Determination of nucleic acids at nanogram level using resonance light scattering technique with Congo Red

Based on the enhancement of the resonance light scattering (RLS) of Congo Red (CR) by nucleic acid, a new quantitative method for nucleic acid is developed. In the Tris-HCl buffer (pH 10.5), the weak light scattering of CR is greatly enhanced by addition of nucleic acid and CTMAB, the maximum peak is at 560 nm and the enhanced intensity of RLS is in proportion to the concentration of nucleic acid. The linear range is 1.0 x 10(-9) to 1.0 x 10(-6) g ml(-1), 7.5 x 10(-8) to 1.0 x 10(-6) g ml(-1) and 7.5 x 10(-8) to 2.5 x 10(-6) g ml(-1) for herring sperm DNA, calf thymus DNA and yeast RNA, and the detection limits are 0.019, 0.89 and 1.2 ng ml(-1) (S/N = 3), respectively. Actual biological samples were satisfactorily determined.     

Nucleic acids analysis with nano-Ag-Tb(III) by a resonance light scattering technique.

The nano-Ag-terbium(III)-mucleic acids system was observed by a resonance light scattering (RLS) technique for the first time, and the quantitative analysis of nucleic acids at nanogram levels was established. Studies showed that the RLS intensity of the nano-Ag-terbium(III) system can be obviously enhanced by nucleic acid, which was characterized by the RLS spectrum and the UV-Vis spectrum. In this system, the nanoparticles were only of a definite size and in a limited particle concentration region. Further research indicated that under the optimum conditions, the enhanced intensity of RLS is in proportion to the concentration of nucleic acids in the ranges of 7.0 x 10(-9) g ml(-1) to 8.0 x 10(-6) g ml(-1) for calf thymus DNA (ctDNA), 2.0 x 10(-8) g ml(-1) to 1.0 x 10(-6) g ml(-1) for fish sperm DNA (fsDNA) and 1.0 x 10(-9) g ml(-1) to 1.0 x 10(-7) g ml(-1) for yeast RNA (yRNA). The detection limits were 1.4 ng ml(-1) for ctDNA, 1.2 ng ml(-1) for fsDNA and 0.85 ng ml(-1) for yRNA, respectively. Synthetic and real samples were determined satisfactorily.     

Interaction of morin-cetyltrimethylammonium bromide with nucleic acids and determination of nucleic acids at nanograms per milliliter levels based on the enhancement of preresonance light scattering

A new preresonance light scattering (PRLS) assay of nucleic acids is presented. At pH 7.30, the weak PRLS of morin-cetyltrimethylammonium bromide (CTMAB) can be greatly enhanced by the addition of nucleic acids, owing to the interaction between the nucleic acid and morin-CTMAB. After the addition of morin and CTMAB to DNA, the zeta potential of DNA decreases and changes from negative to positive, which is due to the formation of an associate, the aggregation of morin on nucleic acids and the electric neutralization between DNA and the cationic surfactant CTMAB. Mechanism studies showed that the enhanced PRLS comes from the aggregation of morin in the presence of nucleic acids and CTMAB. The enhanced intensity of PRLS is in proportion to the concentration of nucleic acids in the range 7.5 x 10(-9)-1.0 x 10(-5) g ml(-1) for calf thymus DNA, 7.5 x 10(-9)-1.0 x 10(-6) g ml(-1) for salmon sperm DNA and 1.0 x 10(-8)-1.0 x 10(-6) g ml(-1) for yeast RNA. The detection limits are 3.4, 6.2 and 4.1 ng ml(-1) for calf thymus DNA, salmon sperm DNA and yeast RNA, respectively. Synthetic samples were analyzed satisfactorily.     

Fluorescence enhancement effect of morin-nucleic acid-L-cysteine-capped nano-ZnS system and the determination of nucleic acid

It is found that L-cysteine-capped nano-ZnS can further enhance the fluorescence intensity of the morin-nucleic acid system. Under optimum conditions, the enhanced intensity of fluorescence is proportional to the concentration of nucleic acid in the range of 7.0 x 10(-8)-1.0 x 10(-5) g mL(-1) for fish sperm DNA (fsDNA) and 9.0 x 10(-8)-5.0 x 10(-6) g mL(-1) for yeast RNA (yRNA). The corresponding detection limits (S/N = 3) are 2.0 x 10(-8) g mL(-1) and 4.0 x 10(-8) g mL(-1), respectively. The interaction mechanisms of morin-nucleic acid-L-cysteine-capped nano-ZnS system are studied by multiple techniques. It is considered that there exists synergistic effects of groove binding and electrostatic interaction between morin, L-cysteine-capped nano-ZnS and nucleic acid, and the complex of morin-L-cysteine-capped nano-ZnS-nucleic acid is formed.     

