• FOCUS: LIFE SCIENCE
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• no.1
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• pp.3.1-3.7
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• 2024

The article discusses the critical role of chromatography in the analysis and purification of proteins in biopharmaceuticals, emphasizing the importance of comprehensive characterization for ensuring their safety and efficacy. It highlights the use of Avantor® ACE® HPLC columns for the separation and purification of proteins, focusing on the analysis of intact proteins using reversed-phase liquid chromatography (RPLC) with fully porous particles. This article also details the application of different mobile phase additives, such as TFA and formic acid, and emphasizes the advantages of using type B ultra-pure silica-based columns for efficiency and peak shape in biomolecule analysis. Additionally, it addresses the challenges of analyzing intact proteins due to slow molecular diffusion and introduces the concept of solid-core (or superficially porous) particles, emphasizing their benefits over traditional porous particles for the analysis of therapeutic proteins. Furthermore, it discusses the development of Avantor® ACE® UltraCore BIO columns, specifically designed for the high-efficiency separation of large biomolecules, such as proteins, and demonstrates their effectiveness in achieving high-resolution separations, even for higher molecular weight proteins like monoclonal antibodies (mAbs). In addition, it underscores the complexity of analyzing and characterizing intact protein biopharmaceuticals, requiring a range of analytical techniques and the use of wide-pore stationary phases, operated at elevated temperatures and with relatively shallow gradients. It highlights the comprehensive range of options offered by Avantor® ACE® wide pore columns, including both fully porous and solid-core particles, bonded with a variety of complementary stationary phase chemistries to optimize selectivity during method development. The use of ultrapure and highly inert base silica is emphasized for enabling the use of lower concentrations of mobile phase modifiers without compromising analyte peak shape, particularly beneficial for LC-MS applications. Then the article concludes by emphasizing the significance of reversed-phase liquid chromatography and its compatibility with mass spectrometry as a valuable tool for the separation and analysis of intact proteins and their closely related variants in biopharmaceuticals.

• Asian-Australasian Journal of Animal Sciences
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• v.33 no.1
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• pp.127-131
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• 2020

Objective: This study was conducted to determine catalytic properties of wheat phytase with exopolyphosphatase activity toward medium-chain and long-chain inorganic polyphosphate (polyP) substrates for comparative purpose. Methods: Exopolyphosphatase assay of wheat phytase toward polyP75 (medium-chain polyP with average 75 phosphate residues) and polyP1150 (long-chain polyP with average 1150 phosphate residues) was performed at pH 5.2 and pH 7.5. Its activity toward these substrates was investigated in the presence of Mg²⁺, Ni²⁺, Co²⁺, Mn²⁺, or ethylenediaminetetraacetic acid (EDTA). Michaelis constant (K_m) and maximum reaction velocity (V_max) were determined from Lineweaver-Burk plot with polyP75 or polyP1150. Monophosphate esterase activity toward p-nitrophenyl phosphate (pNPP) was assayed in the presence of polyP75 or polyP1150. Results: Wheat phytase dephosphorylated polyP75 and polyP1150 at pH 7.5 more effectively than that at pH 5.2. Its exopolyphosphatase activity toward polyP75 at pH 5.2 was 1.4-fold higher than that toward polyP1150 whereas its activity toward polyP75 at pH 7.5 was 1.4-fold lower than that toward polyP1150. Regarding enzyme kinetics, K_m for polyP75 was 1.4-fold lower than that for polyP1150 while V_max for polyP1150 was 2-fold higher than that for polyP75. The presence of Mg²⁺, Ni²⁺, Co²⁺, Mn²⁺, or EDTA (1 or 5 mM) exhibited no inhibitory effect on its activity toward polyP75. Its activity toward polyP1150 was inhibited by 1 mM of Ni²⁺ or Co²⁺ and 5 mM of Ni²⁺, Co²⁺, or Mg²⁺. Ni²⁺ inhibited its activity toward polyP1150 the most strongly among tested additives. Both polyP75 and polyP1150 inhibited the monophosphate esterase activity of wheat phytase toward pNPP in a dose-dependent manner. Conclusion: Wheat phytase with an unexpected exopolyphosphatase activity has potential as a therapeutic tool and a next-generational feed additive for controlling long-chain polyP-induced inappropriate inflammation from Campylobacter jejuni and Salmonella typhimurium infection in public health and animal husbandry.

