Hemp (Cannabis sativa L.) protein: Impact of extraction method and cultivar on structure, function, and nutritional quality.

Hemp (Cannabis sativa L.) is increasingly gaining traction as a novel and sustainable source of plant protein. Accordingly, the aim of this study was to investigate the effectiveness of two protein extraction methods, alkaline extraction coupled with isoelectric precipitation (AE-IEP) and salt extraction coupled with ultrafiltration (SE-UF) in producing hemp protein isolates (pH-HPI and salt-HPI) with high purity and yield. Structural characterization as impacted by extraction method and cultivar was performed and related to functional performance and nutritional quality. Both extraction methods, with carefully selected parameters, resulted in HPI with high purity (86.6-88.1% protein) and protein extraction yields (81.6-87.3%). All HPI samples had poor solubility (∼9-20%) at neutral pH compared to commercial soy protein and pea protein isolates (cSPI, cPPI). A relatively high surface hydrophobicity and low surface charge contributed to such poor solubility of HPI. However, HPI demonstrated similar solubility at acidic pH (50-67%) and comparable gel strength (up to 24 N) to cSPI. Comparing experimental amino acid composition to the theoretical amino acid distribution in hemp protein provided insights to the functional performance of the protein isolates. While pH-HPI demonstrated better functionality than salt-HPI, minimal structural, functional, and nutritional differences were noted among the pH-HPI samples extracted from four different cultivars. Overall, results from this work could be used to guide future attempts to further develop successful protein extraction processes, and to provide valuable insights to propel breeding efforts that target enhanced hemp protein characteristics for food applications.

Impact of cooking on the protein quality of Russet potatoes.

Despite being low in crude protein, on a fresh weight basis, given their overall contribution to the North American diet, potatoes contribute approximately 2%-4% of the population's protein intake. However, the quality of the protein remains ill-defined. To that end, Russet potatoes were secured and subjected to various cooking conditions (raw [control], boiled, baked, microwaved, and fried [3, 6, and 9 min]) to determine the impact of cooking method on protein quality, as determined by amino acid score (AAS) and indices of in vivo true fecal protein digestibility (TFPD%; rodent bioassay) and in vitro protein digestibility (pH-drop, pH-Stat, and simulated gastrointestinal digestion both static and dynamic). The AAS of raw Russet potatoes was 0.67 ± 0.01, with histidine being the limiting AA. Frying led to a significant reduction in the AAS, however, other cooking methods yielded similar results to the raw control. The TFPD% of raw potato was low (40.5% ± 3.9%) and was significantly enhanced to over 80% with all cooking methods. Similar patterns were observed with all in vitro measures, however, all methods yielded higher values for the raw control samples. Final protein digestibility-corrected AAS (PDCAAS; product of AAS and TFPD%) values ranged from 0.27 (raw) to a high of 0.57 (boiled), with cooked values being comparable to other plant-based protein sources, including grains, and some nuts and pulses. In vitro PDCAAS values followed similar trends. This study defined the protein quality of cooked Russet potatoes and provides data for use in defining the quality of total protein consumed in the North American diet.

The in vivo and in vitro protein quality of three hemp protein sources.

In this work, the protein quality of defatted hemp hearts and protein-enriched hemp fractions was determined. Protein quality was assessed using a rodent bioassay to evaluate growth and protein digestibility, while amino acid composition was determined via HPLC. A method for determining in vitro protein digestibility was compared to in vivo methodology and used to generate an in vitro protein quality score. The true protein digestibility of hemp protein 2, a hemp protein concentrate, was significantly lower than that of either defatted hemp hearts or hemp protein 1, a hemp protein concentrate (p < .05). While there was no relationship between the in vivo and in vitro measurements of protein digestibility (R 2 = .293, p = .459), there was a significant correlation between the protein digestibility-corrected amino acid score (PDCAAS) determined in vivo and in vitro PDCAAS (R 2 = .989, p = .005). The protein efficiency ratio of hemp protein 1 was significantly lower than that of either defatted hemp hearts or hemp protein 2 (p < .05). These data highlight the nutritional capacity of hemp protein sources while also demonstrating the relationship between in vivo and in vitro methods for determining protein quality.