The sensitive determination of nucleic acids using resonance light scattering quenching method

It is found that in hexamethylene tetramine (HMTA)-HCl buffer of pH 7.00, nucleic acids can quench the resonance light scattering (RLS) of europium (III) (Eu3+)-2-thenoyltrifluoroacetne (TTA)-1,10-phenanthroline (Phen) system. Based on this, a sensitive method for the determination of nucleic acids is proposed. The experiments indicate that under the optimum conditions, the quenched RLS intensity is in proportion to the concentration of nucleic acids in the range of 1.0x10(-10) to 2.0x10(-6) g ml-1 for fish sperm (fsDNA), 1.0x10(-11) to 1.0x10(-6) g ml-1 for yeast RNA (yRNA), 5.0x10(-11) to 5.0x10(-7) g ml-1 for calf thymus DNA (ctDNA). Their detection limits (S/N=3) are 0.03, 0.006 and 0.002 ng ml-1, respectively. Therefore, the proposed method is the most sensitive RLS method for the determination of nucleic acids so far. The interaction between nucleic acids and Eu3+-TTA-Phen is also discussed.     

Interaction of cetylpyridine bromide with nucleic acids and determination of nucleic acids at nanogram levels based on the enhancement of resonance Rayleigh light scattering

Resonance Rayleigh light scattering (RRLS) spectra of cetylpyridine bromide (CPB)-nucleic acid system and their analytical application have been first studied. The effective factors and optimum conditions of the reaction have been investigated. After CPB and nucleic acid are mixed together, a new absorption peak located at 300 nm appeared, which is due to the formation of new ion associate of CPB-nucleic acid. The new associate can result in two apparent RRLS peaks at 310-400 and 460-480 nm. The RRLS peak of the corrected spectra located at 290-350 nm, which indicate that the RRLS is originated from the absorption of CPB-nucleic acid associate. The peak at 460-480 nm disappears in the corrected RRLS spectra, which indicated that this peak is originated from the strong line emission of the Xe lamp. Under the optimum conditions, the enhanced intensity of RRLS is proportional to the concentration of nucleic acid in the range of 5.0 x 10(-9)-5.0 x 10(-5) g ml(-1) for calf thymus DNA (ctDNA), 1.0 x 10(-8)-4.0 x 10(-5) g ml(-1) for fish sperm DNA (fsDNA) and 1.0 x 10(-8)-5.0 x 10(-5) g ml(-1) for yeast RNA (yRNA). The detection limits (S/N = 3) are 4.3, 8.7 and 7.4 ng ml(-1), respectively. Synthetic samples were determined satisfactorily.     

Flow-injection chemiluminescence determination of ofloxacin and levofloxacin in pharmaceutical preparations and biological fluids.

A novel, rapid and sensitive analytical method is described for determination of ofloxacin and levofloxacin by enhanced chemiluminescence (CL) with flow-injection sampling. The method is based on the CL reaction of the Ce(IV)-Na2S2O4-ofloxacin/levofloxacin-H2SO2 system. The enhanced CL mechanism was developed and the optimum conditions for CL emission were investigated. The CL intensity was correlated linearly (r = 0.9988) with the concentration of ofloxacin (or levofloxacin) in the range of 1.0 x 10(-8) - 1.0 x 10(-7) g ml(-1) and 1.0 x 10(-7) - 6.0 x 10(-6) g ml(-1). The detection limit (S/N = 3) is 7 x 10(-9) g ml(-1). The relative standard derivation (RSD, n = 11) is 2.0% for ofloxacin at 4 x 10(-7) g ml(-1) and for levofloxacin at 6 x 10(-7) g ml(-1). This method has been successfully applied for the determination of ofloxacin and levofloxacin in pharmaceutical preparations and biological fluids with satisfactory results.     

Scopoletine as fluorescence probe for determination of protein

Scopoletine (SLT), 7-hydroxy-6-methoxylcoumarin, is known to possess biological activities such as abirritating and anti-tumor, it can quench intrinsic fluorescence of bovine serum albumin (BSA) and the fluorescence intensity of itself is enhanced. So, SLT is used as fluorescence probe for quantitative determination of protein. The experiments indicate that under optimum conditions, the enhanced intensity of fluorescence is in proportion to the concentration of proteins in a wide range, and their detection limits (S/N=3) are 1.4 x 10(-8)g mL(-1) for BSA and 1.1 x 10(-8)g mL(-1) for HSA, respectively. Samples were satisfactorily determined. The interaction mechanism is also discussed.     

