Non-toxic Food Research Articles

Encapsulation of Lactiplantibacillus plantarum probiotics through cross-linked chitosan and casein for improving their viability, antioxidant and detoxification

In the present study, encapsulation of Lactiplantibacillus plantarum (L.p) was performed using chitosan and casein through calcium phosphate intercrossing. Chitosan and casein both considered as non-toxic and biocompatible food derived components with intrinsic antioxidant properties. Layer by layer strategy was performed for deposition of modified cross-linked chitosan along with casein as the novel protective layers on the surface of probiotics. After confirmation of successful encapsulation, the viability and antioxidant activity of encapsulated L.p was evaluated. The results showed enhanced survival and antioxidant activity of encapsulated L.p compared to free bacteria in simulated digestive conditions. The survival of free and encapsulated L.p was respectively 1.38 ± 0.29 log cfu/ml and 6.99 ± 0.12 log cfu/ml in SGF and 8.54 ± 0.05 log cfu/ml and 7.25 ± 0.23 log cfu/ml in SIJ after 2 h of incubation. HPLC analysis was also used to investigate the detoxification activity of probiotics toward Aflatoxin M1 and obtained results showed encapsulated bacteria could significantly reduce aflatoxin M1 (68.44 ± 0.5 %) compared to free bacteria (43.76 ± 0.54 %). The results of this research suggest that the chitosan/casein mediated encapsulation of L.p with layer-by-layer technology is an effective method to improve the survival and antioxidant properties of probiotics with enhanced detoxification of AFM1.

Abstract 5288: Soy isoflavone genistein reboosts targeted therapy sensitivity in triple negative breast cancer

Abstract Triple-negative breast cancer (TNBC) is a very aggressive type of breast cancer that does not expression important proteins called estrogen, progesterone and HER2. This makes TNBC hard to treat because it does not respond to common targeted treatments. However, recent treatments using the immune system to target key immune proteins such as PD-1 and PD-L1 have been successful in many types of cancer. This form of immunotherapy works by blocking PD-1/PD-L1 pathway and leads to stimulating the body's own immune system to recognize and attack cancer cells, thus providing a potential new treatment option for TNBC patients. Nevertheless, only approximately 30% of TNBC patients may respond to these drugs, suggesting the majority of TNBC patients would not benefit from these innovative immunotherapies. It is believed that low levels of immune cells and PD-L1 expression in tumors are the primary reasons for the lack of response or resistance to anti-PD-1/PD-L1 treatments. The use of nontoxic and healthy food and nutrition to increase treatment response is an emerging field of cancer research. Our studies showed that adding genistein (GE), a bioactive dietary component found in soybean products such as soy milk, soy protein and tofu, could significantly enhance the efficacy of anti-PD-1 immunotherapy in a TNBC mouse model that was previously not responsive to the treatment. Our research also found that soybean GE can increase tumor immune response by significantly recruiting active immune cells into the tumor and increasing PD-L1 expression. Our studies will help to establish a novel nutrition-based intervention targeting TNBC patients who might not qualify for this treatment initially. Eventually, we will develop a novel strategy of administering a tailored dietary plan to resensitize these patients to emerging anti-PD-1/PD-L1 immunotherapies. Citation Format: Taiwo Bankole, Zhenhai Li, Yuanyuan Li. Soy isoflavone genistein reboosts targeted therapy sensitivity in triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5288.

Cyanidin-3-O-Glucoside Combined with PGC1α Inhibitor Promotes Cancer Cell Death by Inducing Excessive Levels of ROS in Cervical Squamous Cell Carcinoma

