Method for biosynthesizing cadmium sulfide quantum dots using acidithiobacillus culture supernatant

Abstract

The present invention discloses a method for biosynthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus. The method comprises the following steps: using the supernatant of Acidithiobacillus cultured to a stable phase as a catalytic reaction system, adjusting its pH to neutral, adding a sulfur source and a cadmium source, and culturing on a shaking table to obtain cadmium sulfide quantum dots; the S source is a sulfur-containing amino acid, the Cd source is a soluble cadmium salt, and the Acidithiobacillus is a member of the genus Thiobacillus that uses inorganic sulfur as an energy source. The present invention utilizes the unique sulfur metabolism mechanism of Acidithiobacillus and can achieve the biocatalytic synthesis of a large number of water-soluble cadmium sulfide quantum dots using only the supernatant of Acidithiobacillus. Compared with other reported biosynthesis methods, the present invention has significant advantages in that it does not require the use of bacterial cells or purified proteases, is simple to operate, has mild preparation conditions, and is low in cost.

Description

Technical Field

[0001] The invention belongs to the field of semiconductor quantum dot material preparation, and particularly relates to a method for biosynthesizing cadmium sulfide quantum dots by utilizing acidithiobacillus culture supernatant. Background Art

[0002] The statements herein merely provide background information related to the present invention and do not necessarily constitute prior art.

[0003] Quantum dots (QDs) are semiconductor nanoparticles composed of a small number of atoms, with a particle size smaller than or close to the exciton Bohr radius of the corresponding semiconductor material. Common QD materials include Group II-VI semiconductor QDs, such as cadmium sulfide (CdS), cadmium selenide (CdSe), cadmium telluride (CdTe), and zinc sulfide (ZnS). By manipulating the shape, structure, and size of QDs, their band gap width, exciton binding energy, and exciton energy blue shift can be tuned. Furthermore, compared to traditional organic fluorescent dyes, QDs offer advantages such as stable fluorescence, high fluorescence intensity, and long lifetime. Consequently, the preparation methods of QDs have attracted extensive attention and research.

[0004] Quantum dots are often synthesized using chemical methods, which offer significant advantages such as controllable particle size and narrow fluorescence emission peaks. However, these methods often involve high temperatures and use highly toxic chemicals, resulting in highly toxic and biocompatible quantum dots. The aqueous synthesis of quantum dots in living organisms, however, is a promising green approach.

[0005] Existing biosynthesis methods for cadmium sulfide quantum dots involve adding thiol compounds such as thioacetamide, water-soluble cadmium salts, and cysteine to a culture medium containing mycelial pellets of Phanerochaete chrysosporium, followed by shaking and culturing. The cadmium sulfide quantum dots are then filtered, rinsed with deionized water, ultrasonically disrupted, centrifuged, and purified. These methods for synthesizing cadmium sulfide quantum dots using bacterial cells require disruption before quantum dot purification. However, cadmium sulfide quantum dots tend to adhere to disrupted cells, complicating subsequent separation and purification.

[0006] Another method uses sulfur-containing amino acids and water-soluble cadmium salts as raw materials and cystathionine gamma lyase (smCSE) protein as a biocatalyst to catalyze the synthesis of cadmium sulfide quantum dots. This operation requires heterologous expression of smCSE protein in Escherichia coli and in vitro protein separation and purification. This method requires a large amount of purified protease, which makes it difficult to achieve convenient and large-scale production of cadmium sulfide quantum dots. Summary of the Invention

[0007] In view of the shortcomings of the prior art, the present invention aims to provide a method for biosynthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus thiophanate.

[0008] In order to achieve the above object, the present invention is implemented through the following technical solutions:

[0009] A method for biosynthesizing cadmium sulfide quantum dots using the supernatant of Acidithiobacillus culture, comprising the following steps: using the supernatant of Acidithiobacillus cultured to a stable phase as a catalytic reaction system, adjusting the pH value to neutral, adding a sulfur source and a Cd source, and culturing on a shaker to obtain cadmium sulfide quantum dots;

[0010] The S source is a sulfur-containing amino acid, and the Cd source is a soluble cadmium salt;

[0011] The acidithiobacillus belongs to the genus Thiobacillus that uses inorganic sulfur as energy.

[0012] In some embodiments, the Acidithiobacillus is selected from the group consisting of Acidithiobacillus thermophilus, Acidithiobacillus sulfuroxidans, and Acidithiobacillus ferrooxidans.