• Korean Journal of Clinical Laboratory Science
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• v.48 no.2
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• pp.118-123
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• 2016

Antimicrobial agents have been used in poultry for treatment of bacterial infections or additives over the past half century. However, increasing antimicrobial resistance has led to selective pressure for therapeutic use in humans and made treatment of bacterial infection more difficult. In this study, we examined the prevalence of plasmid mediated antimicrobial resistant determinants for resistance to ${\beta}-lactam$, quinolone, and aminoglycoside in Enterobacteriaceae isolates obtained from chickens in Gyeongsang provinces, and correlation between the resistant genes and antimicrobial resistance rate was also assessed. A total of 43 Enterobacteriaceae isolates were recovered from 40 chickens at Gyeongsang provinces in Korea. Antimicrobial susceptibility was determined by disk diffusion method. PCR and DNA sequencing were performed to characterize the antimicrobial resistant genes. Of the 43 Enterobacteriaceae isolates tested, 2 isolates harbored $bla_{CTX-M-14}$ gene, and 2 and 5 strains contained qnrS and aac(6')-Ib-cr genes, respectively. A total of 43 isolates displayed a relatively lower susceptible rate ranging between 0.0 and 23.3% to most of the antimicrobial agents, except cefepime, ceftazidime, and cefaclor. We confirmed that plasmid mediated antimicrobial resistant determinants were distributed in Enterobacteriaceae isolates from chickens. Investigation of the genes and monitoring of antimicrobial resistance rate is required to prevent further spreading of antimicrobial resistant genes among Enterobacteriaceae isolates.

• Clinical and Experimental Pediatrics
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• v.51 no.3
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• pp.307-314
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• 2008

Purpose : Capsaicin, the major pungent ingredient in red pepper, has long been used in spices and food additives. It has been recently shown to induce apoptosis in several cell lines through a not well known mechanism. The aim of this study was to investigate the apoptosis-inducing effect of capsaicin on gastric cancer cells, and to provide valuable information concerning the application of capsaicin for therapeutic purposes. Methods : Cultured SNU-668 cells were treated with capsaicin. We analyzed cell survival by trypan blue and crystal violet analysis, cell cytotoxicity by MTT assay, apoptosis by nuclear condensation and DNA fragmentation, bcl-2 and bax mRNA expression by RT-PCR, and the expression of apoptosis related proteins by Western immunoblot analysis. In order to assess whether the growth inhibitory effect of anticancer drugs is enhanced by capsaicin, we investigated the effects of cell cytotoxicity and the expression of apoptosis related proteins of etoposide and adriamycin treated with capsaicin in cells. Results : Capsaicin inhibited growth of SNU-668 cells in a dose-dependent manner. This inhibitory effect of capsaicin on cell growth was mainly due to the induction of apoptosis as evidenced by DNA fragmentation, nuclear condensation and the expression of apoptosis related proteins. Furthermore, capsaicin prominently reduced the ratio of anti-apoptotic Bcl-2 to pro-apoptotic Bax and consequently increased caspase-3 activity. The cells treated with capsaicin were more sensitive to death induced by etoposide and adriamycin than the cells without capsaicin. Conclusion : These results demonstrate that capsaicin efficiently induced apoptosis in SNU-668 cells through a caspase-3-dependent mechanism and sensitizes cancer cells to anticancer drugs toward apoptotic cell death, which may contribute to its anticancer effect and chemosensitizer function against gastric cancer.

• The Journal of Korean Medicine
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• v.21 no.3
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• pp.149-165
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• 2000