Cervical cancer is a global public health problem, particularly in the low-income and middle-income countries. Natural products, such as cyanidin and its derivative cyanidin-3-O-glucoside (C3G), are considered safe and nontoxic food additives, potent therapeutic options for cancer. The present study aims to evaluate the beneficial effects of C3G on cervical squamous cell carcinoma (CSCC) in vitro and explore more potential therapeutic strategies for CSCC. C3G significantly inhibited the cell growth of CSCC cells and induced cancer cell apoptosis in a dose-dependent manner. Meanwhile, C3G promoted cellular reactive oxygen species (ROS) accumulation and decreased mitochondrial membrane potential and mitochondrial mass. The antioxidant regent N-acetyl-L-cysteine (NAC) rescued the effect of C3G on CSCC cells, further demonstrating ROS’s important role in C3G treatment. To explore the underlying mechanism, autophagy-related signaling pathways were investigated, and the results showed that C3G induced cell autophagy but not mitophagy. More importantly, C3G caused a significant activation of mitochondria biogenesis through the peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α) signaling pathway. C3G synergized with PGC-1α inhibitor SR-18292 induced more severe ROS accumulation and showed more potent inhibition of cell proliferation and mitochondrial membrane potential than C3G treatment alone. Thus, the present study suggests a new potential therapeutic strategy for CSCC based on the synergistic effect of C3G and PGC1α inhibitors.

Less brodifacoum in baits results in greater accumulation in the liver of captive Rattus norvegicus in a no-choice trail

AbstractAnticoagulants are widely used as potent cumulative rodenticides; however, they are also associated with adverse environmental effects, such as intoxication of non-target animals. To ensure user safety, in the EU the use of anticoagulants containing active substances of > 30 ppm is restricted to professionals as these substances are toxic to reproduction Kat.1 A or B. Therefore, new products with < 30 ppm anticoagulant substance have been developed and registered. While the biological efficacy of these new products has been evaluated, the effect of their application on liver residues in targeted rodents was not examined before product introduction to the market. Our laboratory no-choice study on wild brown rats (Rattus norvegicus) showed that baits with high (50 ppm) and low (25 ppm) brodifacoum concentrations (with identical nontoxic cereal-based food components) presented 100% efficacy, although the 25 ppm bait exhibited significantly higher palatability and 4.5 times higher liver residues. Rats consumed 13.6 and 167.7 g of the 50 and 25 ppm baits, respectively, which corresponded to 3.16 and 14.91 μg/g of brodifacoum liver residues at their time of death. The experiments demonstrated that a decreased concentration of brodifacoum anticoagulant in rodenticide baits may lead to greater accumulation in rodent liver, thus indicating the urgent need to perform a detailed study of the environmental risks of low-concentration anticoagulant baits.

Ants evade harmful food by active abandonment

Invasive ants, such as the Argentine ant, pose a severe economic and ecological threat. Despite advancements in baiting techniques, effectively managing established ant populations remains a daunting challenge, often ending in failure. Ant colonies employ behavioural immunity against pathogens, raising the question of whether ants can collectively respond to toxic baits. This study investigates whether ant colonies actively abandon palatable but harmful food sources. We provided two sucrose feeders, each generating a new foraging trail, with one transitioning to offering toxic food. Six hours later, ant activity on that path decreases, while activity on the non-toxic food and the trunk trail remains unaffected, excluding factors like population decline or satiation as reasons for the activity decline. Laboratory experiments confirmed that ants remained alive six hours after ingesting toxic food. Ant presence remains low on the toxic food path for days, gradually decreasing along the nearest section of the trunk trail. This abandonment behaviour minimises the entry of harmful food into the nest, acting as a protective social mechanism. The evasion of toxic bait-treated areas likely contributes considerably to control failures. Understanding the behavioural response to toxic baits is essential for developing effective strategies to combat invasive ant species.

Design of SC PEP with enhanced stability against pepsin digestion and increased activity by machine learning and structural parameters modeling

Prolyl endopeptidases from Sphingomonas capsulata (SC PEP) has attracted much attention as promising oral therapy candidate for celiac sprue, however, its low stability in the gastric environment leads to unsatisfactory clinical results. Therefore, improving its stability against pepsin digestion at low pH is crucial for clinical applications, but challenging. In this study, machine learning and physical parameter model were combined to design SC PEP mutants. After iterations, 20 mutants had higher hydrolysis activity in stomach environment, which was up to 14.1-fold compared with wild-type SC PEP. Mutant M24 involving stable and active mutations and pegylated M24 (M24-PEG) had higher activity of hydrolyzing immunogen in bread than wild-type SC PEP in vitro and in vivo, and residual immunogens in simulated gastric environment were only 1/8 and 1/10 of that in the wild-type SC PEP group. The total residual immunogens in the gastrointestinal tract of mice in the M24 and M24-PEG groups were <20 ppm, reaching the standard of non-toxic food. Our results indicate that the combination of M24 (or M24-PEG) with EP-B2 may be a promising candidate for celiac disease, and the strategies developed in this study provide a paradigm for the design of SC PEP stability mutants.