[0013] In some embodiments, the medium used in the culture of Acidithiobacillus is Starkey-S 0 Liquid culture medium or Starkey-K2S4O6 liquid culture medium.

[0014] Preferably, during the cultivation of Acidithiobacillus, the pH value of the culture medium is 1-2.5.

[0015] In some embodiments, the sulfur-containing amino acid is cysteine or glutathione.

[0016] Preferably, the water-soluble cadmium salt is cadmium acetate or cadmium sulfate.

[0017] In some embodiments, the culture temperature of the shaking culture is 30-40°C.

[0018] Preferably, the shaking speed of the shaker culture is 150-180 r / min, and the culture time is 2-3 days.

[0019] In some embodiments, the pH of the Acidithiobacillus supernatant is adjusted to neutral using KOH.

[0020] In some embodiments, in the supernatant of Acidithiobacillus spp., the concentration of soluble cadmium salt is 0.3-0.8 mM, and the concentration of sulfur-containing amino acid is 3-8 mM.

[0021] The beneficial effects achieved by one or more embodiments of the present invention are as follows:

[0022] The present invention utilizes the unique sulfur metabolism mechanism of Acidithiobacillus and can achieve the biocatalytic synthesis of a large number of water-soluble cadmium sulfide quantum dots using only the supernatant of Acidithiobacillus. Compared with other reported biosynthesis methods, the present invention has obvious advantages, does not require the use of bacteria or purified proteases, is simple to operate, has mild preparation conditions, and is low in cost.

[0023] In addition, experiments have confirmed that during the conventional acidophilic culture of Acidithiobacillus, heavy metal elements such as cadmium in solid waste can be effectively leached into the culture supernatant. Therefore, based on the present invention, Acidithiobacillus can be continuously used to leach cadmium-containing supernatant, and further catalytically synthesize CdS quantum dots. It is expected that the conversion of cadmium pollution to cadmium energy (CdS quantum dots) can be achieved using the one-step method of Acidithiobacillus. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] The accompanying drawings, which constitute a part of the present invention, are used to provide a further understanding of the present invention. The exemplary embodiments of the present invention and their descriptions are used to explain the present invention and do not constitute improper limitations on the present invention.

[0025] Figure 1 Shown is a fluorescence photograph of cadmium sulfide quantum dots synthesized in the supernatant of Example 1 of the present invention under a 365nm wavelength ultraviolet lamp;

[0026] Figure 2 Shown is a transmission electron microscope image of cadmium sulfide quantum dots synthesized in Example 1 of the present invention;

[0027] Figure 3 Shown is the X-ray powder diffraction spectrum of cadmium sulfide quantum dots synthesized in Example 1 of the present invention;

[0028] Figure 4 The figure shows the UV-visible absorption spectrum of the cadmium sulfide quantum dots synthesized in Example 1 of the present invention after being desalted by PD-10 and replaced with water solvent;

[0029] Figure 5 The figure shows the emission spectrum of the cadmium sulfide quantum dots synthesized in Example 1 of the present invention after being desalted by PD-10 and replaced with water solvent;

[0030] Figure 6 Shown is a fluorescence photograph of cadmium sulfide quantum dots synthesized in the supernatant of Example 2 of the present invention under a 365nm wavelength ultraviolet lamp;

[0031] Figure 7 Shown is a fluorescence photograph of cadmium sulfide quantum dots synthesized in the supernatant of Example 3 of the present invention under a 365nm wavelength ultraviolet lamp;

[0032] Figure 8 The figure shows the UV-visible absorption spectrum of the cadmium sulfide quantum dots synthesized in Example 3 of the present invention after being desalted by PD-10 and replaced with water solvent;

[0033] Figure 9 The figure shows the emission spectrum of the cadmium sulfide quantum dots synthesized in Example 3 of the present invention after being desalted by PD-10 and replaced with water solvent;

[0034] Figure 10 Shown is the X-ray powder diffraction spectrum of cadmium sulfide quantum dots synthesized in Example 3 of the present invention. DETAILED DESCRIPTION

[0035] It should be noted that the following detailed description is illustrative and is intended to provide further explanation of the present invention. Unless otherwise specified, all technical and scientific terms used in the present invention have the same meaning as commonly understood by those skilled in the art to which the present invention belongs.

[0036] The present invention will be further described below with reference to the embodiments.

