Functionalization of Gold Nanoparticles with Monosaccharide Mannose

 

Sambit Dash¹, Pragna Rao², Ullas Kamath¹, Aparna R Pai³, Prasanna Kumar Reddy Gayam⁴, Ekta Rati⁵, Shivani Karnik⁴, Alex Joseph⁵, Angel Treasa Alex⁴

¹Department of Biochemistry, Melaka Manipal Medical College,

Manipal Academy of Higher Education (MAHE), Manipal, India.

²Manipal Academy of Higher Education (MAHE), Bangalore, India.

¹Department of Biochemistry, Melaka Manipal Medical College,

Manipal Academy of Higher Education (MAHE), Manipal, India.

³Department of Neurology, Kasturba Medical College,

Manipal Academy of Higher Education (MAHE), Manipal, India.

⁴Department of Pharmaceutical Biotechnology, Manipal College of Pharmaceutical Sciences,

Manipal Academy of Higher Education (MAHE), Manipal, India.

⁵Department of Pharmaceutical Chemistry, Manipal College of Pharmaceutical Sciences,

Manipal Academy of Higher Education (MAHE), Manipal, India.

 

ABSTRACT:

Gold nanoparticles have found a wide range of application in biomedical sciences. Unique properties of these metal nanoparticles include surface plasmon resonance and size dependent colour change. Various molecules have been functionalized on the gold nanoparticles surface but carbohydrates have garnered attention due to their properties and their role in living systems. However certain challenges make carbohydrate-gold nanoparticles association difficult to obtain and stabilize. This study was carried out to chemically remodel gold nanoparticles by adding a monosaccharide mannose to its surface. A modified phase transfer method was used to synthesize gold nanoparticles. The surface of the nanoparticles was fixed with cyanuric chloride to serve as a linker. Mannose was then linked to the linker molecule. All three stages of the process, gold nanoparticles, and gold nanoparticles with linker and gold nanoparticles with the carbohydrate were analyzed for size and stability. Zeta potential and UV-vis data exhibited stable gold nanoparticles dispersion, successful binding of linker molecule as well as the carbohydrate. This study shows a simple, cost-effective and robust method of glycomodification of gold nanoparticles surface which can further find use in wide ranging applications. 

 

 

INTRODUCTION:

Gold, a soft and malleable metal, which falls under the category of transition metals, has very low chemical reactivity. But colloidal gold, unlike bulk gold, is highly reactive, the reactivity range of which generates newer application possibility.

 

Owing to the plasmonic properties of gold nanoparticle, observed as colour change during its change in size range, it has found its place in wide variety of applications like drug delivery, targeted delivery of peptides or DNA, clinical chemistry and molecular biology applications. ^1-6 

 

Be it new drug screening, understanding biological processes, clinical diagnosis and treatment, carbohydrates have various functions. Physicochemical and biomedical applications of gold nanoparticles have attracted much attention. Since surface modification to attain functionality is a mainstay in attaining the intended objective, many molecules have been added on to surface of these gold nanoparticles. Of these, attachment of carbohydrate moieties has become a largely used target owing to their molecular properties as well as their action in living systems. ^{7, 8}

 

Two broad methods, namely, physical adsorption and chemical methods can be employed to functionalize gold nanoparticles with target carbohydrate. While physical adsorption has very less output due to low adsorbability chemical methods due to poor solubility and difficulty in interaction with metal nanoparticles make it challenging to functionalize gold nanoparticles with carbohydrates. ⁹ Furthermore, cyclization of carbohydrates attached causes it to lose the property that it aims to utilize. Other methods to functionalize gold nanoparticles with carbohydrates like photochemical reaction are expensive and have low yield.

 

Cyanuric chloride based reactions which employ the several hydroxyl groups of a carbohydrate which remain free, are a set of novel methods to functionalize gold nanoparticles with carbohydrates. Cyanuric chloride by the virtue of having chlorine atoms which can be replaced by hydroxyl or amino group, acts as a good linker. These methods employ lesser steps and are sometimes ‘one pot’ synthesis processes. The aim of this study was to attach monosaccharide mannose on to gold nanoparticles to which the linker cyanuric chloride has been added. ¹⁰

 

MATERIAL AND METHODS:

 

Chemicals and reagents:

Cyanuric chloride, 11-mercaptoundecanoic acid (MUA), 11-mercapto-1-undecanol (MUOH) and HAuCl4 were obtained from Sigma Aldrich, USA. Tween 20, absolute ethanol, sodium citrate, mannose and other regents and solvents were of analytical grade and were obtained from Hi-Media Lab Pvt. Ltd, India. Aqueous solutions were all prepared with double distilled water.

