User:Klare Lazor/Notebook/Chem-496-001/2011/09/07

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Objective

The purpose of this experiment is to determine the ideal pH for the synthesis of Au nanoparticles. For this experiment, Au nanoparticles will be created by reducing HAuCl4 with bovine serum albumen (BSA) in two aqueous solutions. One with a pH of 7.55 and another with a pH of 7(water). Furthermore, the temperature range will be between 74-82 degrees Celsius.

Description

Protocol: Materials needed included Chloroauric Acid (HAuCl4), bovine serum albumen (BSA), and zein protein. BSA(0.0015-0.015mM) and HAuCl4(0.25-10mM) were used to create 10mL aqueous mixtures. Each mixture was transferred to a screw-capped glass bottle. The mixtures were kept in a water thermostat bath a 40, 60, or 80±1˚C for 6 hours. The solutions changed from colorless to pink-purple and finally purple. The samples were then removed from the bath and left to cool overnight. Next, they were purified with distilled water two times in order to remove any excess BSA, using a centrifuge at 10,000-14,000rpm for five minutes. The samples were placed in different temperature baths to observe how temperature affects the synthesis of the gold nanoparticles. In addition, in order to determine the absorbance caused by surface plasmo resonance, UV-visible spectra were taken of each sample using an UV Spectrophotometer with a wavelength range of 200-900nm. Time dependent scans were also taken to collect data on the growth kinetics of AU nanoparticles. Other data was collected using a pH meter, SEM, TEM, X-ray diffraction and AFM. Last, zein protein film formation was carried out using the BSA conjugated nanoparticles. A clear solution of zein (10%w/v) in aqueous ethanol (90%v/v) along with glycerol (30% on zein weight basis) was prepared. Then, the BSA conjugated nanoparticles (10%v/v) was added into this solution. Five grams of this solution was placed in a petri dish and swirled to coat the dish. It was placed in an 40˚C oven for 24 hours to eventually lead to protein film formation. A texture analyzer was then used in order to determine the mechanical properties of the films containing different samples of BSA conjugated Au nanoparticles. (“Protein Films of Bovine Serum Albumen Conjugated Gold Nanoparticles: A Synthetic Route from Bioconjugated Nanoparticles to Biodegrable Protein Films” by Mandeep Singh Bakshi, Harpreet Kaur, Poonam Khullar, Tarlok Singh Banipa, Gurinder Kaur, and Narpinder Singh).

Procedure: In a screw-capped glass bottle, 10mL of BSA(1.5mM) and HAuCl4(0.25mM) were added to 10mL of TRIS that was titrated to a pH of 7.55. The TRIS was already titrated by instructor. A UV-visible spectra was taken of the solution with a wavelength of 200-800nm, as well as of the buffer for a standard. The solution was then placed in a 74-82˚C water thermostat bath for 30 minute intervals. After each interval, a UV-visible spectra was taken of the solution to determine the absorbance caused by the surface plasmo resonance. This was repeated again, however the aqueous solution was water. Stock solutions were created by the instructor.

Data

Calculations: Were calculated by the professor. Similar calculations can be found in notebook from last week.

  • BSA: 0.0015M x 66.76g/mol = 0.10014g/L, so 0.10014g/L x 1L/1000mL x 10mL = 0.0010014 grams of BSA
  • HAuCl4: 0.0025M x 339.7g/mol= 0.084945 g/L, so 0.084945g/L x 1L/1000mL x 10mL = 0.00084945 grams of HAuCl4
  • Aqueous solution #1: 50mM pH 7.55 Tris (prepared by instructor) 10mL
  • Aqueous solution # 2: water 10mL

stock concentrations: chloric acid:2.5mM and bsa:15 micro molar

new concentrations: Chloric acid: .25mM and bsa: 1.5 micro molar


Data Tables:

Image 1: Corrected data plotted at Time (minutes) vs. Absorbance at 550nm for aqueous solution with a pH of 7.55.


Image 2: Corrected data plotted at Wavelength (nm) vs. Absorbance for aqueous solution with pH of 7.55. Series 1: T=0min, series 2: T=30min, series 3: T= 60 min, series 4: T= 90 min, and series 5: T=120 min.

Image 3: Corrected data plotted at Wavelength (nm) vs. Absorbance for aqueous solution with pH of 7.55 zoomed. Series 1: T=0min, series 2: T=30min, series 3: T= 60 min, series 4: T= 90 min, and series 5: T=120min.

Image 4: Corrected data plotted at Time (minutes) vs. Absorbance at 550nm for aqueous solution with pH of 7 (water).


Image 5: Corrected data plotted at Wavelength (nm) vs. Absorbance for aqueous solution with pH of 7 (water). Series 1: T=0min, series 2: T=30min, series 3: T= 60 min, series 4: T= 90 min, and series 5: T=120min.


Image 6: Corrected data plotted at Wavelength (nm) vs. Absorbance for aqueous solution with pH of 7 (water). Series 1: T=0min, series 2: T=30min, series 3: T= 60 min, series 4: T= 90 min, and series 5: T=120min.

Notes

Klare, what do you make of your data? What do you think it means? Is this consistent with what Bashki et al see in their paper for nanoparticle synthesis? Matt Hartings 12:17, 14 September 2011 (EDT)

Contents of Aqueous Solutions Actual Amounts

  • Aqueous Solution #1: 8.15mL of TRIS, 1mL of BSA, and 1 mL of HAuCl4
  • Aqueous Solution #2: 8.12mL of Water, 1mL of BSA, and 1 mL of HAuCl4

Results

  • Aqueous Solution # 1: As observed in the previous lab, solutions with a high pH seem to be poor conditions for the synthesis of gold nanoparticles. Based off of our data, no nanoparticles were produced because there was no significant peak between 520 to 550nm. Colloidal Au NPs usually show absorbance at this wavelength becuase of SPR. I was actually surprised becuase I thought the increase in concentration of BSA would compensate for the high pH, apparently not. Our data was consistent with Bashki, becuase according to Bashki, nanoparticles will most likely be produced at a pH lower than the isoelectric point, pH=4.7. In addition, there seemed to be some error in our data for solution 1. The absorbance seemed to have some error at t=60 minutes. This may be due to the fact that the solution was left to cool to long, or possibility of so foreign material being present in the cubet, ie: fuzz (which we removed).
  • Aqueous Solution # 2: To my surprise, nanoparticles were formed at a pH of 7 between the time frame of 60min-120min. There was a significant peak or change in data at 550nm. I believe this is due to the increase of BSA concentration. If we would of have used the BSA concentration from previous lab, I believe we would not have had similar results. pH of 7 is still greatly above the isoelectric point, but as Bashki stated in artilce the amount of BSA has an affect on whether or not the Au NPs would be produced. According to Bashki, "an increase in BSA at fixed HAuCl4 expecially at 80 degrees Celcius ... facilitate the reduction and simultaneous entrapment of Au NPs in unfolded BSA."

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