Fluorimetric determination of nucleic acids with terbium and lutetium

In this paper, fluorescence-enhancement of Tb-nucleic acids [fish sperm DNA (fsDNA) and yeast RNA (yRNA)] by Lu³⁺ is studied in detail and is applied to determine nucleic acids. The experiments indicated that under the optimum conditions, a linear relationship was obtained between the fluorescence intensity (I_f) and the concentration of nucleic acids. The linear range is 1.2×10⁻⁸-1.0×10⁻⁴ g/ml for DNA and 3.0×10⁻⁸-8.0×10⁻⁴ g/ml for RNA. The detection limits (signal/noise=3) for DNA and RNA were 4.8×10⁻⁹ and 7.0×10⁻⁹ g/ml, respectively. The mechanism of the co-luminescence effect is also discussed.     

Fluorimetric study of the interaction between human serum albumin and quinolones-terbium complex and its application

A highly sensitive spectrofluorimetric method is proposed for determination of human serum albumin (HSA) and some quinolone drugs. Using quinolones-terbium (Tb3+) complex as a fluorescent probe, in the buffer solution of pH 7.8, HSA can remarkably enhance the fluorescence intensity of the quinolones-Tb3+ complex at 545 nm and the enhanced fluorescence intensity of Tb3+ ion is in proportion to the concentration of HSA and quinolone drugs. Optimum conditions for the determination of HSA were also investigated. The linear ranges and limits of detection are 8.0 x 10(-9) to 8.0 x 10(-8) mol L(-1), 4.20 x 10(-9) mol L(-1) (for HSA); 1.0 x 10(-6) to 4.0 x 10(-6) mol L(-1), 1.87 x 10(-8) mol L(-1) (for norfloxacin) and 1.0 x 10(-7) to 1.0 x 10(-6) mol L(-1), 4.82 x 10(-8) mol L(-1) (for enoxacine), respectively. This method is simple, practical and relatively free interference from coexisting substances, as well as much more sensitive than most of the existing assays.     

Resonance double light scattering method for the determination of proteins with morin-CTMAB

A new determination method of proteins with the limit of determination at nanogram levels is proposed by using a common spectrofluorimeter to detect intensity of resonance double line scattering (RDLS). Proteins including bovine serum albumin (BSA), human serum albumin (HSA) can combine with morin and cetyltrimethylammonium briomide (CTMAB) in the pH range 7.0-8.0 and produce enhanced RDLS signal at lambda(ex)/lambda(em) 305.0/610.0 nm. Optimization conditions for the morin-protein-CTMAB interaction were tested. In the studied system, BSA/CTMAB/morin = 1:2:3. The association constant of morin with BSA is 5.2 x 10(4). Under the optimum conditions, the linear range is 7.5 x 10(-8)-1.0 x 10(-5) g/ml for BSA, 2.5 x 10(-8)-5.0 x 10(-6) g/ml for HSA. The detection limits (S/N = 3) are 66.0 ng/ml for BSA and 23.0 ng/ml for HSA, respectively. Four synthetic samples were analyzed satisfactorily.     

姜黄素-La3+-CTMAB-核酸体系的荧光增强效应及其应用

研究了镧离子(La3+)-姜黄素(CU)-十六烷基三甲基溴化铵(CTMAB)-核酸荧光增强体系.建立了测定核酸的新方法.体系的最优条件为:六次甲基四胺(HMTA)-HCl缓冲溶液(pH 5.80)中,1.00× 10-3 mol/L阳离子表面活性剂CTMAB存在下,姜黄素浓度为2.00×10-5 mol/L,La3+的浓度为1.40×10-4 mol/L时,核酸能增强La3+ -CU络合物的荧光强度,而且体系荧光的增强程度与核酸的加入量在一定范围内呈线性关系.fsDNA,ctDNA和yRNA线性范围分别为7.00×10-4～10.00 mg/L,4.00×10-4～10.00 mg/L和7.00×10-4～10.00 mg/L;检出限分别为0.17,0.02和0.14 μg/L.与已报道的核酸的分析方法相比,本方法具有较宽的线性范围和较高的灵敏度.研究表明,核酸对体系荧光的增强源于DNA主链上PO3-4与CU之间的静电结合,以及通过氢键和疏水力进行的沟槽式结合,为探针分子提供了疏水性的微环境,降低了体系的非辐射能量损失,使体系的荧光强度增加.     

