallett-Tapley corresponding author

Charlotte N. Elliott

Department of Chemistry, St. Francis Xavier University, P.O. Box 5000, Antigonish, Nova Scotia Canada, ac.xfts@ttellahg

Find articles by Charlotte N. Elliott

María Cecilia Becerra

Departamento de Ciencias Farmacéuticas, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Córdoba X5000 Argentina, moc.liamg@iccungapmocnimzaj Instituto Multidisciplinario de Biología Vegetal, IMBIV, CONICET, Argentina

Find articles by María Cecilia Becerra

J. Craig Bennett

Department of Physics, Acadia University, P.O. Box 49, Wolfville, Nova Scotia Canada

Find articles by J. Craig Bennett

Lori Graham

Department of Biology, St. Francis Xavier University, P.O. Box 5000, Antigonish, Nova Scotia Canada

Find articles by Lori Graham

M. Jazmin Silvero C.

Departamento de Ciencias Farmacéuticas, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Córdoba X5000 Argentina, moc.liamg@iccungapmocnimzaj Instituto Multidisciplinario de Biología Vegetal, IMBIV, CONICET, Argentina

Find articles by M. Jazmin Silvero C.

Geniece L. Hallett-Tapley

Department of Chemistry, St. Francis Xavier University, P.O. Box 5000, Antigonish, Nova Scotia Canada, ac.xfts@ttellahg Department of Chemistry, St. Francis Xavier University, P.O. Box 5000, Antigonish, Nova Scotia Canada, ac.xfts@ttellahg Departamento de Ciencias Farmacéuticas, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Córdoba X5000 Argentina, moc.liamg@iccungapmocnimzaj Instituto Multidisciplinario de Biología Vegetal, IMBIV, CONICET, Argentina Department of Physics, Acadia University, P.O. Box 49, Wolfville, Nova Scotia Canada Department of Biology, St. Francis Xavier University, P.O. Box 5000, Antigonish, Nova Scotia Canada Created by potrace 1.16, written by Peter Selinger 2001-2019 O stretching for cephalexin and penicillin (attributed to the amide functionality of its structure) is no longer observed (Fig. S3 † ). Similar spectroscopic observations have been reported by Manelli et al. and were considered acceptable confirmation of cephalexin conjugation to the nanoparticle surface. 29 Reported 1 H NMR analysis of cepha@AuNP has shown that these colloids are comprised of an oxidized ATB form. 30 This data supports the hypothesis that the oxophilic nature of β-lactam antibiotics is responsible for Au 3+ reduction, with preferential AuNP binding to the amide group of penicillin and cephalexin, respectively.

IR frequencies corresponding to the ATB were also observed for poly@AuNP and baci@AuNP. The IR data are in agreement with previously reported values. 31,32 Poly@AuNP afforded signals corresponding to C–H stretching (2930 cm −1 ), as well as the amide C Created by potrace 1.16, written by Peter Selinger 2001-2019 O groups (amide I – 1646 cm −1 , amide II – 1525 cm −1 , amide III – 1243 cm −1 ). 33–35 Baci@AuNP afforded similar C–H and amide C Created by potrace 1.16, written by Peter Selinger 2001-2019 O stretching vibrations, as well as additional signals assigned to the C Created by potrace 1.16, written by Peter Selinger 2001-2019 C groups (1540 cm −1 ), aromatic C–C stretches (1660 cm −1 ) and C–O vibrations (1100 cm −1 ).

TEM and dynamic light scattering (DLS) were used to affirm both the size and shape of the AuNP. Representative TEM pictures of ATB@AuNP hybrids are presented in Fig. 3 and corresponding size and morphology data in Table 2 . Each ATB@AuNP colloid presented unique (and reproducible) size and shape characteristics. DLS analysis of peni@AuNP reveals the presence of two size populations, corroborated by TEM. The first group is comprised of a limited number of Au nanostructures that are irregular and polydisperse in size (60.0 ± 8.5 nm). Conversely, the second group is mainly constituted of densely populated, monodisperse, spherical AuNP (5.0 ± 0.2 nm) engulfed in an amorphous matrix ( Fig. 3 ), attributed to penicillin functionalization of the Au surface.

An external file that holds a picture, illustration, etc. Object name is d1ra01316e-f3.jpg
Representative TEM for ATB@AuNP.

DLS analysis of cepha@AuNP revealed three, distinguishable size populations. Closer examination using TEM revealed a higher fraction of these particles assume a triangular (68.5 ± 6.4 nm) or hexagonal (49.9 ± 6.1 nm) morphology, as compared to peni@AuNP. An abundant monodisperse, spherical population was also observed (3.2 ± 0.3 nm).

