Biochemical Techniques — MCQs

Biochemical Techniques Indian Medical PG Practice Questions and MCQs

Question 251: Shadow casting is used in -

A. Light microscopy
B. Electron microscopy (Correct Answer)
C. Fluorescence microscopy
D. Phase contrast microscopy

Explanation: ***Electron microscopy*** - **Shadow casting** is a technique used in **electron microscopy** to enhance contrast and reveal the three-dimensional topography of small structures and molecules by depositing a thin film of heavy metal at an angle. - This process creates areas with more metal (which appears darker) and areas shielded from the metal deposition (appearing lighter, like a shadow), thereby outlining the specimen. *Light microscopy* - **Light microscopy** uses visible light to illuminate specimens and a system of lenses to magnify images, and it does not typically employ shadow casting techniques for contrast enhancement. - While various techniques like staining are used for contrast, the principle of creating shadows by metal deposition is not applicable to light interactions with the sample. *Fluorescence microscopy* - **Fluorescence microscopy** utilizes the property of some substances to emit light of a longer wavelength when excited by light of a shorter wavelength (fluorescence), and it relies on fluorochromes for visualization, not shadow casting. - This technique creates contrast based on specific labels or autofluorescence, highlighting particular structures without direct shadowing. *Phase contrast microscopy* - **Phase contrast microscopy** converts phase shifts in light passing through a transparent specimen into changes in amplitude (brightness), which are then visible as differences in image contrast, and it does not involve metal deposition or shadow casting. - This method is particularly useful for observing live, unstained biological samples by detecting optical path differences.

Question 252: What type of radiation is primarily investigated using spectroscopy?

A. Electromagnetic radiation (Correct Answer)
B. Subatomic particles
C. Alpha radiation
D. Beta radiation

Explanation: ***Electromagnetic radiation*** - **Spectroscopy** is the study of the interaction between matter and **electromagnetic radiation**, covering a wide range from radio waves to gamma rays. - Different forms of **spectroscopy** (e.g., UV-Vis, IR, NMR) analyze how atoms and molecules absorb or emit specific wavelengths of electromagnetic radiation. *Subatomic particles* - While subatomic particles like electrons and protons can be studied using techniques like **mass spectrometry** or **particle detectors**, these are not forms of conventional **spectroscopy**. - **Spectroscopy** primarily focuses on the energy transitions within atoms and molecules induced by electromagnetic waves, not the direct observation of subatomic particle interactions. *Alpha radiation* - **Alpha radiation** consists of **helium nuclei** and is a type of **particulate radiation**, not electromagnetic radiation. - Its study typically involves **particle detectors** to measure its energy and trajectory, rather than spectroscopic methods. *Beta radiation* - **Beta radiation** consists of **high-energy electrons or positrons** and is also a form of **particulate radiation**. - Similar to alpha radiation, its detection and analysis rely on techniques designed for charged particles, not the direct interaction with electromagnetic fields as measured in spectroscopy.

Question 253: Which method is used to separate a mixture of lipids?

A. Electrophoresis
B. Chromatography (Correct Answer)
C. Isoelectric focusing
D. PAGE

Explanation: ***Chromatography*** - **Chromatography** (e.g., thin-layer chromatography, gas chromatography, high-performance liquid chromatography) is widely used to separate lipids based on differences in their **polarity**, **molecular weight**, or **solubility** in various solvents. - This method allows for the isolation and identification of different lipid classes and individual lipid species from a complex mixture. *Electrophoresis* - **Electrophoresis** separates molecules based on their **charge** and **size** in an electric field, making it more commonly used for proteins and nucleic acids. - Lipids are generally **uncharged** or have very low charge, which makes them poorly suited for separation by standard electrophoretic methods without modification. *Isoelectric focusing* - **Isoelectric focusing** is a type of electrophoresis that separates molecules based on their **isoelectric point (pI)**, which is the pH at which a molecule has no net charge. - This technique is primarily used for **proteins** and **peptides**, as lipids typically lack ionizable groups necessary for establishing a distinct pI. *PAGE* - **PAGE** (Polyacrylamide Gel Electrophoresis) is a common method used to separate **proteins** and **nucleic acids** based on their size and charge. - Lipids are **hydrophobic** and do not readily migrate through an aqueous polyacrylamide gel matrix, making PAGE unsuitable for their direct separation.

Question 254: Critical temperature for liquid nitrogen is ?