Study on the resonance light scattering spectrum of berberine-cetyltrimethylammonium bromide system and the determination of nucleic acids at nanogram levels

The interaction of berberine with nucleic acid in the presence of cetyltrimethylammonium bromide (CTMAB) in aqueous solution has been studied by spectrophotometry and resonance light scattering (RLS) spectroscopy. At pH 7.30, the RLS signals of berberine were greatly enhanced by nucleic acid in the region of 300-600 nm characterized by four peaks at 324.0, 386.5, 416.5 and 465.0 nm. The binding properties were examined by using a Scatchard plot based on the measurement of enhanced RLS data at 416.5 nm. Under optimum conditions, the increase of RLS intensity of this system at 416.5 nm is proportional to the concentration of nucleic acid. The linear range is 7.5 x 10(-9)-7.5 x 10(-5) g ml(-1) for calf thymus DNA, 7.5 x 10(-9)-2.5 x 10(-5) g ml(-1) for herring sperm DNA, and 5.0 x 10(-9)-2.5 x 10(-5) g ml(-1) for yeast RNA. The detection limits (S/N = 3) are 2.1 ng ml(-1) for calf thymus DNA, 6.5 ng ml(-1) for herring sperm DNA and 3.5 ng ml(-1) for yeast RNA, respectively. Three synthetic samples were analyzed satisfactorily.     

Post-chemiluminescence phenomenon of NBS-luminol reactions and their applications to flow injection analysis of piroxicam.

A new post-chemiluminescence (PCL) reaction was observed when piroxicam was injected into the reaction mixture after the CL reaction of N-bromosuccinimide (NBS) and luminol. A possible reaction mechanism was proposed based on the studies of the CL kinetic characteristics, the CL spectra, fluorescence spectra and other experiments. A new flow injection CL method for the determination of piroxicam was established based on the PCL reaction. The relative standard deviation (RSD) for the determination of piroxicam was 1.2% (n = 11, c = 2.0 x 10(-6) g/ml). The CL intensity responded linearly to the concentration of piroxicam in the range 1.0 x 10(-7) - 1.0 x 10(-5) g/ml (r = 0.9991). The detection limit was 4.0 x 10(-8) g/ml. The method had been applied to the determination of piroxicam in tablets with satisfactory results.     

Study on the interaction between nucleic acid and Eu3+-oxolinic acid and the determination of nucleic acid using the resonance light scattering technique

At pH 9.75, the resonance light scattering (RLS) intensity of OA–Eu³⁺ system is greatly enhanced by nucleic acid. Based on this phenomenon, a new quantitative method for nucleic acid in aqueous solution has been developed. Under the optimum condition, the enhanced RLS is proportional to the concentration of nucleic acid in the range of 1.0 × 10⁻⁹ to 1.0 × 10⁻⁶ g/ml for herring sperm DNA, 8.0 × 10⁻¹⁰ to 1.0 × 10⁻⁶ g/ml for calf thymus DNA and 1.0 × 10⁻⁹ to 1.0 × 10⁻⁶ g/ml for yeast RNA, and their detection limits are 0.020, 0.011 and 0.010 ng/ml, respectively. Synthetic samples and actual samples were satisfactorily determined. In addition, the interaction mechanism between nucleic acid and OA–Eu³⁺ is also investigated.     

核酸对氯化银胶体溶液共振光散射的猝灭作用及其应用

报道了一种测定水溶液中核酸的方法,该法基于核酸对氯化银溶胶共振射光的猝灭作用。在理想测定条件下,散射光的猝灭程度正比于核酸的浓度,三种核酸（ｃａｌｆ ｔｈｙｍｕｓ ＤＮＡ,ｈｅｒｒｉｎｇ ＤＮＡ ａｎｄ ＹｅａｓｔＲＮＡ）的线性范围分别为０－２０μｇ／Ｌ,０－６０μｇ／Ｌ和０－８０μｇ／Ｌ,检测限分别为０．６５μｇ／Ｌ,１．１μｇ／Ｌ和１．９μｇ／Ｌ。６种合成样品的测定结果令人满意,机理研究结果表明,核酸中的碱基（尤其是嘌呤碱）同银离子具有很强的结合能力,这种结合影响了氯化银的沉淀平衡,导致了氯化银溶胶共振散射光的猝灭。     

Sensitive assay for catecholamines in pharmaceutical samples and blood plasma using flow injection chemiluminescence analysis.