Comparison of methodologies used to define the protein quality of human foods and support regulatory claims.

Protein quality (PQ) is the capacity of a protein to meet the amino acid (AA) requirements of an individual. There are several methodologies for determining the PQ of foods. The protein efficiency ratio is an animal growth bioassay. The protein-digestibility-corrected AA score considers the AA requirements of a reference population, and the true nitrogen digestibility coefficient for each ingredient. The digestible indispensable AA score is based on true ileal AA digestibility and better represents bioavailability of AAs. In vitro techniques for assessment of PQ are available but require validation against a greater range of protein sources. Isotopic methods, such as the indicator AA oxidation and dual tracer techniques measure AA relative bioavailability and digestibility, respectively, but require sophisticated equipment, and may not be cost nor time effective for the industry to adopt. The present review discusses advantages and disadvantages of methodologies for determining PQ of food for humans focused on methods that are or could be adopted by regulatory agencies. Understanding the framework and resources available for PQ determination will help in the selection of appropriate methods depending on the application. Novelty Understanding the framework and resources available for PQ determination will help in the selection of appropriate methods depending on the application.

Thermal processing methods differentially affect the protein quality of Chickpea (Cicer arietinum).

Chickpea is a widely produced pulse crop, but requires processing prior to human consumption. Protein bioavailability and amino acid quantity of chickpea flour can be altered by multiple factors including processing method. For this reason, the protein quality of processed chickpea flour was determined using in vivo and in vitro analyses for processed chickpeas. Processing differentially affected the protein digestibility-corrected amino acid score (PDCAAS) of chickpeas with extruded chickpea (83.8) having a higher PDCAAS score than both cooked (75.2) and baked (80.03). Interestingly, the digestible indispensable amino acid score (DIAAS) value of baked chickpea (0.84) was higher compared to both extruded (0.82) and cooked (0.78). The protein efficiency ratio, another measure of protein quality, was significantly higher for extruded chickpea than baked chickpea (p < .01). In vivo and in vitro analysis of protein quality were well correlated (R 2 = .9339). These results demonstrated that under certain circumstances in vitro methods could replace the use of animals to determine protein quality.

Determination of the protein quality of almonds (Prunus dulcis L.) as assessed by in vitro and in vivo methodologies.

Almonds (Prunus dulcis), such as all nuts, are positioned within the protein foods grouping within the current U.S. Dietary Guidelines. The ability to make claims related to the protein content of almonds, within the United States, requires substantiation via the use of the Protein Digestibility-Corrected Amino Acid Score (PDCAAS). The present study was designed to provide current estimates of PDCAAS, using both in vivo and in vitro assays, of key almond varietals from the 2017 California harvest. Additionally, historical protein and amino acid composition data on 73 separate analyses, performed from 2000 to 2014, were analyzed. Amino acid analysis confirmed lysine as the first-limiting amino acid, generating amino acid scores of 0.53, 0.52, 0.49, and 0.56 for Butte, Independence, Monterey, and Nonpareil varietals, respectively. True fecal protein digestibility coefficients ranged from 85.7% to 89.9% yielding PDCAAS values of 44.3-47.8, being highest for Nonpareil. Similar, albeit lower, results were obtained from the in vitro assessment protocol. Analysis of the historical data again positioned lysine as the limiting amino acid and yielded information on the natural variability present within the protein and amino acid profiles of almonds. Comparison of the 2017 AA profile, averaged across almond varietals, to the historical data provided strong evidence of persistence of amino acid composition and indices of protein quality over time.

Effect of Processing on the In Vitro and In Vivo Protein Quality of Beans (Phaseolus vulgaris and Vicia Faba).