Red beetroot betalains as a novel source of colorent in ice-cream as compared with Red Dye 40 (E129)

Background: Although there is an increasing need for eco-friendly and non-toxic food colorants, plant-based colors have shown to be a promising alternative to synthetic food colors. The natural pigment (betalain) was extracted from red beetroot utilizing a magnetic stirrer shaking apparatus in the current study. Objective: The purpose of this study was to evaluate the effect of betalain (natural pigment) and red dye 40 (synthetic pigment), on the chemical, microbiological, and sensory properties of ice cream. Materials and Methods: Betalain and red dye 40 were added into ice cream at variable concentrations of 50 mg/mL, 100 mg/mL, and 200 mg/mL. Specimens were stored for 70 days and evaluated every ten days. Results: The values of titratable acidity, PH, fat, protein, total count of microorganisms, and sensory characteristics were gave higher scores samples than than in ice cream with red dye 40. Nevertheless, betalain doses of 50, 100, and 200 mg/mL have been reported to have a significant 70-day storage activity in ice cream. When compared to mixtures containing 50, 100, and 200 mg/mL of red dye 40 after 70 days. According to the findings, betalain may be utilized as a natural pigment and food preservative to boost stability during storage.Keywords: Red beetroot,Betalain, Functional foods, Red Dye 40 (E129),Antioxidants, synthetic colors, Storage stability

The Influence of the Toxic Dinoflagellate Alexandrium minutum, Grown under Different N:P Ratios, on the Marine Copepod Acartia tonsa.

HABs pose a threat to coastal ecosystems, the economic sector and human health, and are expanding globally. However, their influence on copepods, a major connector between primary producers and upper trophic levels, remains essentially unknown. Microalgal toxins can eventually control copepod survival and reproduction by deterring grazing and hence reducing food availability. We present several 24-h experiments in which the globally distributed marine copepod, Acartia tonsa, was exposed to different concentrations of the toxic dinoflagellate, Alexandrium minutum, grown under three N:P ratios (4:1, 16:1 and 80:1), with the simultaneous presence of non-toxic food (the dinoflagellate Prorocentrum micans). The different N:P ratios did not affect the toxicity of A. minutum, probably due to the low toxicity of the tested strain. Production of eggs and pellets as well as ingested carbon appeared to be affected by food toxicity. Toxicity levels in A. minutum also had an effect on hatching success and on the toxin excreted in pellets. Overall, A. minutum toxicity affected the reproduction, toxin excretion and, to an extent, the feeding behavior of A. tonsa. This work indicates that even short-term exposure to toxic A. minutum can impact the vital functions of A. tonsa and might ultimately pose serious threats to copepod recruitment and survival. Still, further investigation is required for identifying and understanding, in particular, the long-term effects of harmful microalgae on marine copepods.

Biopolymer Food Packaging Films Incorporated with Essential Oils.

Petroleum-based packaging materials are typically nonbiodegradable, which leads to significant adverse environmental and health issues. Therefore, developing novel efficient, biodegradable, and nontoxic food packaging film materials has attracted increasing attention from researchers. Due to significant research and advanced technology, synthetic additives in packaging materials are progressively replaced with natural substances such as essential oils (EOs). EOs demonstrate favorable antioxidant and antibacterial properties, which would be an economical and effective alternative to synthetic additives. This review summarized the possible antioxidant and antimicrobial mechanisms of various EOs. We analyzed the properties and performance of food packaging films based on various biopolymers incorporated with EOs. The progress in intelligent packaging materials has been discussed as a prospect of food packaging materials. Finally, the current challenges regarding the practical application of EOs-containing biopolymer films in food packaging and areas of future research have been summarized.