A. 36.5°C
B. -20°C
C. -147°C (Correct Answer)
D. -242°C

Explanation: ***-147°C*** - The **critical temperature** is the temperature above which a gas cannot be liquefied, no matter how much pressure is applied. For **liquid nitrogen**, this value is approximately **-147°C**. - At temperatures above **-147°C**, nitrogen exists only in its gaseous phase. *36.5°C* - This temperature is close to **human body temperature** and is not relevant to the critical temperature of nitrogen. - Nitrogen would be in a gaseous state at this temperature and below its critical pressure. *-20°C* - While a low temperature, **-20°C** is still well above nitrogen's **critical temperature**. - At **-20°C**, nitrogen would be a gas unless subjected to very high pressures. *-242°C* - This temperature is below the **critical temperature** of nitrogen, but it is also below its **boiling point** of **-196°C**. - At **-242°C**, nitrogen would be a liquid, but this value is not its critical temperature.

Question 255: What is the process of Hofmann elimination in organic chemistry?

A. E1 elimination reaction favoring tertiary substrates
B. E2 elimination reaction producing the least substituted alkene (Correct Answer)
C. SN1 substitution reaction with carbocation intermediate
D. SN2 substitution reaction with inversion of configuration

Explanation: ***E2 elimination reaction producing the least substituted alkene*** - **Hofmann elimination** is a type of **E2 elimination** reaction where a **quaternary ammonium salt** is heated in the presence of a strong base. - Unlike most E2 reactions which follow **Zaitsev's rule** (producing the most substituted alkene), Hofmann elimination follows the **Hofmann rule**, leading to the formation of the **least substituted (least stable) alkene**. *SN1 substitution reaction with carbocation intermediate* - **SN1 reactions** involve the formation of a **carbocation intermediate** and are typically substitution reactions, not elimination. - Hofmann elimination is an elimination reaction and does not proceed through a carbocation intermediate. *E1 elimination reaction favoring tertiary substrates* - **E1 reactions** are a two-step process involving a **carbocation intermediate** and generally favor **tertiary substrates** and produce the **Zaitsev product**. - Hofmann elimination is a concerted, one-step E2 mechanism and does not involve carbocations. *SN2 substitution reaction with inversion of configuration* - **SN2 reactions** are **bimolecular nucleophilic substitution** reactions that occur in a single step with **inversion of configuration** at the carbon center. - Hofmann elimination is an elimination process resulting in an alkene, not a substitution product, and does not involve inversion of configuration at a stereocenter.

Question 256: Ninhydrin test is used for?

A. Bile salts
B. Amino acids (Correct Answer)
C. Nucleic acid
D. Lipids

Explanation: ***Amino acids*** - The **ninhydrin test** is a chemical test used to detect the presence of **amino acids** and primary and secondary amines. - It produces a **purple-blue color** when it reacts with most amino acids, due to the formation of a colored complex called Ruhemann's purple. *Bile salts* - The detection of **bile salts** typically involves tests like Hay's test or Pettenkofer's test, which are distinct from the ninhydrin reaction. - These tests rely on the physical or chemical properties of bile salts, such as changes in surface tension or specific color reactions with sulfuric acid. *Nucleic acid* - **Nucleic acids** (DNA and RNA) are detected using specific tests like the **diphenylamine test** (for DNA) or orcinol test (for RNA). - These tests target the deoxyribose or ribose sugars present in their structures and result in different color changes compared to ninhydrin. *Lipids* - **Lipids** are typically identified using tests that exploit their nonpolar nature, such as the **emulsion test** or solubility tests in organic solvents. - Their detection does not involve ninhydrin, as they lack the primary or secondary amine groups that react with this reagent.

Question 257: Which technique is used for the separation of proteins based on their mass?

A. Electrophoresis
B. Salting out
C. SDS-PAGE (Correct Answer)
D. Ion exchange chromatography

Explanation: ***Correct Option: SDS-PAGE*** - **SDS-PAGE (Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis)** separates **denatured proteins** almost exclusively by their **molecular mass**. - **SDS** binds to proteins, imparting a uniform negative charge-to-mass ratio, ensuring that separation is primarily based on their size as they migrate through a **polyacrylamide gel**. - This is the gold standard technique for analyzing proteins by molecular weight. *Incorrect Option: Electrophoresis* - This is a general technique that uses an **electric field** to separate molecules based on their **charge** and **size**. - While it can separate proteins, it doesn't exclusively rely on **mass** without additional modifications (like SDS). - Native electrophoresis separates by charge-to-mass ratio, not mass alone. *Incorrect Option: Salting out* - This technique separates proteins based on their **solubility** in high salt concentrations. - Proteins "salt out" or precipitate at different salt concentrations, which is not directly related to their **mass**. - Based on protein surface properties and hydrophobicity. *Incorrect Option: Ion exchange chromatography* - This method separates proteins based on their **net charge** at a particular pH. - Proteins bind to a charged resin and are eluted by changing the **ionic strength** or **pH** of the buffer. - Two types: cation exchange (negative resin) and anion exchange (positive resin).