In this work, the protein quality of different bean types after undergoing the preparatory methods of baking, cooking and extrusion was assayed. Protein quality was assessed using a rodent bioassay to evaluate growth and protein digestibility while amino acid composition was determined via HPLC. In vivo protein digestibility was compared to an in vitro assessment method. The average protein digestibility corrected amino acid score (PDCAAS) for processed beans was higher than the digestible indispensable amino acid score (DIAAS) (61% vs. 45%). Extrusion/cooking of Phaseolus varieties resulted in higher PDCAAS (66% on average) and DIAAS values (61% on average) than baked (52% and 48%) while baked faba beans had higher PDCAAS (66%) and DIAAS (61%) values. A significant correlation was found between PDCAAS and in vitro PDCAAS (R² = 0.7497). This demonstrates which bean processing method will generate the optimal protein quality, which has benefits for both industrial production and individual domestic preparation.

Effect of processing on the in vitro and in vivo protein quality of red and green lentils (Lens culinaris).

In order to determine the effect of extrusion, baking and cooking on the protein quality of red and green lentils, a rodent bioassay was conducted and compared to an in vitro method of protein quality determination. On average, the Protein Digestibility-Corrected Amino Acid Score of red lentils (55.0) was higher than that of green lentils (50.8). Extruded lentil flour had higher scores (63.01 red, 57.09 green) than either cooked (57.40 red, 52.92 green) or baked (53.84 red, 47.14 green) flours. The average Digestible Indispensable Amino Acid Score of red lentils (0.54) was higher than green lentils (0.49). The Protein Efficiency Ratio of the extruded lentil flours (1.30 red, 1.34 green) was higher than that of the baked flour (0.98 red, 1.09 green). A correlation was found between in vivo and in vitro methods of determining protein digestibility (R2=0.8934). This work could influence selection of processing method during product development.

Effect of Processing on the in Vitro and in Vivo Protein Quality of Yellow and Green Split Peas (Pisum sativum).

In order to determine the effect of extrusion, baking, and cooking on the protein quality of yellow and green split peas, a rodent bioassay was conducted and compared to an in vitro method of protein quality determination. The Protein Digestibility-Corrected Amino Acid Score (PDCAAS) of green split peas (71.4%) was higher than that of yellow split peas (67.8%), on average. Similarly, the average Digestible Indispensable Amino Acid Score (DIAAS) of green split peas (69%) was higher than that of yellow split peas (67%). Cooked green pea flour had lower PDCAAS and DIAAS values (69.19% and 67%) than either extruded (73.61%, 70%) or baked (75.22%, 70%). Conversely, cooked yellow split peas had the highest PDCCAS value (69.19%), while extruded yellow split peas had the highest DIAAS value (67%). Interestingly, a strong correlation was found between in vivo and in vitro analysis of protein quality (R2 = 0.9745). This work highlights the differences between processing methods on pea protein quality and suggests that in vitro measurements of protein digestibility could be used as a surrogate for in vivo analysis.

Impact of Processing on the Protein Quality of Pinto Bean (Phaseolus vulgaris) and Buckwheat (Fagopyrum esculentum Moench) Flours and Blends, As Determined by in Vitro and in Vivo Methodologies.

Blending of protein sources can increase protein quality by compensating for limiting amino acids present in individual sources, whereas processing grain flours by extrusion or baking can also alter protein quality. To determine the effect of baking and extrusion on the protein quality of blended flours from buckwheat and pinto beans, a rodent bioassay was performed and compared to an in vitro method of protein quality determination. Overall, extruded products had higher protein efficiency ratio values, increased digestibility, and greater protein digestibility corrected amino acid score (PDCAAS) values than baked products, with the extruded buckwheat/pinto blend having the greatest PDCAAS value of the experimental diets investigated. A correlation was found between both digestibility and PDCAAS values generated from in vitro and in vivo methods. The use of in vitro digestibility analysis should be investigated as a potential replacement for the current rodent assay for nutrient content claim purposes.