As recent developments of Immunology and Nuclear medicine, serum IgE and IgG values are helpful in the diagnosis and evaluation of the therapeutic effects of nasal allergies. But in Korea, air pollution and the increased use of food additives have become leading factors in nasal allergies, It seems to be induced by environmental change, especially industrialization and urbanization, so allergic rhinitis in our environment has changed in accordance with the changes made in the living environment. Therefore this study is attempted in order to observe a clinical analysis which places more importance on allergic rhinitis. We studied 200 patients who had visited Kyunghee Oriental Medical Center with allergic rhinitis from January 1, 1999 to December 31, 1999 The results were as follows: 1. The sex distribution was 114 males(57%) and 86 females(43%). In age distribution, the average age was 25. In males, ages ranged from 3 to 66 years old and the average was 23.81. In females, ages ranged from 4 to 67 years old and the average was 28.57. The peak age was 30～39 years old(24%); under 9 years old and 10～19 years old were each 18%; 20～29 years old was 22%; 40～49 years old was 11 %; over 50 years old 6.5%. The gulf between males and females showed a statistically significant difference(P<0.025). 2. In the age of onset, male' s maximum was 62.5, minimum was 0.25; female s maximum was 59.5, minimum was 0.2. Under 9 years old was the most with 34%(male 24%, female 10%), 10～19 years old was 18%, 20～29 years old was 22.5%, 30～39 years old was 13.50%, over 40 years old was 12%, The gulf between males and females were showed statistically significant difference.(p<0.014) 3. The average duration of the disease was 5.67 years. In male and female, the maximum was 30, the minimum was 0.05; under 5 years old was the most with 62%(male 34.50%, female 27.50%); 6～10 years old was 23%. So, under 10 years old was 85%. There was no statistically significant difference in the duration of disease. 4. Regarding type of residence, 47.50% of patients with allergic rhinitis lived in apartments, 52.50% lived in houses. In males, 29.50% lived in apartments, 27.50% lived in houses. In females, 18% lived in apartments, 25% lived in houses. There was no statistically significant difference in the residence by T-test and chi-test. 5. In the distribution of season, spring is the most with 29.5% of patients, winter 28%, fall 25.5%, and summer 17%. But there was no statistically significant difference. 6. After observing 200 patients with allergic rhinitis, classifying main symptoms into 5 types, sneezing was the main symptom in 177cases(88.50%), nasal obstruction in 176cases(88%), rhinorrhea in 169cases(84.5%), post nasal discharge in 87cases(43.50%), and itching in I04cases(52%). The Cumulus ration is 98.50% and symptoms overlapped with an average 3.57±0.1 times but in an analysis of variance of these symptoms, the gulf between males and females was not recognized as statistically significant by T-test and ANOVA. 7. Patients whose families have allergic diseases account for 90 cases(45%) : 49cases(24.50%) male and 41cases(20.50%) female. There were 4 cases (71.11 %) whose families have allergic rhinitis, 9cases(10%) of asthma, and 7.78% with allergic dermatitis. There were 61 (67.80%) cases of patients whose parents have allergic diseases; cases wherein the patient s child had allergic diseases numbered 13 (14.45%); and cases with a sibling with allergic diseases totalled 16cases (17.80%). There was no statistically significant difference in allergic disease regarding sex, parents, or siblings by chi-test. 8. Blood type: For males, type A is the most common, with 37cases(18.5%), followed by type B with 32cases(16%), type O 28cases(l4%) and type AB 13cases(6.5%). For females, type B is the most common, with 30cases(15%), followed by type O with 23cases(l1.5%), type A with 18cases(9%) and type AB with 13cases(6.5%). There was no statistically significant difference in blood type by chi-test. 9. In the selection of prefered food, most patients prefer cool food, with 98 such cases(49%), tepid food in 54cases(27%) and warm food in 48cases(24%). These showed a statistically significant difference in the selection of prefered food between males and females by chi-test(p<0.009). 10. The state of Past History was classified into II types. chronic hypertrophic rhinitis is the most common with 11cases (18.64%), tonsil and adenoid hypertrophy is 8cases(l3.56%), sinusitis is 6cases(10.17%), nasal septum deviation is 4cases, nasal polyp is 2cases, others are 10cases(l6.95%). No statistically significant difference in past history between males and females was shown, but a statistically significant difference was shown when males and females were compared with total cases by T-test(p<0.002, P<0.0008). 11. Regarding complications, 37 patients (28.91%) had sinusitis: 22cases(17.19%) in male, 15cases(11.72%) in female. Chronic hypertrophic rhinitis was found in 15cases(11.72%). Others are under 10%. There was no statistically significant difference in the type of complications between males and females, but a statistically significant difference was shown when males and females were compared with total cases by T-test(P<0.00l, P<0.007). 12. In the treatment, medication was used 1691 times, an average of 2.58 times. No.34 was used 370 times for 124 cases, an average of 2.98 times. No. 152 was used 318 times for 106 cases, an average of 3.00 times. No.151 was used 307 times for 97cases, an average of 3.16 times. No. 31 was used 117 times for 33 cases, an average of 3.55 times. No 25 was used 116 times for 33 cases, an average of 3.52 times. 13. In the duration of treatment, the most frequent is 1 week(69cases, 34.50%), the maximum is 20weeks, and the minimum is 1week. A treatment period of 2～3 weeks accounted for 32% of cases, a period of 4～5weeks accounted for 13.5%. The gulf between males and females showed a statistically significant difference in the duration of treatment.(p<0.01). There was a statistical significance when the males were compared with total cases by ANOVA(P<0.03). 14. A comparison between before-treatment and after-treatment showed a statistically significant difference in treatment by T-test (p<0.01) and F-test (p<0.0058).