Question 258: Which of the following reagents would be most useful in determining the N-terminal amino acid of a polypeptide?

A. Trypsin
B. Carboxypeptidase
C. Phenylisothiocyanate (Correct Answer)
D. Cyanogen bromide

Explanation: ***Phenylisothiocyanate*** - **Phenylisothiocyanate** (PITC), also known as Edman's reagent, is used in the **Edman degradation** method to identify the N-terminal amino acid. - It sequentially cleaves the **N-terminal amino acid** without hydrolyzing the rest of the peptide chain, allowing for identification by chromatography. *Trypsin* - **Trypsin** is a protease that cleaves peptide bonds at the carboxyl side of **lysine** and **arginine** residues. - It is used for peptide fragmentation, not for determining the N-terminal amino acid. *Carboxypeptidase* - **Carboxypeptidases** are exopeptidases that cleave amino acids from the **C-terminal end** of a polypeptide chain. - They are used to identify the C-terminal amino acid, not the N-terminal. *Cyanogen bromide* - **Cyanogen bromide (CNBr)** is a chemical reagent that specifically cleaves peptide bonds on the C-terminal side of **methionine** residues. - It is used for specific peptide fragmentation and not for N-terminal sequencing.

Question 259: Which of the following methods is least suitable for determining the structure of proteins?

A. SDS Page
B. Crystallography
C. NMR spectrometry
D. High pressure liquid chromatography (HPLC) (Correct Answer)

Explanation: ***High pressure liquid chromatography (HPLC)*** ✓ - **HPLC** is a technique primarily used for **separation, purification, identification, and quantification** of components in a mixture based on their differing affinities for a stationary phase and a mobile phase. - While HPLC can provide valuable information about a protein's **purity, quantity, and molecular characteristics**, it does **not directly determine the three-dimensional structure** of proteins. - HPLC is fundamentally a **chromatographic separation technique**, not a structural determination method, making it the **least suitable** option for elucidating protein structure. *SDS-PAGE* - **SDS-PAGE** (Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis) separates proteins based on **molecular weight** under denaturing conditions. - It provides information about protein **size and purity** but does not reveal the protein's **native three-dimensional structure**. - While useful for characterization, it is not a structure determination technique. *NMR spectrometry* - **NMR spectroscopy** (Nuclear Magnetic Resonance) is a powerful technique that can determine the **detailed three-dimensional structure of proteins** in solution. - Particularly effective for smaller proteins (typically <30-40 kDa), it detects magnetic properties of atomic nuclei, providing **atomic-level structural and dynamic information**. - This is a **standard method for protein structure determination**. *X-ray Crystallography* - **X-ray crystallography** is the **gold standard** for determining the **atomic and molecular structure** of proteins at high resolution. - By analyzing diffraction patterns from protein crystals, researchers can reconstruct **detailed 3D structures** with atomic precision. - This technique has solved the majority of known protein structures and is **highly suitable for structure determination**.

Question 260: If a polymerase chain reaction (PCR) is performed for three cycles on a single DNA molecule, how many copies of the DNA will be produced?

A. 2 copies
B. 3 copies
C. 4 copies
D. 8 copies (Correct Answer)

Explanation: ***8 copies*** - Each cycle of **PCR** theoretically doubles the number of DNA molecules present. - Starting with one DNA molecule, after 3 cycles, the number of copies will be 2^3 = **8 copies**. *2 copies* - This would be the result after only **one cycle** of PCR, not three cycles. - PCR involves an exponential amplification process. *3 copies* - This number does not follow the **exponential amplification** pattern characteristic of PCR. - PCR is not a simple additive process. *4 copies* - This would be the result after **two cycles** of PCR (2^2 = 4), not three cycles. - Each subsequent cycle doubles the product from the previous cycle.

Practice by Chapter

Spectrophotometry and Colorimetry

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Chromatography Techniques

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Electrophoresis and Applications

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Centrifugation and Ultracentrifugation

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Radioisotope Techniques

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Enzyme-Linked Immunosorbent Assay (ELISA)

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Polymerase Chain Reaction (PCR)

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Blotting Techniques: Southern, Northern, Western

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Mass Spectrometry in Biochemistry

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Recombinant DNA Technology

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DNA Sequencing

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Proteomics and Metabolomics

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