The TEM images obtained for polymyxin and bacitracin-functionalized AuNP were considerable different than those obtained for colloids constructed using β-lactam ATBs. TEM images obtained for baci@AuNP show a thin film of an amorphous, spherical structures (Fig. S4 † ). This unique morphological state can be attributed to nanoemulsion that forms following sonication of the bacitracin/Au 3+ solution prior to heating. 36,37 Small, crystalline particles (2.7 ± 0.7 nm) are embedded in the amorphous spheres, as well as larger hexagonal (36.3 ± 6.7 nm) and triangular (21.9 ± 5.0 nm) plates. Similar results were concluded for poly@AuNP, with the spheres encasing primarily smaller, more uniform, crystalline structures (4.7 ± 1.6 nm). Diffraction rings obtained for the encapsulated crystalline structures are consistent with the Au (111) surface. 38–40 EDS analysis of composition of these materials do show some contributions from elemental Au, as expected. A representative EDS analysis for baci@AuNP is shown in Fig. S5. † HR-TEM of the ATB/AuNP samples (Fig. S6–S9 † ) also validate the metallic particles as Au nanospecies. In all cases, HR-TEM images illustrate lattice fringes confirming the crystalline structure of the metallic nanospecies. FFT and FFT filtered images of cepha@AuNP (Fig. S6 † ) reveal diffraction spots corresponding to the Au (111) surface with fringe spacing of 2.4 Å, consistent with the spacing of (111) planes for Au. Similarly, FFT analysis of both peni@ (Fig. S7 † ) and poly@AuNP (Fig. S8 † ) illustrate the crystalline nature of these particles, attributed with the Au (110) surface. HR-TEM of baci@AuNP (Fig. S9 † ) also presents lattice fringes indicative a highly crystalline metallic nanospecies, further corroborated by diffraction ring analysis as the Au (111) surface.

DLS analysis of the bacitracin and polymyxin-protected colloids was unable to accurately illustrate the AuNP populations observed via TEM. Particle sizes of ∼406 nm for baci@AuNP and ∼70 nm for poly@AuNP are well outside the size range of nanostructures observed by TEM. As can be seen in Fig. S4, † the observed nanoemulsion for baci@AuNP is comprised of groups of 3–5 nanodroplets ∼100 nm in diameter. Thus, DLS is most likely identifying the additive diameter of such nanodroplet cohorts, affording inflated DLS size distribution values. A schematic representation of the calculated size distribution and morphologies of each ATB@AuNP is presented in Fig. 4 .

An external file that holds a picture, illustration, etc. Object name is d1ra01316e-f4.jpg
Schematic representation of ATB@AuNP size and morphology distribution.

Assessment of colorimetric bacterial detection

The colorimetric sensor properties of the ATB@AuNP colloids were examined in the presence of three bacterial species: S. aureus (Gram-positive), P. aeruginosa (Gram-negative) and E. coli (Gram-negative). Visual and spectroscopic analysis of the ATB@AuNP colloids before and after exposure to bacteria was used to determine successful pathogen detection. Previously, absorption of prokaryotic cells has been well documented to lie outside the SPB wavelength range monitored in this work. 41–43 Ideally, upon binding to the bacteria, a visible colour change will be noted, as well as a variation in the intensity and λ max of the AuNP SPB. This colour change can be attributed to the binding of the modified ATB surface to the pathogen. Upon binding between the ATB and the bacterial cell wall, agglomeration of the AuNP will result, affording a characteristic red shift in the SPB absorption (Fig. S10 † ). These variables should remain unchanged if binding between ATB@AuNP and bacteria is not possible – most commonly if the ATB ligand is not sensitive to a specific bacterial genus. Importantly, the λ max of the SPB for all four ATB@AuNP colloids remains unchanged after 30 days of storage, showcasing the high stability of these nanocomposites. Representative UV-vis spectra for cepha@AuNP and peni@AuNP are shown in Fig. S11. †