Novel colorimetric, photothermal and up-conversion fluorescence triple-signal sensor for rosmarinic acid detection

Rosmarinic acid (RA) is promising as a natural and nontoxic food additive. However, many analysis methods for RA generally depend on large instruments and single signals for quantitative detection. A new up-conversion fluorescence, colorimetric and photothermal multi-modal sensing strategy is developed for the quantification of RA. β-cyclodextrin (CD) modified citric acid (Cit) wrapped NaYF4:Yb/Er-Cit-CD (Y:Yb/Er-Cit-CD) up-conversion nanocomposite has been synthesized, which emits green fluorescence at 550 nm under 980 nm near-infrared (NIR) excitation. In the presence of oxidized 3,3′,5,5′-tetramethylbenzidine (oxTMB), the green fluorescence is significantly quenched attributed to the fluorescence inner filter effect (IFE) between oxTMB and Y:Yb/Er-Cit-CD. When RA is intervened, blue oxTMB is reduced to colorless 3,3′,5,5′-tetramethylbenzidine (TMB) inducing the recovery of up-conversion fluorescence. At the same time, colorimetric and photothermal signals readout can be easily achieved thanks to the color indication and photothermal effect of the oxTMB. The constructed Y:Yb/Er-Cit-CD/oxTMB sensor displays high sensitivity, visibility and simplicity for RA, and the limits of detection (LOD) for fluorescence, colorimetric and photothermal were 0.004 µmol/L, 0.036 µmol/L and 0.043 µmol/L, respectively. This sensing system is successfully performed for the detection of RA in food samples.

Citric acid cross-linked curcumin/chitosan/chickpea flour film: An active packaging for chicken breast storage

Since there has been an increasing interest in natural, non-toxic, and biodegradable food packages instead of plastic ones, scientists put great effort into eliminating the potential drawbacks of these biodegradable films. In this study, citric acid (CA) was utilized as a crosslinker for the films produced by chitosan (CS), chickpea flour (CF), and active agent curcumin (CUR) by a solvent casting technique. CA was introduced into the film formulations at different concentrations (0, 0.5%, 1%, 1.5% w/v), and the influence on the functional film characteristics were extensively analyzed.Increasing CA concentration in formulation caused a reduction in water vapor permeability (WVP) of the films such that the film with the highest CA concentration (1.5-CUR/CF/CS) was 62.6% more resistant to water vapor than film without CA (0-CUR/CF/CS). When CA concentration in the film formulation was increased, elongation at break (EAB) value increased from 1.64% ± 0.13%, to 11.1%± 1.21% whereas tensile strength (TS) value decreased from 7.83± 0.08 MPa to 3.58± 0.20 MPa. Contrary to expectations, antioxidant activity of the films decreased with increasing CA concentration. X-Ray Diffraction (XRD) patterns of the films proved the amorphous nature and inhibitory characteristic of CA against retrogradation. In addition to all these, CA concentration significantly influenced Thermogravimetric analysis (TGA) pattern of the films with the increasing number of degradation stages. Besides the cross-linker characteristic of CA, it also extensively enhanced the antibacterial characteristic of the films. Films formulated with 1.5% (w/v) CA (1.5-CUR/CF/CS) showed antibacterial effect against both Escherichia coli (E.coli) (ATCC 11229) and Staphylococcus aureus (S.aureus) (ATCC 43300). Finally, chicken breast packed with 1.5-CUR/CF/CS films remained below unacceptable microbial threshold limits even at the end of the 9th -day of storage compared to samples packaged with the film with no antibacterial activity.