[0037] Example 1: S powder culture of thermophilic acidthiobacillus supernatant + glutathione + cadmium salt

[0038] (1) Thermoacidithiobacillus (MTH-04) was inoculated into fresh liquid Starkey medium (pH adjusted to ~2.5) and inorganic sulfur powder was added at 8 g / L as a growth energy source. The culture was shaken at 40°C and 150 rpm. After 7 days of culture, the thermoacidithiobacillus entered the stable phase.

[0039] (2) The culture medium was sterilized by centrifugation at 10,000 rpm, the precipitated bacteria were discarded, and the supernatant was collected as the experimental object; the pH of the supernatant was then adjusted to neutral using strong base KOH.

[0040] (3) 0.5 mM cadmium acetate and 5 mM glutathione were added to the above supernatant, and the culture was continuously shaken on a shaker for 3 days at a temperature of 40°C and a shaking speed of 150 rpm. A large amount of CdS quantum dots were synthesized in the supernatant.

[0041] (4) Since the neutralized Acidithiobacillus culture supernatant itself emits blue-green fluorescence, which interferes with the fluorescence signal of the prepared CdS quantum dots, it is necessary to use a PD-10 column to remove the Starkey culture solution. The specific steps are as follows: the above-mentioned CdS quantum dot-containing reaction solution is centrifuged at 10,000 rpm to remove impurities, and the CdS quantum dots are evenly distributed in the supernatant. The supernatant is collected and the Starkey culture solution is removed using a PD-10 column to obtain CdS quantum dots dissolved in an aqueous solution.

[0042] The fluorescence effect of the prepared cadmium sulfide quantum dots was measured under ultraviolet light (wavelength 365nm), and the fluorescence effect was compared. Figure 1 shown. Figure 1Left (UV light off) is a photo of the supernatant containing CdS quantum dots under natural light, in which no color is visible; Figure 1 The right side (UV lamp on) is a fluorescence photograph of the supernatant containing cadmium sulfide quantum dots under irradiation with a 365nm wavelength UV lamp, and the supernatant appears bright yellow.

[0043] The morphology of the prepared CdS quantum dots was observed by transmission electron microscopy. Figure 2 As shown, cadmium sulfide quantum dots were observed to be spherical and the particle size was about 5 nm.

[0044] The prepared cadmium sulfide quantum dots were tested by X-ray diffractometer, as shown in the attached Figure 3 As shown, characteristic diffraction peaks of cubic crystal appear, and it is concluded that the prepared cadmium sulfide quantum dot sample is cubic crystal.

[0045] The UV absorption and fluorescence performance of the prepared cadmium sulfide quantum dot solution were tested and analyzed by UV spectrophotometer and fluorescence spectrometer. It was found that the prepared cadmium sulfide quantum dots had a wide and strong absorption band in the range of 300nm-400nm. Figure 4 As shown. Under the condition of excitation wavelength λ = 350nm, the cadmium sulfide quantum dot aqueous solution has an obvious emission peak at 500nm of visible light, as shown Figure 5 shown.

[0046] Example 2: S powder culture of thermophilic acidthiobacillus supernatant + cysteine + cadmium salt

[0047] (1) Thermoacidithiobacillus (MTH-04) was inoculated into fresh liquid Starkey medium (pH adjusted to ~2.5) and inorganic sulfur powder was added at 8 g / L as a growth energy source. The culture was shaken at 40°C and 150 rpm. After 7 days of culture, the thermoacidithiobacillus entered the stable phase.

[0048] (2) The culture medium was sterilized by centrifugation at 10,000 rpm, the precipitated bacteria were discarded, and the supernatant was collected as the experimental object; the pH of the supernatant was then adjusted to neutral using strong base KOH.

[0049] (3) 0.5 mM cadmium acetate and 5 mM cysteine were added to the above supernatant, and the culture was continuously shaken on a shaker for 3 days at a temperature of 40°C and a shaking speed of 150 rpm. A large amount of CdS quantum dots were synthesized in the supernatant.

[0050] (4) The reaction solution containing cadmium sulfide quantum dots was centrifuged at 10,000 rpm. Most of the cadmium sulfide quantum dots were deposited at the bottom of the test tube. The fluorescence effect of the prepared cadmium sulfide quantum dots was measured under ultraviolet light (wavelength 365 nm). Figure 6 shown. Figure 6 Left (UV light off) is a photo of CdS quantum dots deposited at the bottom of the tube by centrifugation under natural light, with no color visible; Figure 6 The right side (UV lamp on) is a fluorescence photograph of cadmium sulfide quantum dots deposited at the bottom of the tube by centrifugation under 365nm wavelength UV lamp, showing pink fluorescence.

