This research applied and evaluated a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) using cytochrome b gene to detect pork contamination in meatballs from local markets in Surabaya and Yogyakarta regions, Indonesia. To confirm the effectiveness and specificity of this fragment, thirty nine DNA samples from different meatball shops were isolated and amplified, and then the PCR amplicon was digested by BseDI restriction enzyme to detect the presence of pork in meatballs. BseDI restriction enzyme was able to cleave porcine cytochrome b gene into two fragments (131 bp and 228 bp). Testing the meatballs from the local market showed that nine of twenty meatball shops in Yogyakarta region were detected to have pork contamination, but there was no pork contamination in meatball shops in Surabaya region. In conclusion, specific PCR amplification of cytochrome b gen and cleaved by BseDI restriction enzymes seems to be a powerful technique for the identification of pork presence in meatball because of its simplicity, specificity and sensitivity. Furthermore, pork contamination intended for commercial products of sausage, nugget, steak and meat burger can be checked. The procedure is also much cheaper than other methods based on PCR, immunodiffusion and other techniques that need expensive equipment.

A polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) using BseDI restriction enzyme had been applied for identifying the presence of pork in processed meat (beef sausage and chicken nugget) including before and after frying. Pork sample in various levels (1%, 3%, 5%, 10%, and 25 %) was prepared in a mixture with beef and chicken meats and processed for sausage and nugget. The primers CYTb1 and CYTb2 were designed in the mitochondrial cytochrome b (cyt b) gene and PCR successfully amplifi ed fragments of 359 bp. To distinguish existence of porcine species, the amplifi ed PCR products of mitochondrial DNA were cut by BseDI restriction enzyme. The result showed pig mitochondrial DNA was cut into 131 and 228 bp fragments. The PCR-RFLP species identifi cation assay yielded excellent results for identifi cation of porcine species. It is a potentially reliable technique for pork detection in animal food processed products for Halal authentication.

Access to vitamin B [(R)-pantothenic acid] and both diastereoisomers of α-methyl-substituted vitamin B [(R)- and (S)-3-((R)-2,4-dihydroxy-3,3-dimethylbutanamido)-2-methylpropanoic acid] was achieved using a modular three-step biocatalytic cascade involving 3-methylaspartate ammonia lyase (MAL), aspartate-α-decarboxylase (ADC), β-methylaspartate-α-decarboxylase (CrpG) or glutamate decarboxylase (GAD), and pantothenate synthetase (PS) enzymes. Starting from simple non-chiral dicarboxylic acids (either fumaric acid or mesaconic acid), vitamin B and both diastereoisomers of α-methyl-substituted vitamin B , which are valuable precursors for promising antimicrobials against Plasmodium falciparum and multidrug-resistant Staphylococcus aureus, can be generated in good yields (up to 70 %) and excellent enantiopurity (>99 % ee). This newly developed cascade process may be tailored and used for the biocatalytic production of various vitamin B derivatives by modifying the pantoyl or β-alanine moiety.

The study was designed to determine the potential of collagen hydrolysate produced from the skin of Kacang goat through chymotrypsin hydrolysis to be used as an inhibitor of angiotensin converting enzyme (ACE). This research was conducted in three replications, with the measured parameters include ACE inhibitory potential and collagen hydrolysate fractionation. The results showed that collagen extraction of Kacang goat skin by chymotrypsin hydrolysis yielded 9.74% (dry matter, v/v) collagen, with pH at 6.6. The extracted collagen contained α1, α2, and β collagen chains with molecular weights of 151 kDa, 141 kDa, and 240 kDa, respectively. Furthermore, the collagen hydrolysis produced protein peptides confirmed at molecular weights of 43 to 107 kDa. The hydrolysate fractionation at molecular weights of <3 kDa, 3-5 kDa, and >5 kDa showed proteins concentrations of 2.33 mg/mL, 3.81 mg/mL, and 3.93 mg/mL, respectively. The hydrolysate fractionation with molecular weight <3 kDa showed to have ACE inhibition activity with the IC50 value of 0.47 mg/mL. The study concluded that collagen hydrolysate extracted from the skin of Kacang goat had a promising potential as a source of antihypertensive agent.

Background and Aim: Angiotensin-converting enzyme (ACE) is one of the inhibitory enzymes isolated from animals for the treatment of hypertension. ACE inhibitor (ACE-I) peptides can be obtained by hydrolyzing proteins from various animal tissues, including muscle and connective tissues. However, the study on ACE-I activity from collagen of Kacang goat skin has not been conducted. This study explores the potency of collagen from Kacang goat skin as a source of an antihypertensive agent through ACE inhibition. Thermolysin will hydrolyze collagen and produce the peptide classified antihypertensive bioactive peptides. This study aimed to determine the potential of thermolysin to hydrolyze collagen of Kacang goat skin for ACE-I peptide production and to identify the production of ACE-I peptides. Materials and Methods: Collagen from Kacang goat skin was hydrolyzed with thermolysin and incubated at 37°C for 1 h. Molecular weight (MW) evaluation was performed by SDS PAGE; fractionation peptides at <5 kDa, 3-5 kDa, and <3 kDa were performed by ultrafiltration and ACE-I activity determined by IC50 measurement. Results: Collagen was hydrolyzed by thermolysin, resulting in protein with MW of 117.50-14.60 kDa. The protein content of fractionation at >5 kDa was 3.93±0.72 mg/mL, content of 3-5 kDa was 3.81±0.68 mg/mL, and that of <3 kDa was 2.33±0.38 mg/mL. Fractionation was performed 3 times and one of the results was selected for the ACE-I test. The selected fraction was tested by IC50 measurement with three repetitions and it showed an average enzyme activity at 0.83 μg/mL or 82.94 mg/mL. Conclusion: Thermolysin hydrolysis of collagen from Kacang goat skin showed the potential to produce bioactive peptides, such as ACE-I.