Initial, visual inspection of the colloids following introduction of the three bacterial suspensions clearly showcases the rapid and versatile nature of ATB@AuNP ( Fig. 5 ). The interaction between the bacteria and the ATB@AuNP is expressed as (+) if a colour change is observed, while (−) is reserved for no discernable variation. The anticipated colour changes align with the bacterial sensitivity of the ATB: penicillin (Gram-positive), cephalexin (Gram-positive and some Gram-negative), bacitracin (Gram-positive), polymyxin (Gram-negative). Binding between the bacteria and ATB@AuNP decreases the interparticle distances, as compared to the unbound colloidal solutions. Such an interaction will result in colour changes akin to AuNP agglomeration, producing red-shifts and reduced absorption intensity of the SPB λ max ( Table 2 and Fig. 6 ). Given this, each bacterium possesses the capability to facilitate change of the SPB absorption in relation to its affinity for the ATB ligand. While all nanospecies may be capable of the colorimetric detection, the current focus is on the spherical nanoparticle SPB ( λ max ∼520–560 nm). Responsible for the visible pink hue of the colloids and the most largely affected by bacterial affinities. Though several of the ATB/AuNP are comprised of larger, polyhedron shaped nanospecies, SPB absorption attributed to these do not significantly comprise the UV-visible spectra presented in Fig. 6 . The triangular nanoplates, observed in both cepha and baci-modified AuNP, typically present SPB > 600 nm. 44 Similarly, hexagonal gold nanoplates, seen for peni, cepha and baci@AuNP, commonly present as largely red-shifted SPB (>700 nm). 45 While a SPB at 690 nm is observed for baci/AuNP, likely attributed to the presence of the triangular and hexagonal nanoplates, no variation in this band is observed upon bacterial exposure, while variation in the 557 nm SPB is drastically changed following bacterial coordination.

An external file that holds a picture, illustration, etc. Object name is d1ra01316e-f5.jpg
SPB colour changes of ATB@AuNP upon exposure to S. aureus , P. aeruginosa and E. coli , respectively. A positive sign (+) is indicative of a detectable colour change of the ATB@AuNP, binding of bacteria with the conjugated ATB molecule and successful bacterial detection. A negative sign (−) indicated no perceivable colour change and, thus, no bacterial interaction or detection.
An external file that holds a picture, illustration, etc. Object name is d1ra01316e-f6.jpg
UV-visible spectra illustrating positive detection of bacteria for peni@AuNP (A), cepha@AuNP (B), baci@AuNP (C) and poly@AuNP (D). S. aureus is used as an example in A, B and C, while E. coli is used in D.

Fig. 5 displays several colour variations following introduction of bacteria to the AuNP colloid. In summary, peni@AuNP exhibits a substantial colour change from pink to blue-violet following exposure to S. aureus , while no detectable change was observed upon exposure to P. aeruginosa or E. coli . A similar colour change was observed for cepha@AuNP in the presence of S. aureus and E. coli . This observation is coincident with the known antibacterial activity of penicillin G (benzylpenicillin) and cephalexin, respectively, through attachment to the penicillin-binding proteins in Gram-positive bacteria. 46

The baci@AuNP responded to S. aureus , evidenced by a tangible colour change from pink to violet. This result is most likely ascribed to the undecaprenyl pyrophosphate binding capability of bacitracin (a polypeptide antibiotic) that hinders cell wall biosynthesis, 47 a feature of some Gram-positive bacteria. On the other hand, SPB variations of poly@AuNP were detected in the presence of P. aeruginosa and E . coli. This transformation can be credited to the selective binding of polymyxin to lipopolysaccharides predominantly present in Gram-negative bacteria. 48 All three bacteria implemented in this work produce differing colorimetric patterns, enabling rapid bacterial detection in the presence of an appropriate ATB@AuNP control (in the absence of bacteria).

UV-visible spectroscopy was also used to confirm bacterial detection. As is shown in Fig. 6 , all four of the colorimetric changes perceived by the naked eye were also observed in the corresponding spectroscopic data. A red shift (shift to longer wavelengths) of the λ max of the AuNP SPB was detected concomitant with the positive visual responses. This wavelength variation can be attributed to AuNP aggregation. This experimental result further substantiates reduced interparticle distance immediately following binding of the ATB@AuNP and bacteria. Assays employing the four ATB@AuNP using lower bacteria concentrations (10 2 CFU mL −1 ; Fig. S12 † ) also afforded a decrease in absorption intensity and red-shifting of the SPB, showcasing the sensitivity and advantage of the described methodology.

To date, few studies have exploited the bacterial detection capability of pharmaceutical-conjugated nanoparticles. The current design focuses on hybrid stability and implementing ATB functional groups that binds to specific sites on the bacterial envelopes. A similar technique has been explored for magnetic nanoparticles, combining nanospecies with fluorescent molecules and vancomycin for the rapid detection low bacteria concentrations. 49 However, several drawbacks of this method include low sensitivity of the fluorescence spectroscopic analysis and the multiple step synthetic design. A complimentary report ascribes successful bacteria identification, in part, to a visible AuNP colour change (from red to blue). 50 This colour change occurs upon binding of the H2 receptors of the cysteine@AuNP and the bacterial wall. While a step in the right direction, this technique lacks selectivity given the known aptitude of cysteine to bind to a variety interfering molecules present in reaction media. This shortcoming can be addressed through the current methodology given the specificity of the ATB for specific bacteria. Combined with the well-established stability of the ATB under a number of conditions, 51 the current method offers several advantages compared to previously published routes. The purpose of this work is to ensure that the ATB protectant is able to effectively interact with bacteria upon exposure. While minor modifications of the ATB surface protectants can be expected, as evidenced by FTIR, the primary binding modes of biological molecules (van der Waals and coulombic) 20 are not favourable to significant structural variations. Furthermore, the positive bacterial detection suggests no substantial changes to the active portion of the ATB have occurred throughout the synthetic procedure. However, limitations for the proposed method have yet to be fully recognized. Future directions should be focused on the detection of other strains of clinical importance, such as Mycobacterium – a bacterium that experiences slow growth in vitro . Also, the possibility of false positives due to nanoparticle aggregation in a myriad biological fluids or food liquids should be take into account.