[0051] Example 3 K2S4O6 culture of thermophilic acidthiobacillus supernatant + glutathione + cadmium salt

[0052] (1) Thermoacidithiobacillus (MTH-04) was inoculated into fresh liquid Starkey medium (pH adjusted to ~2.5) and potassium tetrathionate (K2S4O6) was added as a growth energy source at 3 g / L. The culture was shaken at 40°C and 150 rpm. After 4 days of culture, the thermoacidithiobacillus entered the stable phase.

[0053] (2) The culture medium was sterilized by centrifugation at 10,000 rpm, the precipitated bacteria were discarded, and the supernatant was collected as the experimental object; the pH of the supernatant was then adjusted to neutral using strong base KOH.

[0054] (3) 0.5 mM cadmium acetate and 5 mM glutathione were added to the above supernatant, and the culture was continuously shaken on a shaker for 3 days at a temperature of 40°C and a shaking speed of 150 rpm. A large amount of CdS quantum dots were synthesized in the supernatant.

[0055] (4) The cadmium sulfide quantum dot reaction solution was centrifuged at 10,000 rpm to remove impurities. The CdS quantum dots were evenly distributed in the supernatant. The supernatant was collected and the inorganic salt components in the Starkey culture medium were removed using a PD-10 column to obtain CdS quantum dots dissolved in an aqueous solution.

[0056] The fluorescence effect of the prepared cadmium sulfide quantum dots was measured under ultraviolet light (wavelength 365nm), and the fluorescence effect was compared. Figure 7 shown. Figure 7 Left (UV light off) is a photo of the supernatant containing CdS quantum dots under natural light, in which no color is visible; Figure 7 The right side (UV lamp on) is a fluorescence photograph of the supernatant containing cadmium sulfide quantum dots under irradiation with a 365nm wavelength UV lamp, and the supernatant appears yellow.

[0057] The UV absorption and fluorescence performance of the prepared cadmium sulfide quantum dot solution were tested and analyzed by UV spectrophotometer and fluorescence spectrometer. It was found that the prepared cadmium sulfide quantum dots had a wide and strong absorption band in the range of 300nm-400nm. Figure 8As shown. Under the condition of excitation wavelength λ = 350nm, the cadmium sulfide quantum dot aqueous solution has an obvious emission peak at 550-600nm of visible light, as shown Figure 9 shown.

[0058] The prepared cadmium sulfide quantum dots were tested by X-ray diffractometer, such as Figure 10 As shown, characteristic diffraction peaks of cubic crystal appear, and it is concluded that the prepared cadmium sulfide quantum dot sample is cubic crystal.

[0059] Example 4: Supernatant of Acidithiobacillus ferrooxidans cultured with S powder + cysteine + cadmium salt

[0060] (1) Acidithiobacillus ferrooxidans (ATCC 23270) was inoculated into fresh liquid 9K medium (pH adjusted to ~2.5) and inorganic sulfur powder was added as a growth energy source at 8 g / L. The culture was shaken at 30°C and 180 rpm. After 7 days of culture, Acidithiobacillus ferrooxidans entered the stable phase.

[0061] (2) The culture medium was sterilized by centrifugation at 10,000 rpm, the precipitated bacteria were discarded, and the supernatant was collected as the experimental object; the pH of the supernatant was then adjusted to neutral using strong base KOH.

[0062] (3) 0.5 mM cadmium acetate and 5 mM cysteine were added to the above supernatant, and the temperature was 40°C and the shaking speed was 150 rpm. After continuous shaking and incubation for 3 days, a large amount of CdS quantum dots could be synthesized in the supernatant.

[0063] (4) The reaction solution containing cadmium sulfide quantum dots was centrifuged at 10,000 rpm, and most of the cadmium sulfide quantum dots were deposited at the bottom of the test tube.

[0064] The supernatant of chemoautotrophic bacteria such as Acidithiobacillus, which use low-valent sulfur as energy, cultivated to the stable period can be used as a catalytic system for preparing cadmium sulfide quantum dots, including thermophilic Acidithiobacillus, Acidithiobacillus ferrooxidans and other bacteria; using sulfur powder or K2S4O6 as energy, the supernatant of Acidithiobacillus can be used as a catalytic system for preparing cadmium sulfide quantum dots.

