Pet food palatability evaluation: A review of standard assay techniques and interpretation of results with a primary focus on limitations

Citation: Aldrich, G. C., & Koppel, K. (2015). Pet food palatability evaluation: A review of standard assay techniques and interpretation of results with a primary focus on limitations. Animals, 5(1), 43-55. doi:10.3390/ani5010043The pet food industry continues to grow steadily as a result of new innovative products. Quality control and product development tests for pet foods are typically conducted through palatability testing with dogs and cats. Palatability is the measure of intake of a food that indicates acceptance or the measure of preference of one food over another. Pet food palatability is most commonly measured using a single-bowl or a two-bowl assay. While these tests answer some questions about the animals’ perception of the food, there are many limitations as well. This review addresses some of these limitations and indicates opportunities for future research. © 2015 by the authors; licensee MDPI, Basel, Switzerlan

The impact of rendered protein meal oxidation level on shelf-life, sensory characteristics, and acceptability in extruded pet food

Citation: Chanadang, S., Koppel, K., & Aldrich, G. (2016). The impact of rendered protein meal oxidation level on shelf-life, sensory characteristics, and acceptability in extruded pet food. Animals, 6(8). doi:10.3390/ani6080044Pet foods are expected to have a shelf-life for 12 months or more. Sensory analysis can be used to determine changes in products and to estimate products’ shelf-life. The objectives of this study were to (1) investigate how increasing levels of oxidation in rendered protein meals used to produce extruded pet food affected the sensory properties and (2) determine the effect of shelf-life on pet owners’ acceptability of extruded pet food diet formulated without the use of preservative. Pet food diets contained beef meat bone meal (BMBM) and chicken byproduct meal (CBPM) in which the oxidation was retarded with ethoxyquin, mixed tocopherols, or none at all, and then extruded into dry pet foods. These samples represented low, medium, and high oxidation levels, respectively. Samples were stored for 0, 3, 6, 9, and 12 months at ambient temperature. Each time point, samples were evaluated by six highly trained descriptive panelists for sensory attributes related to oxidation. Samples without preservatives were chosen for the acceptability test, since the differences in sensory characteristics over storage time were more distinguishable in those samples. Pet owners evaluated samples for aroma, appearance and overall liking. Descriptive sensory analysis detected significant changes in oxidized-related sensory characteristics over storage time. However, the differences for CBPM samples were more pronounced and directional. The consumer study showed no differences in pet owners’ acceptability for BMBM samples. However, the noticeable increase in aroma characteristics (rancid aroma 0.33-4.21) in CBPM samples over storage time did have a negative effect on consumer’s liking (overall liking 5.52-4.95). © 2016 by the authors; licensee MDPI, Basel, Switzerland

Effect of a dry acidulant coating on the palatability of dry extruded dog food

Citation: Jeffrey, A. M., Aldrich, G. C., Huss, A. R., Knueven, C. J., & Jones, C. K. (2016). Effect of a dry acidulant coating on the palatability of dry extruded dog food. Journal of Animal Science, 94, 114-114. doi:10.2527/msasas2016-242In the pet food industry, Salmonella is getting greater scrutiny because it is considered a “reasonably foreseeable hazard” with the implementation of the Food Safety Modernization Act. Specifically, there is zero tolerance for any serotype of Salmonella in pet foods. Salmonella contamination was responsible for 78% of the Class I recalls in pet food according to the most recent Reportable Food Registry Report (FDA, 2015). One potential method of Salmonella mitigation shown to be effective was through coating the exterior of the kibble with a powdered dry acidulant, such as sodium bisulfate (SBS; Jones-Hamilton, Co.). Sodium bisulfate coating on both dog and cat kibbles was shown to provide complete mitigation of Salmonella within 14-d storage (Jeffrey et al., 2014). However, it is thought that the use of dry acidulant with a palatant for coating kibble may negatively impact palatability of a dry dog food. Therefore, the objective of this experiment was to determine if the use of a dry acidulant, SBS, would influence the palatability of a dry dog food. A single dry extruded all life stages dog food was collected from a commercial pet food manufacturer before the coating step. The kibble was coated with either 2.2% spray dried chicken liver + 0.2% SBS or 2.2% spray dried chicken liver + 0.2% powdered silica (control). A total of 20 beagles were used in a standard 2-bowl forced choice palatability test method for 2 d. Dogs were fed 400 g of both diets once per day, with bowls rotated daily to address side bias. Results were analyzed using the GLIMMIX procedure of SAS (Cary, NC). The inclusion of SBS did not affect daily preference of diet (P = 0.23). Furthermore, there was no effect of day (P = 0.18) or the interaction of treatment × day (P = 0.98). These results demonstrate that palatability is not affected by the inclusion of SBS with a palatant in the coating of dog food kibble. Considering that the inclusion of SBS has been shown to be effective at mitigating Salmonella in pet food and no negative effects on palatability were observed, the use of a dry acidulant in a dog food coating gives the industry a promising method to control Salmonella contamination of finished dog foods