Efficacy of Eight Anticoagulant Food Baits in House Mouse (Mus musculus): Comparison of Choice and No-Choice Laboratory Testing Approaches

House mouse (Mus musculus) is a cosmopolitan pest in agricultural facilities, commodity stores and rural and urban environments. It is regularly controlled by anticoagulant-based baits. Since the number of registered active ingredients is limited, the producers are trying to develop new formulations with increased tamper-resistance, attractiveness and palatability. Reliable and economical methods for laboratory screening/testing are needed. Therefore, this work compared simple no-choice with more economically demanding choice feeding tests for laboratory evaluation of efficacy of rodenticide anticoagulant-based preparations in wild populations of house mouse. We analyzed mice survival and bait intake of the various rodenticide commercial preparations differing in a toxic active substance (warfarin, bromadiolone, brodifacoum, difethialone) and nontoxic food lure components. In most cases, we found insignificant differences in survival of house mice treated with eight test baits within the no-choice and choice feeding tests. We rejected the tested hypothesis that there would be significant differences in all tested preparations for two different testing approaches.

Perillaldehyde mitigates virulence factors and biofilm formation of Pseudomonas aeruginosa clinical isolates, by acting on the quorum sensing mechanism in vitro.

The incidence of biofilm linked catheter-associated urinary tract infections is increasing worldwide and Pseudomonas aeruginosa is one of the major causes. Perillaldehyde (PLD): as a natural, widely used flavouring agent, has been reported to possess various pharmacological properties. We hypothesized that PLD can inhibit biofilm formation and virulence factor (VF) production by P. aeruginosa by hampering the quorum sensing (QS) system(s). Minimum inhibitory concentration (MIC) of PLD was assessed for standard strain and two multi-drug resistant catheter isolates of P. aeruginosa utilizing the microdilution method. Microtiter plate assay, crystal violet staining and scanning electron microscopy were used to evaluate the biofilm inhibition property. CFU was utilized to assess the antifouling property of PLD. Detection of VFs and expression analysis of virulence determinants were applied to investigate the anti-virulence activity. Gene expression and molecular docking studies were also executed to explore the QS inhibition and binding of PLD with QS receptors. In the present study, PLD has significantly inhibited biofilm formation and antivirulence activity at sub-MIC levels (2.5 and 3.5 mM) in all the tested strains. In addition, molecular docking studies revealed a significant affinity towards QS receptors. Perillaldehyde, being a non-toxic food flavouring agent, significantly inhibited biofilm formation and exhibited antifouling property. PLD exhibited significantly reduced levels of VFs (p < 0.001) and their respective genetic determinants (p < 0.001). Gene expression analysis and molecular docking studies confirmed the interactions of PLD to the QS receptors, indicating the plausible mechanism for the anti-virulence property. This study identified the anti-virulence potential of PLD and provided mechanistic insights. PLD can be a suitable, non-toxic candidate for countering biofilms and associated pathogens, contributing to the prevention of biofilm-associated nosocomial infections.

Facile fabrication of thermostable and colorimetric starch-based waterproof coating with edible organic materials

With the attention to pollution and human health, nontoxic food grade superhydrophobic coating as a strategy to reduce food waste has aroused wide interest. Herein, a food grade colorimetric starch-based waterproof coating was achieved using starch nanoparticle (SNP), stearic acid (STA) and anthocyanin. The as-prepared coating exhibited considerable thermostability and remarkable water repellency due to the low-surface-energy pomegranate pulp-like micro/nanostructure assembled by STA and SNP. The certain mechanical and chemical durability, prominent self-cleaning property and liquid food repellency in the coating had been thoroughly verified. Additionally, anthocyanin was creatively introduced to the coating via SNP loading, which endowed the coating with color response to different pH solutions. Thus, the coating is applicable to freshness monitoring without being disabled by water. This thermostable and colorimetric starch-based waterproof coating shows promising application prospects in advanced bio-based food-contact material field.