 

Preparation of Mannose linked Gold Nanoparticles (GNP):

Gold nanoparticles were prepared based on a method described by Xiao Wang et. al.¹⁰  An aqueous solution of HAuCl₄ (0.25 mM, 100 mL) was used to synthesize gold nanoparticles until colour of the solution became dark purple and finally light red.⁹ To avoid aggregation, Tween 20 (0.1%, v/v) was used which was stored at 4֯C. A mixture of MUOH and MUA in ethanol solution was added to the stored nanoparticles following which a harvest of the precipitate containing MUOH/MUA-modified GNPs was made. After re-suspending the prepared gold nanoparticles, cyanuric chloride was added. Further, mannose was added to the admixture. Post centrifugation, at 12000 rpm for 20 min, the precipitate was harvested, and washed which was followed by re-suspension using sonication. The nanoparticles were characterized using particle size, zeta potential, UV-visible spectroscopy and visible colour change.^{11, 12}

 

RESULTS AND DISCUSSION:

Gold nanoparticle was synthesized by reducing the gold salt and then a modified phase transfer method was used to attach the hydroxyl group. This hydroxyl group was then replaced by linker cyanuric chloride and then the carbohydrate mannose was attached to the linker. Immobilization chemistry includes three steps. In the first step, hydroxyl terminal are created on GNPs. In the second step, cyanuric chloride is anchored onto these OH terminals to form Gold nanoparticle linked with Cyanuric chloride (GNP-CC). In the last step, the carbohydrate mannose was immobilized on the surface of cyanuric chloride layer to form GNP-CC linked with mannose (GNP-CC-Mannose).  We synthesized gold nanoparticle of the average size 43.07 nm with zeta potential of - 11.2 mV. The addition of cyanuric chloride group increased the size and an average diameter of 57.02 nm was obtained, with zeta potential of -11.2 mV. On addition of mannose, the overall size of the molecule increased to an average of 257.7 nm, with a zeta potential of -28.9 mV, exhibiting good stability (Table 1, Fig 1 and 2). The successful introduction of each of these chemical is characterized by change in the UV spectra as shown in Fig 1.

 

Table 1: Size range and zeta-potential of gold nanoparticles and remodelled gold nanoparticles

Sample No	Size (nm)	Percentage Intensity	Zeta-potential (mV)
GNP	43.07	70.2	- 11.2
GNP- CC	57.02	59.8	- 11.2
GNP-CC-Mannose	257.7	100	- 28.9

 

Figure 1: Average Particle Size distribution of GNP-CC-Mannose

 

Figure 2: Average Zeta Potential of GNP-CC-Mannose

 

Attachment of OH terminals on GNPs causes a shift in absorption from 543 nm to 545 nm, while carbohydrate attached nanoparticles exhibited a maximum absorbance at 522 nm showing a significant hypsochromic shift. This shift to lower wavelength may be due to increase in hydrophilicity of the surface layer causing a decrease in aggregation (Fig 3). There is also a significant change in the colour of nanoparticles observed during these three stages, as shown in Fig 4. The visible colour change of the gold nanoparticle and its two subsequent modifications, from a shade of red to blue via violet, indicates shift in surface plasmon absorption to lower energies.

 

Figure 3: UV spectra of GNP at various stages of functionalization. 1 - Gold nanoparticle, wavelength 543 nm. 2 - Cyanuric chloride linked gold nanoparticle, wavelength 545 nm, 3 - Gold nanoparticle with mannose attached, wavelength 522 nm)

 

Figure 4: Colour of nanoparticles observed during the three stages (i: Gold nanoparticles, ii: GNP-CC, iii: GNP-CC- mannose)

 

CONCLUSION:

Metal nanoparticles have been used as a tether for carbohydrates since sometime and it has been applied in various detection techniques. However, these interactions are often unstable and less soluble. We have been able to bind mannose successfully and have a stable chemically modified gold nanoparticle by a cost effective and quick method.  The inherent property of gold nanoparticles to exhibit colour change in congruence with size change encourages its applications in various detection techniques. Mannose tethered gold nanoparticles have been used to detect E Coli proteins, mannose receptor positive cancer tissue, etc, and has a potential to detect proteins in a carbohydrate-protein interaction.^{13, 14} Its function as a frequency amplifier can be further explored. Limitation of the study includes analysis of the modified gold nanoparticles by scanning electron microscopy.

 

ACKNOWLEDGEMENT:

The authors are grateful to the authorities of MAHE, KMC and MCOPS Manipal for the facilities.

 

CONFLICT OF INTEREST:

The authors declare no conflict of interest.

 

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Accepted on 20.02.2021           © RJPT All right reserved

Research J. Pharm. and Tech 2021; 14(12):6281-6284.