Anuja Kulkarni

Over the years, human lifestyle and food habits have changed drastically. This current century has also seen an increased awareness about personal well-being, raising consumer demands for obtaining functional ingredients in concentrated forms to meet nutrient requirements and combat diseases. The hunt for these ‘nutraceuticals’ has triggered exploratory research for obtaining such natural bioactive compounds. While there has been a significant advancement in obtaining bioactives from microbial and animal sources, they face severe apprehensions on ethical grounds. Moreover, the product titres obtained from these routes are considerably lower and fail to suffice as a reliable resource. Therefore, it becomes inevitable to unleash the underutilized potential of agro-food wastes, which is rich in various bioactive compounds, like lipids, proteins, fibres, flavonoids, phytochemicals, etc. However, these by-products and agro-food wastes are ascribed with complexities and recalcitrant natures owing to the diverse segments which generate them together in enormous amounts every day. These adversities, therefore, challenge their sustainable and complete utilisation. Moreover, the uncertainties associated with the qualitative and quantitative aspects of agro-food wastes also limit their applicability. This chapter discusses recent advances and possibilities of developing selective strategies for recovery, encapsulation, and applying nutraceuticals derived from agro-food wastes.

Background Methemoglobin is the reduced form of haemoglobin that is normally found in the blood in levels < 1%. Methemoglobinemia can occur as a congenital or acquired disease. Two types of recessive congenital methaemoglobinemia (RCM) are caused by the NADH-dependent cytochrome b5 reductase enzyme deficiency of the CYB5R3 gene. RCM-I is characterized by higher methaemoglobin levels (> 2 g/dL), causing only cyanosis, whereas RCM-II is associated with cyanosis with neurological impairment. Methods Routine haematological investigations were done by standard method. The methaemoglobin level was evaluated by the potassium ferricyanide assay. NADH-cytochrome b5 reductase (cytb5r) enzyme activities were measured by standard methods, and molecular analysis was performed by polymerase chain reaction (PCR) followed by DNA sequencing. The interpretation of mutation effect and the molecular modeling were performed by using specific software DEEP VIEW SWISS-PDB VIEWER and Pymol molecular graphics program. Results The present study discovered three novel homozygous pathogenic variants of CYB5R3 causing RCM I and II in four unrelated Indian patients. In patient-1 and patient-2 of RCM type I caused due to novel c.175C>T (p.Arg59Cys) and other reported c.469T>C (p.Phe157Ser) missense pathogenic variants respectively, whereas patient-3 and patient-4 presented with the RCM type II are related to developmental delay with cyanosis since birth due to a novel homozygous (g.25679_25679delA) splice-site deletion and novel homozygous c.824_825insC (p.Pro278ThrfsTer367) single nucleotide insertion. The CYB5R3 transcript levels were estimated by qRT-PCR in the splice-site deletion, which was 0.33fold of normal healthy control. The insertion of nucleotide C resulted in a frameshift of termination codon are associated with neurological impairment. Conclusions Molecular diagnosis of RCM can help to conduct genetic counselling for novel mutations and, subsequently, prenatal diagnosis of high-risk genetic disorders.

Scope: Chickpea (Cicer arietinum) allergy has frequently been reported particularly in Spain and India. Nevertheless, chickpea allergens are poorly characterized. The authors aim to identify and characterize potential allergens from chickpea.Methods and Results: Candidate proteins are selected by an in silico approach or immunoglobuline E (IgE)‐testing. Potential allergens are prepared as recombinant or natural proteins and characterized for structural integrity by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE), circular dichroism (CD)‐spectroscopy, and mass spectrometry (MS) analysis. IgE‐sensitization pattern of Spanish chickpea allergic and German peanut and birch pollen sensitized patients are investigated using chickpea extracts and purified proteins. Chickpea allergic patients show individual and heterogeneous IgE‐sensitization profiles with extracts from raw and boiled chickpeas. Chickpea proteins pathogenesis related protein family 10 (PR‐10), a late embryogenesis abundant protein (LEA/DC‐8), and a vicilin‐containing fraction, but not 2S albumin, shows IgE reactivity with sera from chickpea, birch pollen, and peanut sensitized patients. Remarkably, allergenic vicilin, DC‐8, and PR‐10 are detected in the extract of boiled chickpeas.Conclusion: Several IgE‐reactive chickpea allergens are identified. For the first time a yet not classified DC‐8 protein is characterized as minor allergen (Cic a 1). Finally, the data suggest a potential risk for peanut allergic patients by IgE cross‐reactivity with homologous chickpea proteins.