Organic farming: assessment of the perception of practicing tribal farmers in Jharkhand

Organic farming aims to produce healthy and non-toxic food for consumers using natural inputs to eliminate anysubstance from farming system which is harmful to human as well as soil health. Organic farming is not a new concept but a traditional farming system with modern scientific techniques. Realizing the long-term positive impact of organic farming, Government of India is also promoting the organic farming on a large scale. In line with this, three villages of Angara block of Ranchi District have been developed as fully organic villages. To assess the general, attitudinal and control belief among tribal farmers towards organic farming, a study was conducted in two Participatory Guarantee System (PGS) certified villages namely Dhurleta and Budhakocha of Ranchi district in Jharkhand state during the year 2019–20. Data for the study was collected through personal interview of the tribal farmers practicing organic farming using structured interview schedule developed for the purpose. A total of 50 farmers (25 farmers from each village) selected through systematic random sampling method formed the sample of the study. Findings of the study showed that about 76.80% farmers believed that organic farming is good while 61% of the respondents had favourable attitudinal belief towards organic farming. Majority of the population agreed that disease, pest control, marketing and complexity in certification are the major setbacks in large scale adoption of organic farming.

Determination of tyrosinase-cyanidin-3-O-glucoside and (\u2212/+)-catechin binding modes reveal mechanistic differences in tyrosinase inhibition

Tyrosinase, exquisitely catalyzes the phenolic compounds into brown or black pigment, inhibition is used as a treatment for dermatological or neurodegenerative disorders. Natural products, such as cyanidin-3-O-glucoside and (−/+)-catechin, are considered safe and non-toxic food additives in tyrosinase inhibition but their ambiguous inhibitory mechanism against tyrosinase is still elusive. Thus, we presented the mechanistic insights into tyrosinase with cyanidin-3-O-glucoside and (−/+)-catechin using computational simulations and in vitro assessment. Initial molecular docking results predicted ideal docked poses (− 9.346 to − 5.795 kcal/mol) for tyrosinase with selected flavonoids. Furthermore, 100 ns molecular dynamics simulations and post-simulation analysis of docked poses established their stability and oxidation of flavonoids as substrate by tyrosinase. Particularly, metal chelation via catechol group linked with the free 3-OH group on the unconjugated dihydropyran heterocycle chain was elucidated to contribute to tyrosinase inhibition by (−/+)-catechin against cyanidin-3-O-glucoside. Also, predicted binding free energy using molecular mechanics/generalized Born surface area for each docked pose was consistent with in vitro enzyme inhibition for both mushroom and murine tyrosinases. Conclusively, (−/+)-catechin was observed for substantial tyrosinase inhibition and advocated for further investigation for drug development against tyrosinase-associated diseases.

Bactericidal and fungicidal activities of novel ProtecTeaV formulations - alcohol-based hand hygiene and surface disinfectant prototypes containing epigallocatechin-3-gallate-palmitate (EC16).

Currently used alcohol-based hand sanitizers and surgical hand rubs are not effective against alcohol-resistant microorganisms. We reported previously that nontoxic antioxidant food additive compounds derived from green tea polyphenols, particularly epigallocatechin-3-gallate-palmitate (EC16), are suitable in alcohol formulations to effectively inactivate nonenveloped viruses and bacterial spores. However, whether EC16 influences the bactericidal and fungicidal activity of alcohol is not clear. The objective of the current study was to determine the bactericidal and fungicidal activities of ProtecTeaV hand sanitizer and surface spray prototypes containing EC16. The prototypes were tested according to the guidelines from the Food and Drug Administration (FDA) and the Environmental Protection Agency (EPA). As expected, EC16 did not reduce the bactericidal and fungicidal activities of ethanol. The hand sanitizer gel formulation was equally effective as 70% ethanol and met the tested standard, and the surface spray prototype met the EPA performance standard. EC16 can be combined with ethanol without reducing antibacterial or antifungal activity, and the ProtecTeaV prototypes could be further developed into novel hand hygiene and surface disinfectant products with virucidal, bactericidal, fungicidal and sporicidal activities.

ε-poly-L-lysine Affects the Vegetative Growth, Pathogenicity and Expression Regulation of Necrotrophic Pathogen Sclerotinia sclerotiorum and Botrytis cinerea.