Vinylogous Dehydration by a Polyketide Dehydratase Domain in Curacin Biosynthesis

Polyketide synthase (PKS) enzymes continue to hold great promise as synthetic biology platforms for the production of novel therapeutic agents, biofuels, and commodity chemicals. Dehydratase (DH) catalytic domains play an important role during poly­ketide biosynthesis through the dehydration of the nascent poly­ketide intermediate to provide olefins. Our understanding of the detailed mechanistic and structural underpinning of DH domains that control substrate specificity and selectivity remains limited, thus hindering our efforts to rationally re-engineer PKSs. The curacin pathway houses a rare plurality of possible double bond permutations containing conjugated olefins as well as both <i>cis</i>- and <i>trans</i>-olefins, providing an unrivaled model system for poly­ketide dehydration. All four DH domains implicated in curacin biosynthesis were characterized <i>in vitro</i> using synthetic substrates, and activity was measured by LC-MS/MS analysis. These studies resulted in complete kinetic characterization of the <i>all-trans</i>-trienoate-forming CurK-DH, whose <i>k</i>_cat of 72 s^–1 is more than 3 orders of magnitude greater than that of any previously reported PKS DH domain. A novel stereospecific mechanism for diene formation involving a vinylogous enolate intermediate is proposed for the CurJ and CurH DHs on the basis of incubation studies with truncated substrates. A synthetic substrate was co-crystallized with a catalytically inactive Phe substitution in the His-Asp catalytic dyad of CurJ-DH to elucidate substrate–enzyme interactions. The resulting complex suggested the structural basis for dienoate formation and provided the first glimpse into the enzyme–substrate interactions essential for the formation of olefins in poly­ketide natural products. This examination of both canonical and non-canonical dehydration mechanisms reveals hidden catalytic activity inherent in some DH domains that may be leveraged for future applications in synthetic biology

Functional Characterization of a Dehydratase Domain from the Pikromycin Polyketide Synthase

Metabolic engineering of polyketide synthase (PKS) pathways represents a promising approach to natural products discovery. The dehydratase (DH) domains of PKSs, which generate an α,β-unsaturated bond through a dehydration reaction, have been poorly studied compared with other domains, likely because of the simple nature of the chemical reaction they catalyze and the lack of a convenient assay to measure substrate turnover. Herein we report the first steady-state kinetic analysis of a PKS DH domain employing LC–MS/MS analysis for product quantitation. PikDH2 was selected as a model DH domain. Its substrate specificity and mechanism were interrogated with a systematic series of synthetic triketide substrates containing a nonhydrolyzable thioether linkage as well as by site-directed mutagenesis, evaluation of the pH dependence of the catalytic efficiency (<i>V</i>_max/<i>K</i>_M), and kinetic characterization of a mechanism-based inhibitor. These studies revealed that PikDH2 converts d-alcohol substrates to <i>trans</i>-olefin products. The reaction is reversible with equilibrium constants ranging from 1.2 to 2. Moreover, the enzyme activity is robust, and PikDH2 was used on a preparative scale for the chemoenzymatic synthesis of unsaturated triketide products. PikDH2 was shown to possess remarkably strict substrate specificity and is unable to turn over substrates that are epimeric at the β-, γ-, or δ-position. We also demonstrated that PikDH2 has a key ionizable group with a p<i>K</i>_a of 7.0 and can be irreversibly inactivated through covalent modification by a mechanism-based inhibitor, which provides a foundation for future structural studies to elucidate substrate–protein interactions