276
anti- parallel beta sheet, and turns while measurement in the near-UV region
(250–330 nm) provides the detailed fingerprint of tertiary structure of aromatic
amino acid residues (Tryptophan, Tyrosine, and Phenylanaline) and dihedral angles
of disulfide bonds can be used for quality control (Siligardi and Hussain 2017).
Protein molecules exhibit absorption in the ultraviolet region of the spectrum by
peptide bonds (amid chromophores), side chains in proteins, and any prosthetic
groups (Johnson 1992). For example, CD spectra of α-helical proteins have negative
band near 222 nm due to the strong hydrogen-bonding environment of this conformation. A second transition at 190 nm is split into a negative band near 208 nm and
a positive band near 192 nm. Both bands are reduced in intensity in short helices.
Antiparallel β-pleated sheets show a negative band at 216 nm, a positive band
between 195 and 200 nm, and a negative band near 175 nm. However, the position
and magnitude of these bands is variable, resulting in less accurate predictions for
β-structure than for a-helices by CD (Pelton and McLean 2000). Collagens are
unique class of proteins having triple helical structure with conformation of each
strand resembling that of poly-L-proline in an extanded helical conformation where
all bonds are trans to each other (poly-L-proline II). CD spectra associated with
some of the common protein secondary structures are shown below (Fig. 1):
Mass Spectrometry (MS)
MS is a unique spectroscopy technique based on ionization to produce gas phase
protonated ions by an electrostatic field (mass analyzer) and their subsequent separation in a mass spectrometer according to their mass-to-charge ratio (m/z). The
charged ions generated by ionisation are then counted on the electrode and presented in dependence on their m/z ratio. This technique provides information about
molecular weights and chemical structures of the peptides, proteins, carbohydrates,
oligonucleotides, natural products, and drug metabolites (Biemann 2014).
Improvements in instrumentation, advances in on- line separation techniques and
in data processing have contributed to great expansion of MS in food-related analysis. Post era 1981 saw the rise of so-called “soft ionization” techniques such as fast
atom bombardment technique (FAB) followed by the electrospray ionization (ESI)
and matrix-assisted laser desorption/ ionization (MALDI) techniques that made
analyses by mass spectrometry possible for biomolecules, with high accuracy
(0.01%).The common feature of these ionization techniques is ionization without
fragmentation, accurate mass determination, broad applicability and picomole to
femtomole sensitivity. A brief discussion on FAB-, ESI-and and MALDI-MS is
being given below (Leonil et al. 2000).
M. Manzoor et al.
anti- parallel beta sheet, and turns while measurement in the near-UV region
(250–330 nm) provides the detailed fingerprint of tertiary structure of aromatic
amino acid residues (Tryptophan, Tyrosine, and Phenylanaline) and dihedral angles
of disulfide bonds can be used for quality control (Siligardi and Hussain 2017).
Protein molecules exhibit absorption in the ultraviolet region of the spectrum by
peptide bonds (amid chromophores), side chains in proteins, and any prosthetic
groups (Johnson 1992). For example, CD spectra of α-helical proteins have negative
band near 222 nm due to the strong hydrogen-bonding environment of this conformation. A second transition at 190 nm is split into a negative band near 208 nm and
a positive band near 192 nm. Both bands are reduced in intensity in short helices.
Antiparallel β-pleated sheets show a negative band at 216 nm, a positive band
between 195 and 200 nm, and a negative band near 175 nm. However, the position
and magnitude of these bands is variable, resulting in less accurate predictions for
β-structure than for a-helices by CD (Pelton and McLean 2000). Collagens are
unique class of proteins having triple helical structure with conformation of each
strand resembling that of poly-L-proline in an extanded helical conformation where
all bonds are trans to each other (poly-L-proline II). CD spectra associated with
some of the common protein secondary structures are shown below (Fig. 1):
Mass Spectrometry (MS)
MS is a unique spectroscopy technique based on ionization to produce gas phase
protonated ions by an electrostatic field (mass analyzer) and their subsequent separation in a mass spectrometer according to their mass-to-charge ratio (m/z). The
charged ions generated by ionisation are then counted on the electrode and presented in dependence on their m/z ratio. This technique provides information about
molecular weights and chemical structures of the peptides, proteins, carbohydrates,
oligonucleotides, natural products, and drug metabolites (Biemann 2014).
Improvements in instrumentation, advances in on- line separation techniques and
in data processing have contributed to great expansion of MS in food-related analysis. Post era 1981 saw the rise of so-called “soft ionization” techniques such as fast
atom bombardment technique (FAB) followed by the electrospray ionization (ESI)
and matrix-assisted laser desorption/ ionization (MALDI) techniques that made
analyses by mass spectrometry possible for biomolecules, with high accuracy
(0.01%).The common feature of these ionization techniques is ionization without
fragmentation, accurate mass determination, broad applicability and picomole to
femtomole sensitivity. A brief discussion on FAB-, ESI-and and MALDI-MS is
being given below (Leonil et al. 2000).
M. Manzoor et al.