[0065] Based on the above results, it can be seen that the addition of thiol amino acids such as glutathione or cysteine can promote the production of CdS quantum dots, but there are differences in the dispersibility of CdS quantum dots produced by the two amino acids. Glutathione has good water solubility, and the CdS quantum dots prepared with glutathione have better dispersion in the supernatant. After centrifugation at 10,000 rpm, most of the CdS quantum dots remain dispersed in the supernatant. In contrast, cysteine has poor water solubility, resulting in moderate dispersion of CdS quantum dots in the supernatant. After centrifugation at 10,000 rpm, most of the CdS quantum dots aggregate as a precipitate at the bottom of the tube.

[0066] Based on the above results, it can be seen that the present invention provides a simple biological method for synthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus. Acidithiobacillus is an autotrophic bacterium that uses inorganic sulfur powder and low-valent sulfur as energy. It has a unique sulfur metabolism system. Studies have found that various forms of sulfur-containing compounds and phosphorus-containing compounds are present in the culture supernatant due to bacterial metabolism. Therefore, the present invention has found that these substances in the supernatant can effectively catalyze the synthesis of cadmium sulfide quantum dots without the need for bacterial cells.

[0067] It is important to note that the supernatant of Acidithiobacillus cultured to the logarithmic phase is highly acidic, with a pH as low as around 1. When using the supernatant, the pH must be adjusted to neutral before adding cadmium salts. The supernatant of Acidithiobacillus cultured to the stationary phase exhibits a distinct blue-green fluorescence, which is absent below a pH of 2.5. However, after the pH is adjusted to neutral, it exhibits distinct blue-green fluorescence under ultraviolet light (365nm). This spontaneous blue-green fluorescence overlaps with the fluorescence emitted by the smaller cadmium sulfide quantum dots synthesized in the early stages, causing some interference. Using gel-based columns such as PD-10 can effectively separate the cadmium sulfide quantum dots from the supernatant, thereby eliminating interference from the supernatant's spontaneous blue-green fluorescence.

[0068] The foregoing description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Those skilled in the art will readily appreciate that various modifications and variations of the present invention are possible. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present invention are intended to be within the scope of protection of the present invention.

Claims

1. A method for biosynthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus thiophanate, characterized in that: The process comprises the following steps: culturing Acidithiobacillus ( Acidithiobacillus ) The supernatant was used as the catalytic reaction system, and after adjusting its pH to neutral, S source and Cd source were added thereto, and the mixture was shaken and cultured to obtain cadmium sulfide quantum dots; The S source is glutathione or sulfur-containing amino acids, and the Cd source is a soluble cadmium salt; The acidithiobacillus is a species of the genus Thiobacillus that uses inorganic sulfur as an energy source; The acidithiobacillus is selected from the group consisting of acidithiobacillus thermophilus, acidithiobacillus sulfuroxidans and acidithiobacillus ferrooxidans.

2. The method for biosynthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus according to claim 1, characterized in that: The medium used in the culture of Acidithiobacillus was Starkey-S 0 Liquid culture medium or Starkey-K2S4O6 liquid culture medium.

3. The method for biosynthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus according to claim 2, characterized in that: During the cultivation of Acidithiobacillus, the pH value of the culture medium is 1-2.

5.

4. The method for biosynthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus according to claim 1, characterized in that: The sulfur-containing amino acid is cysteine.

5. The method for biosynthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus according to claim 1, characterized in that: The water-soluble cadmium salt is cadmium acetate or cadmium sulfate.

6. The method for biosynthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus according to claim 1, characterized in that: The culture temperature for shaking culture is 30~40℃.

7. The method for biosynthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus according to claim 6, characterized in that: The shaking speed of the shaker culture is 150~180 r / min, and the culture time is 2~3 days.

8. The method for biosynthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus according to claim 1, characterized in that: The pH of the Acidithiobacillus supernatant was adjusted to neutral using KOH.

9. The method for biosynthesizing cadmium sulfide quantum dots using the culture supernatant of Acidithiobacillus according to claim 1, characterized in that: In the supernatant of Acidithiobacillus spp., the concentration of soluble cadmium salts was 0.3-0.8 mM, and the concentration of glutathione or sulfur-containing amino acids was 3-8 mM.

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