Microbial secondary metabolites produced by Streptomyces are applied to control plant diseases. The metabolite, ε-poly-l-lysine (ε-PL), is a non-toxic food preservative, but the potential application of this compound as a microbial fungicide in agriculture is rarely reported. In this study, the effect and mode of action of ε-PL on two necrotrophic pathogenic fungi, Sclerotinia sclerotiorum and Botrytis cinerea, were investigated. The results showed that ε-PL effectively inhibited the mycelial growth of S. sclerotiorum and B. cinerea with EC50 values of 283 μg/mL and 281 μg/mL, respectively. In addition, ε-PL at the dose of 150 and 300 μg/mL reduced S. sclerotiorum sclerotia formation. The results of the RNA-seq and RT-qPCR validation indicated that ε-PL significantly regulated the gene expression of critical differential expressed genes (DEGs) involved in fungal growth, metabolism, pathogenicity, and induced an increase in the expression of the fungal stress responses and the detoxification genes. These results provided new insights for understanding the modes of action of ε-PL on S. sclerotiorum and B. cinerea and improved the sustainable management of these plant diseases.

Acute Toxicity Study of Porang (Amorphophallus oncophyllus) Flour Macerated with Strobilanthes crispus in Wistar Rats

BACKGROUND: Porang (Amorphophallus oncophyllus) is a local tuber food that high in bioactive compound glucomannan. It uses are limited due to oxalate acid content which poses health risks. Strobilanthes crispus leaves could reduce the level of calcium oxalate in porang. However, there is still no study to prove its safety. AIM: This study aimed to investigate the acute oral toxicity study of porang (A. oncophyllus) macerated with S. crispus based on observation of mortality rate (LD50), the changes in behavior during 72 h, renal and hepatic function such as urinary protein, serum glutamic oxaloacetic transaminase (SGOT), and serum glutamic pyruvic transaminase (SGPT) levels of Wistar rat (Rattus norvegicus) METHODS: An acute toxicity test was conducted based on the Organization of Economic Co-Operation and Development 420 Fixed-Dose Procedure Guideline that consists of preliminary and main studies. For the preliminary study, rats were divided into control and five treatment groups with the dosage of 50, 300, 2000, and 5000 mg/kg body weight (BW) for each natural porang flour (NPF) and S. crispus-macerated porang flour (SPF). For the main study, rats were divided into four groups, those were NPF and SPF with the dosage of 2000 and 5000 mg/kg BW. Levels of urinary protein and blood serum SGOT and SGPT levels were measured at 0, 24, and 72 after treatment. RESULTS: The acute toxicity study showed that porang and porang macerated with S. crispus were not toxic until the highest dose of 5000 mg/kg BW. It was proved by the absence of LD50, no change in behavior, no weight losses, and also the results of biochemical tests, such as urinary protein, SGOT, and SGPT which were still in the normal range. CONCLUSIONS: Porang flour and SPF were concluded as non-toxic food based on acute toxicity study.

Tracing the Fate of Pollen Substitute Patties in Western Honey Bee (Hymenoptera: Apidae) Colonies.

Commercial beekeepers need healthy, productive honey bee (Apis mellifera L.) colonies, even when the landscape lacks adequate pollen forage to sustain the colonies. As a result, many commercial beekeepers spend significant money and labor on the use of pollen substitutes in their colonies. However, there is little consensus in the literature about the benefits and drawbacks of pollen substitute use on honey bee colony health. In order to understand this critically, it is important to know first how honey bees distribute pollen substitute patties throughout their colonies. We traced the fate of three commercially available pollen substitute patties (MegaBee, UltraBee, AP23) dyed with a nontoxic food coloring (Brilliant Blue FCF) and undyed as negative controls, a dyed positive control (fondant), and a dyed consumption control (Crayola Model Magic Clay) in 44 honey bee colonies. Using spectrophotometry and visual inspection, we analyzed adult bee guts, larval guts, bee bread stores and colony debris underneath the hive for presence of the dye. Our data suggest that (1) a proportion of adult bees ingest the patty, (2) adult bees likely do not feed patty directly to larvae, (3) adult bees do not store patty like bee bread, and (4) only a very small proportion of patty is lost as debris. Collectively our data suggest that honey bee colonies use pollen substitute patties. However, patties likely do not replace the function of natural pollen entirely in terms of larval provisioning and long-term storage as bee bread.