Conclusions

The current contribution describes the straightforward and efficient synthesis of AuNP functionalized with a series of cost-effective and commercial antibiotics: penicillin, cephalexin, bacitracin and polymyxin. Care was taken to include ATB that encompass common strains of both Gram-positive ( S. aureus ) and Gram-negative ( P. aeruginosa and E. coli ) bacteria. The ATB@AuNP synthesis was monitored using a number of experimental techniques. UV-visible spectroscopy clearly showcases effective synthesis of the ATB functionalized AuNP, with surface plasmon bands associated with the nanospecies appearing between 520–550 nm. The size of the gold nanospecies was corroborated through the use of DLS and TEM, while the identity of the metallic nanospecies was determined through a cooperative study using HR-TEM, diffraction patterning and EDS. Closer analysis of the nanostructures revealed the formation of multi-population AuNP colloids, with penicillin-, cephalexin- and bacitracin-coordination facilitating small quantities of larger, polyhedral particles (hexagonal, triangular). However, the major AuNP population constituted small, monodisperse spherical AuNP on the average of 2–5 nm. Polymyxin-doped AuNP afforded a singular particle population, spherical and monodisperse, on the order of 4 nm. Baci@ and poly@AuNP colloids also presented as spherical amorphous structures, a result of antibiotic emulsification, which were found to encapsulate the observed gold nanostructures.

UV-visible data illustrated considerable deviations of the SPB absorption wavelength and intensity, but could also be detected by the naked eye, without an instrument. These observations support the capacity of ATB@AuNP to selectively detect a series of bacterial contaminants using a simple colorimetric technique observable with the naked eye. Given this, development of this technique will continue to be explored, with focus on expansion towards a more portable and higher through-put detection methods ( i.e. , reaction test strips).

Author contributions

Manuscript preparation, review, and redaction: M. J. Silvero, M. C. Becerra, L. Graham, G. L. Hallett-Tapley; experimental conception: M. J. Silvero; formal analysis and investigation: M. J. Silvero, C. N. Elliott, L. Graham, J. C. Bennett, G. L. Hallett-Tapley; supervision: G. L. Hallett-Tapley, M. J. Silvero; funding acquisition and encouraging to the research: G. L. Hallett-Tapley. All authors have approved the final version of the manuscript.

Conflicts of interest

The authors declare no conflict of interest.

Acknowledgments

CNE would like to thank the Scotia Scholar Undergraduate Research Awards (ResearchNS) for financial support. MCB would like to thank SecyT for financial support (2018–2021) MJS would like to thank the W. F. Visiting James Research Chair Fund and StFX University for financial support. MJS and MCB are members of CONICET (IMBIV). GHT would like to acknowledge the Natural Sciences and Engineering Research Council of Canada Discovery Grant Program (NSERC-DG), the Canada Foundation for Innovation (CFI), ResearchNS and the StFX University Council for Research (UCR) for gracious financial support.

Notes

† Electronic supplementary information (ESI) available: Experimental design, photographs of ATB@AuNP colloids, TEM, HR-TEM, stability and bacterial concentration UV-visible data, diffraction patterning and EDS images, equations used in the approximation of [AuNP]. See DOI: 10.1039/d1ra01316e

Notes and references

Articles from RSC Advances are provided here courtesy of Royal Society of Chemistry
 
推荐文章
考研的馒头  ·  未定义标识符FILE_INFO_BY_HANDLE_CLASS怎么解决？ - Microsoft Q&A
2 月前
火星上的蚂蚁  ·  An Optimized Single-Molecule Pull-Down Assay for Quantification of Protein Phosphorylation (Scientif
8 月前
绅士的竹笋  ·  存储常见问题_容器服务 Kubernetes 版 ACK-阿里云帮助中心
1 年前
俊逸的创口贴  ·  SQL Server。按XML列的节点排序
1 年前
今天看啥   ·   Py中国   ·   codingpro   ·   藏经阁   ·   小百科   ·   link之家   ·   卧龙AI搜索
删除内容请联系邮箱 2879853325@qq.com
Code - 代码工具平台
© 2024 ~ 沪ICP备11025650号