A rapid and sensitive chemiluminescence (CL) method using flow injection analysis was described for the determination of three catecholamines: dopamine, adrenaline and dobutamine, based on their greatly enhancing effects on the CL reaction of luminol-potassium periodate in basic solutions. Under the optimized conditions, the calibration graphs relating the increase of CL intensity to the concentration of the analytes were linear. The present method allows for the determination of dopamine, adrenaline, and dobutamine over the range of 1.0 x 10(-10) - 1.0 x 10(-7) g/ml. The relative standard deviations for measurements (n=11) of dopamine, adrenaline and dobutamine were 2.9, 2.3 and 1.8% when the concentrations of three catecholamines were at 1.0 x 10(-9) g/ml, respectively. The detection limits of the method were 2.0 x 10(-11) g/ml dopamine, 1.0 x 10(-11) g/ml adrenaline and 4.0 x 10(-11) g/ml dobutamine. The method was successfully applied to the determination of three catecholamines in pharmaceutical samples and blood plasma.     

Nucleic acids determination using the complex of eriochrome black T and silver nanoparticles in a resonance light scattering technique

A novel method for the determination of nucleic acids by using silver nanoparticle (AgNPs)-eriochrome black T (EBT) as a resonance light scattering (RLS) probe has been developed. Under optimum conditions, there are linear relationships between the quenching extent of RLS intensity and the concentration of nucleic acids in the range of 4.0×10(-9)-4.0×10(-7), 4.0×10(-7)-1.6×10(-6) g mL(-1) for fish sperm DNA (fsDNA) and 4.0×10(-8)-2.0×10(-6) g mL(-1) for yeast RNA (yRNA). Their detection limits (S/N=3) are 2.0 ng mL(-1) and 21 ng mL(-1), respectively. The results indicate that AgNPs can form wirelike aggregates and nanoslices in the presence of the EBT. Whereas, when nucleic acids are added into the AgNPs-EBT system, the dynamic balance of AgNPs-EBT system is destroyed and the nanoparticles undergo dispersion again, leading to the RLS intensity of AgNPs-EBT system quenching. Meanwhile, the conformation of fsDNA is changed by the synergistic effect of AgNPs and EBT.     

The second-order scattering study of the Tb(III)-RNA and determination of RNA

The second-order scattering technique (SOS), using a common spectrofluorometer, was first developed as a sensitive instrumental analysis method for determination of the ribonucleic acid (RNA). The results indicate that RNA had a weak SOS peak and the Tb(III) ion can greatly enhance the SOS intensity of RNA with the maximum peak located at 612.0 nm. Mechanism study shows that the peak results from the long-range assembly of Tb(III) ion on the molecular surface of RNA. At the pH 7.50 and with cetyltrimethylammonium bromide (CTMAB) (6.0 x 10(-5)M), the enhanced SOS intensity was in proportion to the concentration of RNA in the range of 2.0 x 10(-8) to 2.0 x 10(-5)g/ml. The detection limit was 1.96 ng/ml. The relative standard deviation (five replicates) was within +/-5% in the linear range. This method has been used satisfactorily for the determination of both synthetic and real samples. In comparison with most other methods for the determination of ribonucleic acids, this method is more sensitive.     

Chemiluminescence of tryptophan and histidine in Ru(bpy)3(2+)-KMnO4 aqueous solution

Amino acids with different chemical structures have different abilities in terms of increasing the intensity of chemiluminescence (CL) of tris(2,2'-bipyridine)ruthenium(II) [Ru(bpy)3(2+)]. In a flow system, CL caused by the reaction between Ru(bpy)3(3+) and 15 amino acids was observed, but only tryptophan (Trp) and histidine (His) enhanced the intensity obviously, and so the CL of Trp and His and their molecular groups was studied. A calculation of the ionization potentials (IPs) of their N atom indicated that the CL intensities of these compounds depended on their IPs. In addition, the flow system was used for the determination of Trp and His, and the detection limits were 3 x 10(-8) mol L(-1) for His and 2.5 x 10(-9) mol L(-1) for Trp. The calibration curves for the two amino acids were 1.0 x 10(-7) to 5.0 x 10(-3) mol L(-1) for His and 1.0 x 10(-8) to 1.0 x 10(-4) mol L(-1) for Trp. The proposed approach was applied to the determination of His in Ganoderma.